@article{pmid39318036,

title = {PSAP-Genomic-Regions: A Method Leveraging Population Data to Prioritize Coding and Non-Coding Variants in Whole Genome Sequencing for Rare Disease Diagnosis},

author = {Marie-Sophie C Ogloblinsky and Ozvan Bocher and Chaker Aloui and Anne-Louise Leutenegger and Ozan Ozisik and Anaïs Baudot and Elisabeth Tournier-Lasserve and Helen Castillo-Madeen and Daniel Lewinsohn and Donald F Conrad and Emmanuelle Génin and Gaëlle Marenne},

doi = {10.1002/gepi.22593},

issn = {1098-2272},

year  = {2024},

date = {2024-09-01},

urldate = {2024-09-01},

journal = {Genet Epidemiol},

abstract = {The introduction of Next-Generation Sequencing technologies in the clinics has improved rare disease diagnosis. Nonetheless, for very heterogeneous or very rare diseases, more than half of cases still lack molecular diagnosis. Novel strategies are needed to prioritize variants within a single individual. The Population Sampling Probability (PSAP) method was developed to meet this aim but only for coding variants in exome data. Here, we propose an extension of the PSAP method to the non-coding genome called PSAP-genomic-regions. In this extension, instead of considering genes as testing units (PSAP-genes strategy), we use genomic regions defined over the whole genome that pinpoint potential functional constraints. We conceived an evaluation protocol for our method using artificially generated disease exomes and genomes, by inserting coding and non-coding pathogenic ClinVar variants in large data sets of exomes and genomes from the general population. PSAP-genomic-regions significantly improves the ranking of these variants compared to using a pathogenicity score alone. Using PSAP-genomic-regions, more than 50% of non-coding ClinVar variants were among the top 10 variants of the genome. On real sequencing data from six patients with Cerebral Small Vessel Disease and nine patients with male infertility, all causal variants were ranked in the top 100 variants with PSAP-genomic-regions. By revisiting the testing units used in the PSAP method to include non-coding variants, we have developed PSAP-genomic-regions, an efficient whole-genome prioritization tool which offers promising results for the diagnosis of unresolved rare diseases.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid39122675,

title = {Inherited defects of piRNA biogenesis cause transposon de-repression, impaired spermatogenesis, and human male infertility},

author = {Birgit Stallmeyer and Clara Bühlmann and Rytis Stakaitis and Ann-Kristin Dicke and Farah Ghieh and Luisa Meier and Ansgar Zoch and David MacKenzie MacLeod and Johanna Steingröver and Özlem Okutman and Daniela Fietz and Adrian Pilatz and Antoni Riera-Escamilla and Miguel J Xavier and Christian Ruckert and Sara Di Persio and Nina Neuhaus and Ali Sami Gurbuz and Ahmet Şalvarci and Nicolas Le May and Kevin McEleny and Corinna Friedrich and Godfried van der Heijden and Margot J Wyrwoll and Sabine Kliesch and Joris A Veltman and Csilla Krausz and Stéphane Viville and Donald F Conrad and Dónal O'Carroll and Frank Tüttelmann},

doi = {10.1038/s41467-024-50930-9},

issn = {2041-1723},

year  = {2024},

date = {2024-08-01},

urldate = {2024-08-01},

journal = {Nat Commun},

volume = {15},

number = {1},

pages = {6637},

abstract = {piRNAs are crucial for transposon silencing, germ cell maturation, and fertility in male mice. Here, we report on the genetic landscape of piRNA dysfunction in humans and present 39 infertile men carrying biallelic variants in 14 different piRNA pathway genes, including PIWIL1, GTSF1, GPAT2, MAEL, TDRD1, and DDX4. In some affected men, the testicular phenotypes differ from those of the respective knockout mice and range from complete germ cell loss to the production of a few morphologically abnormal sperm. A reduced number of pachytene piRNAs was detected in the testicular tissue of variant carriers, demonstrating impaired piRNA biogenesis. Furthermore, LINE1 expression in spermatogonia links impaired piRNA biogenesis to transposon de-silencing and serves to classify variants as functionally relevant. These results establish the disrupted piRNA pathway as a major cause of human spermatogenic failure and provide insights into transposon silencing in human male germ cells.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38997255,

title = {AXDND1 is required to balance spermatogonial commitment and for sperm tail formation in mice and humans},

author = {Brendan J Houston and Joseph Nguyen and D Jo Merriner and Anne E O'Connor and Alexandra M Lopes and Liina Nagirnaja and Corinna Friedrich and Sabine Kliesch and Frank Tüttelmann and Kenneth I Aston and Donald F Conrad and Robin M Hobbs and Jessica E M Dunleavy and Moira K O'Bryan},

doi = {10.1038/s41419-024-06874-5},

issn = {2041-4889},

year  = {2024},

date = {2024-07-01},

urldate = {2024-07-01},

journal = {Cell Death Dis},

volume = {15},

number = {7},

pages = {499},

abstract = {Dynein complexes are large, multi-unit assemblies involved in many biological processes via their critical roles in protein transport and axoneme motility. Using next-generation sequencing of infertile men presenting with low or no sperm in their ejaculates, we identified damaging variants in the dynein-related gene AXDND1. We thus hypothesised that AXDND1 is a critical regulator of male fertility. To test this hypothesis, we produced a knockout mouse model. Axdnd1 males were sterile at all ages but presented with an evolving testis phenotype wherein they could undergo one round of histologically replete spermatogenesis followed by a rapid depletion of the seminiferous epithelium. Marker experiments identified a role for AXDND1 in maintaining the balance between differentiation-committed and self-renewing spermatogonial populations, resulting in disproportionate production of differentiating cells in the absence of AXDND1 and increased sperm production during initial spermatogenic waves. Moreover, long-term spermatogonial maintenance in the Axdnd1 knockout was compromised, ultimately leading to catastrophic germ cell loss, destruction of blood-testis barrier integrity and immune cell infiltration. In addition, sperm produced during the first wave of spermatogenesis were immotile due to abnormal axoneme structure, including the presence of ectopic vesicles and abnormalities in outer dense fibres and microtubule doublet structures. Sperm output was additionally compromised by a severe spermiation defect and abnormal sperm individualisation. Collectively these data identify AXDND1 as an atypical dynein complex-related protein with a role in protein/vesicle transport of relevance to spermatogonial function and sperm tail formation in mice and humans. This study underscores the importance of studying the consequences of gene loss-of-function on both the establishment and maintenance of male fertility.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38570187,

title = {Genetic mutation of  results in male infertility due to abnormal sperm tail composition},

author = {Brendan J Houston and D Jo Merriner and G Gemma Stathatos and Joseph H Nguyen and Anne E O'Connor and Alexandra M Lopes and Donald F Conrad and Mark Baker and Jessica Em Dunleavy and Moira K O'Bryan},

doi = {10.26508/lsa.202302452},

issn = {2575-1077},

year  = {2024},

date = {2024-06-01},

urldate = {2024-06-01},

journal = {Life Sci Alliance},

volume = {7},

number = {6},

abstract = {The transition zone is a specialised gate at the base of cilia/flagella, which separates the ciliary compartment from the cytoplasm and strictly regulates protein entry. We identified a potential new regulator of the male germ cell transition zone, CEP76. We demonstrated that CEP76 was involved in the selective entry and incorporation of key proteins required for sperm function and fertility into the ciliary compartment and ultimately the sperm tail. In the mutant, sperm tails were shorter and immotile as a consequence of deficits in essential sperm motility proteins including DNAH2 and AKAP4, which accumulated at the sperm neck in the mutant. Severe annulus, fibrous sheath, and outer dense fibre abnormalities were also detected in sperm lacking CEP76. Finally, we identified that CEP76 dictates annulus positioning and structure. This study suggests CEP76 as a male germ cell transition zone protein and adds further evidence to the hypothesis that the spermatid transition zone and annulus are part of the same functional structure.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38614076,

title = {Toward clinical exomes in diagnostics and management of male infertility},

author = {Kristiina Lillepea and Anna-Grete Juchnewitsch and Laura Kasak and Anu Valkna and Avirup Dutta and Kristjan Pomm and Olev Poolamets and Liina Nagirnaja and Erik Tamp and Eisa Mahyari and Vladimir Vihljajev and Stanislav Tjagur and Sofia Papadimitriou and Antoni Riera-Escamilla and Nassim Versbraegen and Ginevra Farnetani and Helen Castillo-Madeen and Mailis Sütt and Viljo Kübarsepp and Sven Tennisberg and Paul Korrovits and Csilla Krausz and Kenneth I Aston and Tom Lenaerts and Donald F Conrad and Margus Punab and Maris Laan},

doi = {10.1016/j.ajhg.2024.03.013},

issn = {1537-6605},

year  = {2024},

date = {2024-05-01},

journal = {Am J Hum Genet},

volume = {111},

number = {5},

pages = {877--895},

abstract = {Infertility, affecting ∼10% of men, is predominantly caused by primary spermatogenic failure (SPGF). We screened likely pathogenic and pathogenic (LP/P) variants in 638 candidate genes for male infertility in 521 individuals presenting idiopathic SPGF and 323 normozoospermic men in the ESTAND cohort. Molecular diagnosis was reached for 64 men with SPGF (12%), with findings in 39 genes (6%). The yield did not differ significantly between the subgroups with azoospermia (20/185, 11%), oligozoospermia (18/181, 10%), and primary cryptorchidism with SPGF (26/155, 17%). Notably, 19 of 64 LP/P variants (30%) identified in 28 subjects represented recurrent findings in this study and/or with other male infertility cohorts. NR5A1 was the most frequently affected gene, with seven LP/P variants in six SPGF-affected men and two normozoospermic men. The link to SPGF was validated for recently proposed candidate genes ACTRT1, ASZ1, GLUD2, GREB1L, LEO1, RBM5, ROS1, and TGIF2LY. Heterozygous truncating variants in BNC1, reported in female infertility, emerged as plausible causes of severe oligozoospermia. Data suggested that several infertile men may present congenital conditions with less pronounced or pleiotropic phenotypes affecting the development and function of the reproductive system. Genes regulating the hypothalamic-pituitary-gonadal axis were affected in >30% of subjects with LP/P variants. Six individuals had more than one LP/P variant, including five with two findings from the gene panel. A 4-fold increased prevalence of cancer was observed in men with genetic infertility compared to the general male population (8% vs. 2%; p = 4.4 × 10). Expanding genetic testing in andrology will contribute to the multidisciplinary management of SPGF.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38577799,

title = {The human infertility single-cell testis atlas (HISTA): an interactive molecular scRNA-Seq reference of the human testis},

author = {Eisa Mahyari and Katinka A Vigh-Conrad and Clément Daube and Ana C Lima and Jingtao Guo and Douglas T Carrell and James M Hotaling and Kenneth I Aston and Donald F Conrad},

doi = {10.1111/andr.13637},

issn = {2047-2927},

year  = {2024},

date = {2024-04-01},

urldate = {2024-04-01},

journal = {Andrology},

abstract = {BACKGROUND: Single-cell RNA-seq (scRNA-Seq) has been widely adopted to study gene expression of the human testis. Several datasets of scRNA-Seq from human testis have been generated from different groups processed with different informatics pipelines. An integrated atlas of scRNA-Seq expression constructed from multiple donors, developmental ages, and fertility states would be widely useful for the testis research community.nnOBJECTIVE: To describe the generation and use of the human infertility single-cell testis atlas (HISTA), an interactive web tool for understanding human spermatogenesis through scRNA-Seq analysis.nnMETHODS: We obtained scRNA-Seq datasets derived from 12 donors, including healthy adult controls, juveniles, and several infertility cases, and reprocessed these data using methods to remove batch effects. Using Shiny, an open-source environment for data visualization, we created numerous interactive tools for exploring the data, some of which support simple statistical hypothesis testing. We used the resulting HISTA browser and its underlying data to demonstrate HISTA's value for testis researchers.nnRESULTS: A primary application of HISTA is to search by a single gene or a set of genes; thus, we present various analyses that quantify and visualize gene expression across the testis cells and pathology. HISTA also contains machine-learning-derived gene modules ("components") that capture the entire transcriptional landscape of the testis tissue. We show how the use of these components can simplify the highly complex data in HISTA and assist with the interpretation of genes with unknown functions. Finally, we demonstrate the diverse ways HISTA can be used for new data analysis, including hypothesis testing.nnDISCUSSION AND CONCLUSIONS: HISTA is a research environment that can help scientists organize and understand the high-dimensional transcriptional landscape of the human testis. HISTA has already contributed to published testis research and can be updated as needed with input from the research community or downloaded and modified for individual needs.},

keywords = {Human Reproduction, Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38297832,

title = {Heterozygous loss-of-function SMC3 variants are associated with variable growth and developmental features},

author = {Morad Ansari and Kamli N W Faour and Akiko Shimamura and Graeme Grimes and Emeline M Kao and Erica R Denhoff and Ana Blatnik and Daniel Ben-Isvy and Lily Wang and Benjamin M Helm and Helen Firth and Amy M Breman and Emilia K Bijlsma and Aiko Iwata-Otsubo and Thomy J L de Ravel and Vincent Fusaro and Alan Fryer and Keith Nykamp and Lara G Stühn and Tobias B Haack and G Christoph Korenke and Panayiotis Constantinou and Kinga M Bujakowska and Karen J Low and Emily Place and Jennifer Humberson and Melanie P Napier and Jessica Hoffman and Jane Juusola and Matthew A Deardorff and Wanqing Shao and Shira Rockowitz and Ian Krantz and Maninder Kaur and Sarah Raible and Victoria Dortenzio and Sabine Kliesch and Moriel Singer-Berk and Emily Groopman and Stephanie DiTroia and Sonia Ballal and Siddharth Srivastava and Kathrin Rothfelder and Saskia Biskup and Jessica Rzasa and Jennifer Kerkhof and Haley McConkey and Bekim Sadikovic and Sarah Hilton and Siddharth Banka and Frank Tüttelmann and Donald F Conrad and Anne O'Donnell-Luria and Michael E Talkowski and David R FitzPatrick and Philip M Boone},

doi = {10.1016/j.xhgg.2024.100273},

issn = {2666-2477},

year  = {2024},

date = {2024-04-01},

journal = {HGG Adv},

volume = {5},

number = {2},

pages = {100273},

abstract = {Heterozygous missense variants and in-frame indels in SMC3 are a cause of Cornelia de Lange syndrome (CdLS), marked by intellectual disability, growth deficiency, and dysmorphism, via an apparent dominant-negative mechanism. However, the spectrum of manifestations associated with SMC3 loss-of-function variants has not been reported, leading to hypotheses of alternative phenotypes or even developmental lethality. We used matchmaking servers, patient registries, and other resources to identify individuals with heterozygous, predicted loss-of-function (pLoF) variants in SMC3, and analyzed population databases to characterize mutational intolerance in this gene. Here, we show that SMC3 behaves as an archetypal haploinsufficient gene: it is highly constrained against pLoF variants, strongly depleted for missense variants, and pLoF variants are associated with a range of developmental phenotypes. Among 14 individuals with SMC3 pLoF variants, phenotypes were variable but coalesced on low growth parameters, developmental delay/intellectual disability, and dysmorphism, reminiscent of atypical CdLS. Comparisons to individuals with SMC3 missense/in-frame indel variants demonstrated an overall milder presentation in pLoF carriers. Furthermore, several individuals harboring pLoF variants in SMC3 were nonpenetrant for growth, developmental, and/or dysmorphic features, and some had alternative symptomatologies with rational biological links to SMC3. Analyses of tumor and model system transcriptomic data and epigenetic data in a subset of cases suggest that SMC3 pLoF variants reduce SMC3 expression but do not strongly support clustering with functional genomic signatures of typical CdLS. Our finding of substantial population-scale LoF intolerance in concert with variable growth and developmental features in subjects with SMC3 pLoF variants expands the scope of cohesinopathies, informs on their allelic architecture, and suggests the existence of additional clearly LoF-constrained genes whose disease links will be confirmed only by multilayered genomic data paired with careful phenotyping.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38359823,

title = {C19ORF84 connects piRNA and DNA methylation machineries to defend the mammalian germ line},

author = {Ansgar Zoch and Gabriela Konieczny and Tania Auchynnikava and Birgit Stallmeyer and Nadja Rotte and Madeleine Heep and Rebecca V Berrens and Martina Schito and Yuka Kabayama and Theresa Schöpp and Sabine Kliesch and Brendan Houston and Liina Nagirnaja and Moira K O'Bryan and Kenneth I Aston and Donald F Conrad and Juri Rappsilber and Robin C Allshire and Atlanta G Cook and Frank Tüttelmann and Dónal O'Carroll},

doi = {10.1016/j.molcel.2024.01.014},

issn = {1097-4164},

year  = {2024},

date = {2024-03-01},

journal = {Mol Cell},

volume = {84},

number = {6},

pages = {1021--1035.e11},

abstract = {In the male mouse germ line, PIWI-interacting RNAs (piRNAs), bound by the PIWI protein MIWI2 (PIWIL4), guide DNA methylation of young active transposons through SPOCD1. However, the underlying mechanisms of SPOCD1-mediated piRNA-directed transposon methylation and whether this pathway functions to protect the human germ line remain unknown. We identified loss-of-function variants in human SPOCD1 that cause defective transposon silencing and male infertility. Through the analysis of these pathogenic alleles, we discovered that the uncharacterized protein C19ORF84 interacts with SPOCD1. DNMT3C, the DNA methyltransferase responsible for transposon methylation, associates with SPOCD1 and C19ORF84 in fetal gonocytes. Furthermore, C19ORF84 is essential for piRNA-directed DNA methylation and male mouse fertility. Finally, C19ORF84 mediates the in vivo association of SPOCD1 with the de novo methylation machinery. In summary, we have discovered a conserved role for the human piRNA pathway in transposon silencing and C19ORF84, an uncharacterized protein essential for orchestrating piRNA-directed DNA methylation.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37723288,

title = {Improved phenotypic classification of male infertility to promote discovery of genetic causes},

author = {Margot J Wyrwoll and Godfried W van der Heijden and Csilla Krausz and Kenneth I Aston and Sabine Kliesch and Robert McLachlan and Liliana Ramos and Donald F Conrad and Moira K O'Bryan and Joris A Veltman and Frank Tüttelmann},

doi = {10.1038/s41585-023-00816-0},

issn = {1759-4820},

year  = {2024},

date = {2024-02-01},

journal = {Nat Rev Urol},

volume = {21},

number = {2},

pages = {91--101},

abstract = {An increasing number of genes are being described in the context of non-syndromic male infertility. Linking the underlying genetic causes of non-syndromic male infertility with clinical data from patients is important to establish new genotype-phenotype correlations. This process can be facilitated by using universal nomenclature, but no standardized vocabulary is available in the field of non-syndromic male infertility. The International Male Infertility Genomics Consortium aimed at filling this gap, providing a standardized vocabulary containing nomenclature based on the Human Phenotype Ontology (HPO). The "HPO tree" was substantially revised compared with the previous version and is based on the clinical work-up of infertile men, including physical examination and hormonal assessment. Some causes of male infertility can already be suspected based on the patient's clinical history, whereas in other instances, a testicular biopsy is needed for diagnosis. We assembled 49 HPO terms that are linked in a logical hierarchy and showed examples of morphological features of spermatozoa and testicular histology of infertile men with identified genetic diagnoses to describe the phenotypes. This work will help to record patients' phenotypes systematically and facilitate communication between geneticists and andrologists. Collaboration across institutions will improve the identification of patients with the same phenotypes, which will promote the discovery of novel genetic causes for non-syndromic male infertility.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38654924,

title = {Undiagnosed RASopathies in infertile men},

author = {Anna-Grete Juchnewitsch and Kristjan Pomm and Avirup Dutta and Erik Tamp and Anu Valkna and Kristiina Lillepea and Eisa Mahyari and Stanislav Tjagur and Galina Belova and Viljo Kübarsepp and Helen Castillo-Madeen and Antoni Riera-Escamilla and Lisanna Põlluaas and Liina Nagirnaja and Olev Poolamets and Vladimir Vihljajev and Mailis Sütt and Nassim Versbraegen and Sofia Papadimitriou and Robert I McLachlan and Keith A Jarvi and Peter N Schlegel and Sven Tennisberg and Paul Korrovits and Katinka Vigh-Conrad and Moira K O'Bryan and Kenneth I Aston and Tom Lenaerts and Donald F Conrad and Laura Kasak and Margus Punab and Maris Laan},

doi = {10.3389/fendo.2024.1312357},

issn = {1664-2392},

year  = {2024},

date = {2024-01-01},

journal = {Front Endocrinol (Lausanne)},

volume = {15},

pages = {1312357},

abstract = {RASopathies are syndromes caused by congenital defects in the Ras/mitogen-activated protein kinase (MAPK) pathway genes, with a population prevalence of 1 in 1,000. Patients are typically identified in childhood based on diverse characteristic features, including cryptorchidism (CR) in >50% of affected men. As CR predisposes to spermatogenic failure (SPGF; total sperm count per ejaculate 0-39 million), we hypothesized that men seeking infertility management include cases with undiagnosed RASopathies. Likely pathogenic or pathogenic (LP/P) variants in 22 RASopathy-linked genes were screened in 521 idiopathic SPGF patients (including 155 CR cases) and 323 normozoospermic controls using exome sequencing. All 844 men were recruited to the ESTonian ANDrology (ESTAND) cohort and underwent identical andrological phenotyping. RASopathy-specific variant interpretation guidelines were used for pathogenicity assessment. LP/P variants were identified in  (two),  (three),  (one),  (one),  (one),  (one), and  (one). The findings affected six of 155 cases with CR and SPGF, three of 366 men with SPGF only, and one (of 323) normozoospermic subfertile man. The subgroup "CR and SPGF" had over 13-fold enrichment of findings compared to controls (3.9%  0.3%; Fisher's exact test,  = 5.5 × 10). All ESTAND subjects with LP/P variants in the Ras/MAPK pathway genes presented congenital genitourinary anomalies, skeletal and joint conditions, and other RASopathy-linked health concerns. Rare forms of malignancies (schwannomatosis and pancreatic and testicular cancer) were reported on four occasions. The Genetics of Male Infertility Initiative (GEMINI) cohort (1,416 SPGF cases and 317 fertile men) was used to validate the outcome. LP/P variants in  (three),  (three), and  (one) were identified in six SPGF cases (including 4/31 GEMINI cases with CR) and one normozoospermic man. Undiagnosed RASopathies were detected in total for 17 ESTAND and GEMINI subjects, 15 SPGF patients (10 with CR), and two fertile men. Affected RASopathy genes showed high expression in spermatogenic and testicular somatic cells. In conclusion, congenital defects in the Ras/MAPK pathway genes represent a new congenital etiology of syndromic male infertility. Undiagnosed RASopathies were especially enriched among patients with a history of cryptorchidism. Given the relationship between RASopathies and other conditions, infertile men found to have this molecular diagnosis should be evaluated for known RASopathy-linked health concerns, including specific rare malignancies.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37937809,

title = {Deleterious genetic changes in AGTPBP1 result in teratozoospermia with sperm head and flagella defects},

author = {Yu-Hua Lin and Ya-Yun Wang and Tsung-Hsuan Lai and Jih-Lung Teng and Chi-Wei Lin and Chih-Chun Ke and I-Shing Yu and Hui-Ling Lee and Chying-Chyuan Chan and Chi-Hua Tung and Donald F Conrad and Moira K O'Bryan and Ying-Hung Lin},

doi = {10.1111/jcmm.18031},

issn = {1582-4934},

year  = {2024},

date = {2024-01-01},

journal = {J Cell Mol Med},

volume = {28},

number = {2},

pages = {e18031},

abstract = {Approximately 10%-15% of couples worldwide are infertile, and male factors account for approximately half of these cases. Teratozoospermia is a major cause of male infertility. Although various mutations have been identified in teratozoospermia, these can vary among ethnic groups. In this study, we performed whole-exome sequencing to identify genetic changes potentially causative of teratozoospermia. Out of seven genes identified, one, ATP/GTP Binding Protein 1 (AGTPBP1), was characterized, and three missense changes were identified in two patients (Affected A: p.Glu423Asp and p.Pro631Leu; Affected B: p.Arg811His). In those two cases, severe sperm head and tail defects were observed. Moreover, AGTPBP1 localization showed a fragmented pattern compared to control participants, with specific localization in the neck and annulus regions. Using murine models, we found that AGTPBP1 is localized in the manchette structure, which is essential for sperm structure formation. Additionally, in Agtpbp1-null mice, we observed sperm head and tail defects similar to those in sperm from AGTPBP1-mutated cases, along with abnormal polyglutamylation tubulin and decreasing △-2 tubulin levels. In this study, we established a link between genetic changes in AGTPBP1 and human teratozoospermia for the first time and identified the role of AGTPBP1 in deglutamination, which is crucial for sperm formation.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38073178,

title = {Genome sequencing of Pakistani families with male infertility identifies deleterious genotypes in SPAG6, CCDC9, TKTL1, TUBA3C, and M1AP},

author = {Muhammad Riaz Khan and Arvand Akbari and Thomas J Nicholas and Helen Castillo-Madeen and Muhammad Ajmal and Taqweem Ul Haq and Maris Laan and Aaron R Quinlan and Jasvinder S Ahuja and Aftab Ali Shah and Donald F Conrad},

doi = {10.1111/andr.13570},

issn = {2047-2927},

year  = {2023},

date = {2023-12-01},

urldate = {2023-12-01},

journal = {Andrology},

abstract = {BACKGROUND: There are likely to be hundreds of monogenic forms of human male infertility. Whole genome sequencing (WGS) is the most efficient way to make progress in mapping the causative genetic variants, and ultimately improve clinical management of the disease in each patient. Recruitment of consanguineous families is an effective approach to ascertain the genetic forms of many diseases.nnOBJECTIVES: To apply WGS to large consanguineous families with likely hereditary male infertility and identify potential genetic cases.nnMATERIALS AND METHODS: We recruited seven large families with clinically diagnosed male infertility from rural Pakistan, including five with a history of consanguinity. We generated WGS data on 26 individuals (3-5 per family) and analyzed the resulting data with a computational pipeline to identify potentially causal single nucleotide variants, indels, and copy number variants.nnRESULTS: We identified plausible genetic causes in five of the seven families, including a homozygous 10 kb deletion of exon 2 in a well-established male infertility gene (M1AP), and biallelic missense substitutions (SPAG6, CCDC9, TUBA3C) and an in-frame hemizygous deletion (TKTL1) in genes with emerging relevance.nnDISCUSSION AND CONCLUSION: The rate of genetic findings using the current approach (71%) was much higher than what we recently achieved using whole-exome sequencing (WES) of unrelated singleton cases (20%). Furthermore, we identified a pathogenic single-exon deletion in M1AP that would be undetectable by WES. Screening more families with WGS, especially in underrepresented populations, will further reveal the types of variants underlying male infertility and accelerate the use of genetics in the patient management.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid38062027,

title = {TAD evolutionary and functional characterization reveals diversity in mammalian TAD boundary properties and function},

author = {Mariam Okhovat and Jake VanCampen and Kimberly A Nevonen and Lana Harshman and Weiyu Li and Cora E Layman and Samantha Ward and Jarod Herrera and Jackson Wells and Rory R Sheng and Yafei Mao and Blaise Ndjamen and Ana C Lima and Katinka A Vigh-Conrad and Alexandra M Stendahl and Ran Yang and Lev Fedorov and Ian R Matthews and Sarah A Easow and Dylan K Chan and Taha A Jan and Evan E Eichler and Sandra Rugonyi and Donald F Conrad and Nadav Ahituv and Lucia Carbone},

doi = {10.1038/s41467-023-43841-8},

issn = {2041-1723},

year  = {2023},

date = {2023-12-01},

urldate = {2023-12-01},

journal = {Nat Commun},

volume = {14},

number = {1},

pages = {8111},

abstract = {Topological associating domains (TADs) are self-interacting genomic units crucial for shaping gene regulation patterns. Despite their importance, the extent of their evolutionary conservation and its functional implications remain largely unknown. In this study, we generate Hi-C and ChIP-seq data and compare TAD organization across four primate and four rodent species and characterize the genetic and epigenetic properties of TAD boundaries in correspondence to their evolutionary conservation. We find 14% of all human TAD boundaries to be shared among all eight species (ultraconserved), while 15% are human-specific. Ultraconserved TAD boundaries have stronger insulation strength, CTCF binding, and enrichment of older retrotransposons compared to species-specific boundaries. CRISPR-Cas9 knockouts of an ultraconserved boundary in a mouse model lead to tissue-specific gene expression changes and morphological phenotypes. Deletion of a human-specific boundary near the autism-related AUTS2 gene results in the upregulation of this gene in neurons. Overall, our study provides pertinent TAD boundary evolutionary conservation annotations and showcases the functional importance of TAD evolution.},

keywords = {NHP Genetics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37984997,

title = {A naturally occurring variant of  causes maternal germline hypermutation in primates},

author = {Alexandra M Stendahl and Rashesh Sanghvi and Samuel Peterson and Karina Ray and Ana C Lima and Raheleh Rahbari and Donald F Conrad},

doi = {10.1101/gr.277977.123},

issn = {1549-5469},

year  = {2023},

date = {2023-12-01},

journal = {Genome Res},

volume = {33},

number = {12},

pages = {2053--2059},

abstract = {As part of an ongoing genome sequencing project at the Oregon National Primate Research Center, we identified a rhesus macaque with a rare homozygous frameshift mutation in the gene methyl-CpG binding domain 4, DNA glycosylase (). MBD4 is responsible for the repair of C > T deamination mutations at CpG dinucleotides and has been linked to somatic hypermutation and cancer predisposition in humans. We show here that MBD4-associated hypermutation also affects the germline: The six offspring of the -null dam have a fourfold to sixfold increase in de novo mutation burden. This excess burden was predominantly C > T mutations at CpG dinucleotides consistent with  loss of function in the dam. There was also a significant excess of C > T at CpA sites, indicating an important, unappreciated role for MBD4 to repair deamination in CpA contexts. The -null dam developed sustained eosinophilia later in life, but we saw no other signs of neoplastic processes associated with  loss of function in humans nor any obvious disease in the hypermutated offspring. This work provides the first evidence for a genetic factor causing hypermutation in the maternal germline of a mammal and adds to the very small list of naturally occurring variants known to modulate germline mutation rates in mammals.},

keywords = {NHP Genetics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37244752,

title = {Rhesus macaque fetal and placental growth demographics: A resource for laboratory animal researchers},

author = {Victoria H J Roberts and Jenna N Castro and Brady M Wessel and Donald F Conrad and Anne D Lewis and Jamie O Lo},

doi = {10.1002/ajp.23526},

issn = {1098-2345},

year  = {2023},

date = {2023-08-01},

urldate = {2023-08-01},

journal = {Am J Primatol},

volume = {85},

number = {8},

pages = {e23526},

abstract = {Rhesus macaques (Macaca mulatta) are amongst the most common nonhuman primate species used in biomedical research. These animals provide a precious resource for translational studies and opportunities to maximize rhesus data use are encouraged. Here we compile data produced from 10 years of investigator-driven pregnancy studies conducted at the Oregon National Primate Research Center (ONPRC). All pregnancies were generated within the consistent and reproducible protocols of the ONPRC time-mated breeding program. The data included are from control animals who did not experience in utero perturbations or experimental manipulations. A total of 86 pregnant rhesus macaques were delivered by cesarean section over a range of gestational days (G) 50 to G159 (where term is G165 ± 10 days in the rhesus macaque), with subsequent immediate tissue harvesting following standardized protocols. Fetal and placental growth measures, and all major organ weights are reported. All data are presented relative to gestational age for the entire cohort and in addition, data are stratified by fetal sex. The outcome is a large reference resource for use by laboratory animal researchers in future comparative fetal development studies.},

keywords = {NHP Genetics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37459509,

title = {In vivo versus in silico assessment of potentially pathogenic missense variants in human reproductive genes},

author = {Xinbao Ding and Priti Singh and Kerry Schimenti and Tina N Tran and Robert Fragoza and Jimmaline Hardy and Kyle E Orwig and Marta Olszewska and Maciej K Kurpisz and Alexander N Yatsenko and Donald F Conrad and Haiyuan Yu and John C Schimenti},

doi = {10.1073/pnas.2219925120},

issn = {1091-6490},

year  = {2023},

date = {2023-07-01},

journal = {Proc Natl Acad Sci U S A},

volume = {120},

number = {30},

pages = {e2219925120},

abstract = {Infertility is a heterogeneous condition, with genetic causes thought to underlie a substantial fraction of cases. Genome sequencing is becoming increasingly important for genetic diagnosis of diseases including idiopathic infertility; however, most rare or minor alleles identified in patients are variants of uncertain significance (VUS). Interpreting the functional impacts of VUS is challenging but profoundly important for clinical management and genetic counseling. To determine the consequences of these variants in key fertility genes, we functionally evaluated 11 missense variants in the genes , , ,  and  by generating genome-edited mouse models. Nine variants were classified as deleterious by most functional prediction algorithms, and two disrupted a protein-protein interaction (PPI) in the yeast two hybrid (Y2H) assay. Though these genes are essential for normal meiosis or spermiogenesis in mice, only one variant, observed in the  gene of a male infertility patient, compromised fertility or gametogenesis in the mouse models. To explore the disconnect between predictions and outcomes, we compared pathogenicity calls of missense variants made by ten widely used algorithms to 1) those annotated in ClinVar and 2) those evaluated in mice. All the algorithms performed poorly in terms of predicting the effects of human missense variants modeled in mice. These studies emphasize caution in the genetic diagnoses of infertile patients based primarily on pathogenicity prediction algorithms and emphasize the need for alternative and efficient in vitro or in vivo functional validation models for more effective and accurate VUS description to either pathogenic or benign categories.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37267208,

title = {Consensus label propagation with graph convolutional networks for single-cell RNA sequencing cell type annotation},

author = {Daniel P Lewinsohn and Katinka A Vigh-Conrad and Donald F Conrad and Cory B Scott},

doi = {10.1093/bioinformatics/btad360},

issn = {1367-4811},

year  = {2023},

date = {2023-06-01},

urldate = {2023-06-01},

journal = {Bioinformatics},

volume = {39},

number = {6},

abstract = {MOTIVATION: Single-cell RNA sequencing (scRNA-seq) data, annotated by cell type, is useful in a variety of downstream biological applications, such as profiling gene expression at the single-cell level. However, manually assigning these annotations with known marker genes is both time-consuming and subjective.nnRESULTS: We present a Graph Convolutional Network (GCN)-based approach to automate the annotation process. Our process builds upon existing labeling approaches, using state-of-the-art tools to find cells with highly confident label assignments through consensus and spreading these confident labels with a semi-supervised GCN. Using simulated data and two scRNA-seq datasets from different tissues, we show that our method improves accuracy over a simple consensus algorithm and the average of the underlying tools. We also compare our method to a nonparametric neighbor majority approach, showing comparable results. We then demonstrate that our GCN method allows for feature interpretation, identifying important genes for cell type classification. We present our completed pipeline, written in PyTorch, as an end-to-end tool for automating and interpreting the classification of scRNA-seq data.nnAVAILABILITY AND IMPLEMENTATION: Our code for conducting the experiments in this paper and using our model is available at https://github.com/lewinsohndp/scSHARP.},

keywords = {Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37053313,

title = {The origins and functional effects of postzygotic mutations throughout the human life span},

author = {Nicole B Rockweiler and Avinash Ramu and Liina Nagirnaja and Wing H Wong and Michiel J Noordam and Casey W Drubin and Ni Huang and Brian Miller and Ellen Z Todres and Katinka A Vigh-Conrad and Antonino Zito and Kerrin S Small and Kristin G Ardlie and Barak A Cohen and Donald F Conrad},

doi = {10.1126/science.abn7113},

issn = {1095-9203},

year  = {2023},

date = {2023-04-01},

urldate = {2023-04-01},

journal = {Science},

volume = {380},

number = {6641},

pages = {eabn7113},

abstract = {Postzygotic mutations (PZMs) begin to accrue in the human genome immediately after fertilization, but how and when PZMs affect development and lifetime health remain unclear. To study the origins and functional consequences of PZMs, we generated a multitissue atlas of PZMs spanning 54 tissue and cell types from 948 donors. Nearly half the variation in mutation burden among tissue samples can be explained by measured technical and biological effects, and 9% can be attributed to donor-specific effects. Through phylogenetic reconstruction of PZMs, we found that their type and predicted functional impact vary during prenatal development, across tissues, and through the germ cell life cycle. Thus, methods for interpreting effects across the body and the life span are needed to fully understand the consequences of genetic variants.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid36997603,

title = {DDX3Y is likely the key spermatogenic factor in the AZFa region that contributes to human non-obstructive azoospermia},

author = {Ann-Kristin Dicke and Adrian Pilatz and Margot J Wyrwoll and Margus Punab and Christian Ruckert and Liina Nagirnaja and Kenneth I Aston and Donald F Conrad and Sara Di Persio and Nina Neuhaus and Daniela Fietz and Maris Laan and Birgit Stallmeyer and Frank Tüttelmann},

doi = {10.1038/s42003-023-04714-4},

issn = {2399-3642},

year  = {2023},

date = {2023-03-01},

journal = {Commun Biol},

volume = {6},

number = {1},

pages = {350},

abstract = {Non-obstructive azoospermia, the absence of sperm in the ejaculate due to disturbed spermatogenesis, represents the most severe form of male infertility. De novo microdeletions of the Y-chromosomal AZFa region are one of few well-established genetic causes for NOA and are routinely analysed in the diagnostic workup of affected men. So far, it is unclear which of the three genes located in the AZFa chromosomal region is indispensible for germ cell maturation. Here we present four different likely pathogenic loss-of-function variants in the AZFa gene DDX3Y identified by analysing exome sequencing data of more than 1,600 infertile men. Three of the patients underwent testicular sperm extraction and revealed the typical AZFa testicular Sertoli cell-only phenotype. One of the variants was proven to be de novo. Consequently, DDX3Y represents the AZFa key spermatogenic factor and screening for variants in DDX3Y should be included in the diagnostic workflow.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid37234866,

title = {Genomic study of  variants: prevalence and allelic heterogeneity in men with spermatogenic failure},

author = {Sidra Qureshi and Jimmaline J Hardy and Christopher Pombar and Andrea J Berman and Agnieszka Malcher and Tara Gingrich and Rachel Hvasta and Jannah Kuong and Sarah Munyoki and Kathleen Hwang and Kyle E Orwig and Jawad Ahmed and Marta Olszewska and Maciej Kurpisz and Donald F Conrad and Muhammad Jaseem Khan and Alexander N Yatsenko},

doi = {10.3389/fgene.2023.1134849},

issn = {1664-8021},

year  = {2023},

date = {2023-01-01},

journal = {Front Genet},

volume = {14},

pages = {1134849},

abstract = { Human spermatogenesis is a highly intricate process that requires the input of thousands of testis-specific genes. Defects in any of them at any stage of the process can have detrimental effects on sperm production and/or viability. In particular, the function of many meiotic proteins encoded by germ cell specific genes is critical for maturation of haploid spermatids and viable spermatozoa, necessary for fertilization, and is also extremely sensitive to even the slightest change in coding DNA.  Here, using whole exome and genome approaches, we identified and reported novel, clinically significant variants in testis-expressed gene 15 (), in unrelated men with spermatogenic failure (SPGF).  TEX15 mediates double strand break repair during meiosis. Recessive loss-of-function (LOF)  mutations are associated with SPGF in humans and knockout male mice are infertile. We expand earlier reports documenting heterogeneous allelic pathogenic  variants that cause a range of SPGF phenotypes from oligozoospermia (low sperm) to nonobstructive azoospermia (no sperm) with meiotic arrest and report the prevalence of 0.6% of  variants in our patient cohort. Among identified possible LOF variants, one homozygous missense substitution c.6835G>A (p.Ala2279Thr) co-segregated with cryptozoospermia in a family with SPGF. Additionally, we observed numerous cases of inferred  compound heterozygous variants in  among unrelated individuals with varying degrees of SPGF. Variants included splice site, insertions/deletions (indels), and missense substitutions, many of which resulted in LOF effects (i.e., frameshift, premature stop, alternative splicing, or potentially altered posttranslational modification sites).  In conclusion, we performed an extensive genomic study of familial and sporadic SPGF and identified potentially damaging TEX15 variants in 7 of 1097 individuals of our combined cohorts. We hypothesize that SPGF phenotype severity is dictated by individual  variant's impact on structure and function. Resultant LOFs likely have deleterious effects on crossover/recombination in meiosis. Our findings support the notion of increased gene variant frequency in SPGF and its genetic and allelic heterogeneity as it relates to complex disease such as male infertility.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid36572685,

title = {Diverse monogenic subforms of human spermatogenic failure},

author = {Liina Nagirnaja and Alexandra M Lopes and Wu-Lin Charng and Brian Miller and Rytis Stakaitis and Ieva Golubickaite and Alexandra Stendahl and Tianpengcheng Luan and Corinna Friedrich and Eisa Mahyari and Eloise Fadial and Laura Kasak and Katinka Vigh-Conrad and Manon S Oud and Miguel J Xavier and Samuel R Cheers and Emma R James and Jingtao Guo and Timothy G Jenkins and Antoni Riera-Escamilla and Alberto Barros and Filipa Carvalho and Susana Fernandes and João Gonçalves and Christina A Gurnett and Niels Jørgensen and Davor Jezek and Emily S Jungheim and Sabine Kliesch and Robert I McLachlan and Kenan R Omurtag and Adrian Pilatz and Jay I Sandlow and James Smith and Michael L Eisenberg and James M Hotaling and Keith A Jarvi and Margus Punab and Ewa Rajpert-De Meyts and Douglas T Carrell and Csilla Krausz and Maris Laan and Moira K O'Bryan and Peter N Schlegel and Frank Tüttelmann and Joris A Veltman and Kristian Almstrup and Kenneth I Aston and Donald F Conrad},

doi = {10.1038/s41467-022-35661-z},

issn = {2041-1723},

year  = {2022},

date = {2022-12-01},

urldate = {2022-12-01},

journal = {Nat Commun},

volume = {13},

number = {1},

pages = {7953},

abstract = {Non-obstructive azoospermia (NOA) is the most severe form of male infertility and typically incurable. Defining the genetic basis of NOA has proven challenging, and the most advanced classification of NOA subforms is not based on genetics, but simple description of testis histology. In this study, we exome-sequenced over 1000 clinically diagnosed NOA cases and identified a plausible recessive Mendelian cause in 20%. We find further support for 21 genes in a 2-stage burden test with 2072 cases and 11,587 fertile controls. The disrupted genes are primarily on the autosomes, enriched for undescribed human "knockouts", and, for the most part, have yet to be linked to a Mendelian trait. Integration with single-cell RNA sequencing data shows that azoospermia genes can be grouped into molecular subforms with synchronized expression patterns, and analogs of these subforms exist in mice. This analysis framework identifies groups of genes with known roles in spermatogenesis but also reveals unrecognized subforms, such as a set of genes expressed across mitotic divisions of differentiating spermatogonia. Our findings highlight NOA as an understudied Mendelian disorder and provide a conceptual structure for organizing the complex genetics of male infertility, which may provide a rational basis for disease classification.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid36214638,

title = {SATINN: an automated neural network-based classification of testicular sections allows for high-throughput histopathology of mouse mutants},

author = {Ran Yang and Alexandra M Stendahl and Katinka A Vigh-Conrad and Madison Held and Ana C Lima and Donald F Conrad},

doi = {10.1093/bioinformatics/btac673},

issn = {1367-4811},

year  = {2022},

date = {2022-11-01},

urldate = {2022-11-01},

journal = {Bioinformatics},

volume = {38},

number = {23},

pages = {5288--5298},

abstract = {MOTIVATION: The mammalian testis is a complex organ with a cellular composition that changes smoothly and cyclically in normal adults. While testis histology is already an invaluable tool for identifying and describing developmental differences in evolution and disease, methods for standardized, digital image analysis of testis are needed to expand the utility of this approach.nnRESULTS: We developed SATINN (Software for Analysis of Testis Images with Neural Networks), a multi-level framework for automated analysis of multiplexed immunofluorescence images from mouse testis. This approach uses residual learning to train convolutional neural networks (CNNs) to classify nuclei from seminiferous tubules into seven distinct cell types with an accuracy of 81.7%. These cell classifications are then used in a second-level tubule CNN, which places seminiferous tubules into one of 12 distinct tubule stages with 57.3% direct accuracy and 94.9% within ±1 stage. We further describe numerous cell- and tubule-level statistics that can be derived from wild-type testis. Finally, we demonstrate how the classifiers and derived statistics can be used to rapidly and precisely describe pathology by applying our methods to image data from two mutant mouse lines. Our results demonstrate the feasibility and potential of using computer-assisted analysis for testis histology, an area poised to evolve rapidly on the back of emerging, spatially resolved genomic and proteomic technologies.nnAVAILABILITY AND IMPLEMENTATION: The source code to reproduce the results described here and a SATINN standalone application with graphic-user interface are available from http://github.com/conradlab/SATINN.nnSUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid36150389,

title = {The piRNA-pathway factor FKBP6 is essential for spermatogenesis but dispensable for control of meiotic LINE-1 expression in humans},

author = {Margot J Wyrwoll and Channah M Gaasbeek and Ieva Golubickaite and Rytis Stakaitis and Manon S Oud and Liina Nagirnaja and Camille Dion and Emad B Sindi and Harry G Leitch and Channa N Jayasena and Anu Sironen and Ann-Kristin Dicke and Nadja Rotte and Birgit Stallmeyer and Sabine Kliesch and Carlos H P Grangeiro and Thaís F Araujo and Paul Lasko and  and Kathleen D'Hauwers and Roos M Smits and Liliana Ramos and Miguel J Xavier and Don F Conrad and Kristian Almstrup and Joris A Veltman and Frank Tüttelmann and Godfried W van der Heijden},

doi = {10.1016/j.ajhg.2022.09.002},

issn = {1537-6605},

year  = {2022},

date = {2022-10-01},

journal = {Am J Hum Genet},

volume = {109},

number = {10},

pages = {1850--1866},

abstract = {Infertility affects around 7% of the male population and can be due to severe spermatogenic failure (SPGF), resulting in no or very few sperm in the ejaculate. We initially identified a homozygous frameshift variant in FKBP6 in a man with extreme oligozoospermia. Subsequently, we screened a total of 2,699 men with SPGF and detected rare bi-allelic loss-of-function variants in FKBP6 in five additional persons. All six individuals had no or extremely few sperm in the ejaculate, which were not suitable for medically assisted reproduction. Evaluation of testicular tissue revealed an arrest at the stage of round spermatids. Lack of FKBP6 expression in the testis was confirmed by RT-qPCR and immunofluorescence staining. In mice, Fkbp6 is essential for spermatogenesis and has been described as being involved in piRNA biogenesis and formation of the synaptonemal complex (SC). We did not detect FKBP6 as part of the SC in normal human spermatocytes, but small RNA sequencing revealed that loss of FKBP6 severely impacted piRNA levels, supporting a role for FKBP6 in piRNA biogenesis in humans. In contrast to findings in piRNA-pathway mouse models, we did not detect an increase in LINE-1 expression in men with pathogenic FKBP6 variants. Based on our findings, FKBP6 reaches a "strong" level of evidence for being associated with male infertility according to the ClinGen criteria, making it directly applicable for clinical diagnostics. This will improve patient care by providing a causal diagnosis and will help to predict chances for successful surgical sperm retrieval.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35850260,

title = {DDB1- and CUL4-associated factor 12-like protein 1 (Dcaf12l1) is not essential for male fertility in mice},

author = {Brendan J Houston and Alexandra M Lopes and Maris Laan and Liina Nagirnaja and Anne E O'Connor and D Jo Merriner and Joseph Nguyen and Margus Punab and Antoni Riera-Escamilla and Csilla Krausz and Kenneth Ivan Aston and Donald F Conrad and Moira K O'Bryan},

doi = {10.1016/j.ydbio.2022.07.006},

issn = {1095-564X},

year  = {2022},

date = {2022-10-01},

journal = {Dev Biol},

volume = {490},

pages = {66--72},

abstract = {Male infertility is a common condition affecting at least 7% of men worldwide and is often genetic in origin. Using whole exome sequencing, we recently discovered three hemizygous, likely damaging variants in DDB1- and CUL4-associated factor 12-like protein 1 (DCAF12L1) in men with azoospermia. DCAF12L1 is located on the X-chromosome and as identified by single cell sequencing studies, its expression is enriched in human testes and specifically in Sertoli cells and spermatogonia. However, very little is known about the role of DCAF12L1 in spermatogenesis, thus we generated a knockout mouse model to further explore the role of DCAF12L1 in male fertility. Knockout mice were generated using CRISPR/Cas9 technology to remove the entire coding region of Dcaf12l1 and were assessed for fertility over a broad range of ages (2-8 months of age). Despite outstanding genetic evidence in men, loss of DCAF12L1 had no discernible impact on male fertility in mice, as highlighted by breeding trials, histological assessment of the testis and epididymis, daily sperm production and evaluation of sperm motility using computer assisted methods. This disparity is likely due to the parallel evolution, and subsequent divergence, of DCAF12 family members in mice and men or the presence of compounding environmental factors in men.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35679955,

title = {Zinc finger RNA binding protein 2 (ZFR2) is not required for male fertility in the mouse},

author = {Lachlan M Cauchi and Brendan J Houston and Liina Nagirnaja and Anne E O'Connor and D Jo Merriner and Kenneth I Aston and Peter N Schlegel and Don F Conrad and Richard Burke and Moira K O'Bryan},

doi = {10.1016/j.ydbio.2022.05.020},

issn = {1095-564X},

year  = {2022},

date = {2022-09-01},

journal = {Dev Biol},

volume = {489},

pages = {55--61},

abstract = {BACKGROUND: Thousands of genes are expressed during spermatogenesis and male infertility has a strong genetic component. Within this study, we focus on the role of Zfr2 in male fertility, a gene previously implicated in human male fertility. To date, very little is known about the role of ZFR2 in either humans or mice. To this end, the requirement for ZFR2 in male fertility was assessed using a knockout mouse model.nnRESULTS: Zfr2 was found to be expressed in the testes of both humans and mice. Deletion of Zfr2 was achieved via removal of exon 2 using CRISPR-Cas9 methods. The absence of Zfr2 did not result in a reduction in any fertility parameters assessed. Knockout males were capable of fostering litter sizes equal to wild type males, and there were no effects of Zfr2 knockout on sperm number or motility. We note Zfr2 knockout females were also fertile.nnCONCLUSIONS: The absence of Zfr2 alone is not sufficient to cause a reduction in male fertility in mice.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35809576,

title = {Large-scale analyses of the X chromosome in 2,354 infertile men discover recurrently affected genes associated with spermatogenic failure},

author = {Antoni Riera-Escamilla and Matthias Vockel and Liina Nagirnaja and Miguel J Xavier and Albert Carbonell and Daniel Moreno-Mendoza and Marc Pybus and Ginevra Farnetani and Viktoria Rosta and Francesca Cioppi and Corinna Friedrich and Manon S Oud and Godfried W van der Heijden and Armin Soave and Thorsten Diemer and Elisabet Ars and Josvany Sánchez-Curbelo and Sabine Kliesch and Moira K O'Bryan and Eduard Ruiz-Castañe and  and Fernando Azorín and Joris A Veltman and Kenneth I Aston and Donald F Conrad and Frank Tüttelmann and Csilla Krausz},

doi = {10.1016/j.ajhg.2022.06.007},

issn = {1537-6605},

year  = {2022},

date = {2022-08-01},

journal = {Am J Hum Genet},

volume = {109},

number = {8},

pages = {1458--1471},

abstract = {Although the evolutionary history of the X chromosome indicates its specialization in male fitness, its role in spermatogenesis has largely been unexplored. Currently only three X chromosome genes are considered of moderate-definitive diagnostic value. We aimed to provide a comprehensive analysis of all X chromosome-linked protein-coding genes in 2,354 azoospermic/cryptozoospermic men from four independent cohorts. Genomic data were analyzed and compared with data in normozoospermic control individuals and gnomAD. While updating the clinical significance of known genes, we propose 21 recurrently mutated genes strongly associated with and 34 moderately associated with azoospermia/cryptozoospermia not previously linked to male infertility (novel). The most frequently affected prioritized gene, RBBP7, was found mutated in ten men across all cohorts, and our functional studies in Drosophila support its role in germ stem cell maintenance. Collectively, our study represents a significant step towards the definition of the missing genetic etiology in idiopathic severe spermatogenic failure and significantly reduces the knowledge gap of X-linked genetic causes of azoospermia/cryptozoospermia contributing to the development of future diagnostic gene panels.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35535697,

title = {Actionable secondary findings following exome sequencing of 836 non-obstructive azoospermia cases and their value in patient management},

author = {Laura Kasak and Kristiina Lillepea and Liina Nagirnaja and Kenneth I Aston and Peter N Schlegel and João Gonçalves and Filipa Carvalho and Daniel Moreno-Mendoza and Kristian Almstrup and Michael L Eisenberg and Keith A Jarvi and Moira K O'Bryan and Alexandra M Lopes and Donald F Conrad and  and Margus Punab and Maris Laan},

doi = {10.1093/humrep/deac100},

issn = {1460-2350},

year  = {2022},

date = {2022-06-01},

journal = {Hum Reprod},

volume = {37},

number = {7},

pages = {1652--1663},

abstract = {STUDY QUESTION: What is the load, distribution and added clinical value of secondary findings (SFs) identified in exome sequencing (ES) of patients with non-obstructive azoospermia (NOA)?nnSUMMARY ANSWER: One in 28 NOA cases carried an identifiable, medically actionable SF.nnWHAT IS KNOWN ALREADY: In addition to molecular diagnostics, ES allows assessment of clinically actionable disease-related gene variants that are not connected to the patient's primary diagnosis, but the knowledge of which may allow the prevention, delay or amelioration of late-onset monogenic conditions. Data on SFs in specific clinical patient groups, including reproductive failure, are currently limited.nnSTUDY DESIGN, SIZE, DURATION: The study group was a retrospective cohort of patients with NOA recruited in 10 clinics across six countries and formed in the framework of the international GEMINI (The GEnetics of Male INfertility Initiative) study.nnPARTICIPANTS/MATERIALS, SETTING, METHODS: ES data of 836 patients with NOA were exploited to analyze SFs in 85 genes recommended by the American College of Medical Genetics and Genomics (ACMG), Geisinger's MyCode, and Clinical Genome Resource. The identified 6374 exonic variants were annotated with ANNOVAR and filtered for allele frequency, retaining 1381 rare or novel missense and loss-of-function variants. After automatic assessment of pathogenicity with ClinVar and InterVar, 87 variants were manually curated. The final list of confident disease-causing SFs was communicated to the corresponding GEMINI centers. When patient consent had been given, available family health history and non-andrological medical data were retrospectively assessed.nnMAIN RESULTS AND THE ROLE OF CHANCE: We found a 3.6% total frequency of SFs, 3.3% from the 59 ACMG SF v2.0 genes. One in 70 patients carried SFs in genes linked to familial cancer syndromes, whereas 1 in 60 cases was predisposed to congenital heart disease or other cardiovascular conditions. Retrospective assessment confirmed clinico-molecular diagnoses in several cases. Notably, 37% (11/30) of patients with SFs carried variants in genes linked to male infertility in mice, suggesting that some SFs may have a co-contributing role in spermatogenic impairment. Further studies are needed to determine whether these observations represent chance findings or the profile of SFs in NOA patients is indeed different from the general population.nnLIMITATIONS, REASONS FOR CAUTION: One limitation of our cohort was the low proportion of non-Caucasian ethnicities (9%). Additionally, as comprehensive clinical data were not available retrospectively for all men with SFs, we were not able to confirm a clinico-molecular diagnosis and assess the penetrance of the specific variants.nnWIDER IMPLICATIONS OF THE FINDINGS: For the first time, this study analyzed medically actionable SFs in men with spermatogenic failure. With the evolving process to incorporate ES into routine andrology practice for molecular diagnostic purposes, additional assessment of SFs can inform about future significant health concerns for infertility patients. Timely detection of SFs and respective genetic counseling will broaden options for disease prevention and early treatment, as well as inform choices and opportunities regarding family planning. A notable fraction of SFs was detected in genes implicated in maintaining genome integrity, essential in both mitosis and meiosis. Thus, potential genetic pleiotropy may exist between certain adult-onset monogenic diseases and NOA.nnSTUDY FUNDING/COMPETING INTEREST(S): This work was supported by the Estonian Research Council grants IUT34-12 and PRG1021 (M.L. and M.P.); National Institutes of Health of the United States of America grant R01HD078641 (D.F.C., K.I.A. and P.N.S.); National Institutes of Health of the United States of America grant P50HD096723 (D.F.C. and P.N.S.); National Health and Medical Research Council of Australia grant APP1120356 (M.K.O'B., D.F.C. and K.I.A.); Fundação para a Ciência e a Tecnologia (FCT)/Ministério da Ciência, Tecnologia e Inovação grant POCI-01-0145-FEDER-007274 (A.M.L., F.C. and J.G.) and FCT: IF/01262/2014 (A.M.L.). J.G. was partially funded by FCT/Ministério da Ciência, Tecnologia e Ensino Superior (MCTES), through the Centre for Toxicogenomics and Human Health-ToxOmics (grants UID/BIM/00009/2016 and UIDB/00009/2020). M.L.E. is a consultant for, and holds stock in, Roman, Sandstone, Dadi, Hannah, Underdog and has received funding from NIH/NICHD. Co-authors L.K., K.L., L.N., K.I.A., P.N.S., J.G., F.C., D.M.-M., K.A., K.A.J., M.K.O'B., A.M.L., D.F.C., M.P. and M.L. declare no conflict of interest.nnTRIAL REGISTRATION NUMBER: N/A.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35022746,

title = {Human INHBB Gene Variant (c.1079T>C:p.Met360Thr) Alters Testis Germ Cell Content, but Does Not Impact Fertility in Mice},

author = {Brendan J Houston and Anne E O'Connor and Degang Wang and Georgia Goodchild and D Jo Merriner and Haitong Luan and Don F Conrad and Liina Nagirnaja and Kenneth I Aston and Sabine Kliesch and Margot J Wyrwoll and Corinna Friedrich and Frank Tüttelmann and Craig Harrison and Moira K O'Bryan and Kelly Walton},

doi = {10.1210/endocr/bqab269},

issn = {1945-7170},

year  = {2022},

date = {2022-03-01},

journal = {Endocrinology},

volume = {163},

number = {3},

abstract = {Testicular-derived inhibin B (α/β B dimers) acts in an endocrine manner to suppress pituitary production of follicle-stimulating hormone (FSH), by blocking the actions of activins (β A/B/β A/B dimers). Previously, we identified a homozygous genetic variant (c.1079T>C:p.Met360Thr) arising from uniparental disomy of chromosome 2 in the INHBB gene (β B-subunit of inhibin B and activin B) in a man suffering from infertility (azoospermia). In this study, we aimed to test the causality of the p.Met360Thr variant in INHBB and testis function. Here, we used CRISPR/Cas9 technology to generate InhbbM364T/M364T mice, where mouse INHBB p.Met364 corresponds with human p.Met360. Surprisingly, we found that the testes of male InhbbM364T/M364T mutant mice were significantly larger compared with those of aged-matched wildtype littermates at 12 and 24 weeks of age. This was attributed to a significant increase in Sertoli cell and round spermatid number and, consequently, seminiferous tubule area in InhbbM364T/M364T males compared to wildtype males. Despite this testis phenotype, male InhbbM364T/M364T mutant mice retained normal fertility. Serum hormone analyses, however, indicated that the InhbbM364T variant resulted in reduced circulating levels of activin B but did not affect FSH production. We also examined the effect of this p.Met360Thr and an additional INHBB variant (c.314C>T: p.Thr105Met) found in another infertile man on inhibin B and activin B in vitro biosynthesis. We found that both INHBB variants resulted in a significant disruption to activin B in vitro biosynthesis. Together, this analysis supports that INHBB variants that limit activin B production have consequences for testis composition in males.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid35013161,

title = {A de novo paradigm for male infertility},

author = {M S Oud and R M Smits and H E Smith and F K Mastrorosa and G S Holt and B J Houston and P F de Vries and B K S Alobaidi and L E Batty and H Ismail and J Greenwood and H Sheth and A Mikulasova and G D N Astuti and C Gilissen and K McEleny and H Turner and J Coxhead and S Cockell and D D M Braat and K Fleischer and K W M D'Hauwers and E Schaafsma and  and L Nagirnaja and D F Conrad and C Friedrich and S Kliesch and K I Aston and A Riera-Escamilla and C Krausz and C Gonzaga-Jauregui and M Santibanez-Koref and D J Elliott and L E L M Vissers and F Tüttelmann and M K O'Bryan and L Ramos and M J Xavier and G W van der Heijden and J A Veltman},

doi = {10.1038/s41467-021-27132-8},

issn = {2041-1723},

year  = {2022},

date = {2022-01-01},

journal = {Nat Commun},

volume = {13},

number = {1},

pages = {154},

abstract = {De novo mutations are known to play a prominent role in sporadic disorders with reduced fitness. We hypothesize that de novo mutations play an important role in severe male infertility and explain a portion of the genetic causes of this understudied disorder. To test this hypothesis, we utilize trio-based exome sequencing in a cohort of 185 infertile males and their unaffected parents. Following a systematic analysis, 29 of 145 rare (MAF < 0.1%) protein-altering de novo mutations are classified as possibly causative of the male infertility phenotype. We observed a significant enrichment of loss-of-function de novo mutations in loss-of-function-intolerant genes (p-value = 1.00 × 10) in infertile men compared to controls. Additionally, we detected a significant increase in predicted pathogenic de novo missense mutations affecting missense-intolerant genes (p-value = 5.01 × 10) in contrast to predicted benign de novo mutations. One gene we identify, RBM5, is an essential regulator of male germ cell pre-mRNA splicing and has been previously implicated in male infertility in mice. In a follow-up study, 6 rare pathogenic missense mutations affecting this gene are observed in a cohort of 2,506 infertile patients, whilst we find no such mutations in a cohort of 5,784 fertile men (p-value = 0.03). Our results provide evidence for the role of de novo mutations in severe male infertility and point to new candidate genes affecting fertility.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid34498060,

title = {A systematic review of the validated monogenic causes of human male infertility: 2020 update and a discussion of emerging gene-disease relationships},

author = {Brendan J Houston and Antoni Riera-Escamilla and Margot J Wyrwoll and Albert Salas-Huetos and Miguel J Xavier and Liina Nagirnaja and Corinna Friedrich and Don F Conrad and Kenneth I Aston and Csilla Krausz and Frank Tüttelmann and Moira K O'Bryan and Joris A Veltman and Manon S Oud},

doi = {10.1093/humupd/dmab030},

issn = {1460-2369},

year  = {2021},

date = {2021-12-01},

journal = {Hum Reprod Update},

volume = {28},

number = {1},

pages = {15--29},

abstract = {BACKGROUND: Human male infertility has a notable genetic component, including well-established diagnoses such as Klinefelter syndrome, Y-chromosome microdeletions and monogenic causes. Approximately 4% of all infertile men are now diagnosed with a genetic cause, but a majority (60-70%) remain without a clear diagnosis and are classified as unexplained. This is likely in large part due to a delay in the field adopting next-generation sequencing (NGS) technologies, and the absence of clear statements from field leaders as to what constitutes a validated cause of human male infertility (the current paper aims to address this). Fortunately, there has been a significant increase in the number of male infertility NGS studies. These have revealed a considerable number of novel gene-disease relationships (GDRs), which each require stringent assessment to validate the strength of genotype-phenotype associations. To definitively assess which of these GDRs are clinically relevant, the International Male Infertility Genomics Consortium (IMIGC) has identified the need for a systematic review and a comprehensive overview of known male infertility genes and an assessment of the evidence for reported GDRs.nnOBJECTIVE AND RATIONALE: In 2019, the first standardised clinical validity assessment of monogenic causes of male infertility was published. Here, we provide a comprehensive update of the subsequent 1.5 years, employing the joint expertise of the IMIGC to systematically evaluate all available evidence (as of 1 July 2020) for monogenic causes of isolated or syndromic male infertility, endocrine disorders or reproductive system abnormalities affecting the male sex organs. In addition, we systematically assessed the evidence for all previously reported possible monogenic causes of male infertility, using a framework designed for a more appropriate clinical interpretation of disease genes.nnSEARCH METHODS: We performed a literature search according to the PRISMA guidelines up until 1 July 2020 for publications in English, using search terms related to 'male infertility' in combination with the word 'genetics' in PubMed. Next, the quality and the extent of all evidence supporting selected genes were assessed using an established and standardised scoring method. We assessed the experimental quality, patient phenotype assessment and functional evidence based on gene expression, mutant in-vitro cell and in-vivo animal model phenotypes. A final score was used to determine the clinical validity of each GDR, across the following five categories: no evidence, limited, moderate, strong or definitive. Variants were also reclassified according to the American College of Medical Genetics and Genomics-Association for Molecular Pathology (ACMG-AMP) guidelines and were recorded in spreadsheets for each GDR, which are available at imigc.org.nnOUTCOMES: The primary outcome of this review was an overview of all known GDRs for monogenic causes of human male infertility and their clinical validity. We identified a total of 120 genes that were moderately, strongly or definitively linked to 104 infertility phenotypes.nnWIDER IMPLICATIONS: Our systematic review curates all currently available evidence to reveal the strength of GDRs in male infertility. The existing guidelines for genetic testing in male infertility cases are based on studies published 25 years ago, and an update is far overdue. The identification of 104 high-probability 'human male infertility genes' is a 33% increase from the number identified in 2019. The insights generated in the current review will provide the impetus for an update of existing guidelines, will inform novel evidence-based genetic testing strategies used in clinics, and will identify gaps in our knowledge of male infertility genetics. We discuss the relevant international guidelines regarding research related to gene discovery and provide specific recommendations to the field of male infertility. Based on our findings, the IMIGC consortium recommend several updates to the genetic testing standards currently employed in the field of human male infertility, most important being the adoption of exome sequencing, or at least sequencing of the genes validated in this study, and expanding the patient groups for which genetic testing is recommended.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid34626582,

title = {Comparative single-cell analysis of biopsies clarifies pathogenic mechanisms in Klinefelter syndrome},

author = {Eisa Mahyari and Jingtao Guo and Ana C Lima and Daniel P Lewinsohn and Alexandra M Stendahl and Katinka A Vigh-Conrad and Xichen Nie and Liina Nagirnaja and Nicole B Rockweiler and Douglas T Carrell and James M Hotaling and Kenneth I Aston and Donald F Conrad},

doi = {10.1016/j.ajhg.2021.09.001},

issn = {1537-6605},

year  = {2021},

date = {2021-10-01},

urldate = {2021-10-01},

journal = {Am J Hum Genet},

volume = {108},

number = {10},

pages = {1924--1945},

abstract = {Klinefelter syndrome (KS), also known as 47, XXY, is characterized by a distinct set of physiological abnormalities, commonly including infertility. The molecular basis for Klinefelter-related infertility is still unclear, largely because of the cellular complexity of the testis and the intricate endocrine and paracrine signaling that regulates spermatogenesis. Here, we demonstrate an analysis framework for dissecting human testis pathology that uses comparative analysis of single-cell RNA-sequencing data from the biopsies of 12 human donors. By comparing donors from a range of ages and forms of infertility, we generate gene expression signatures that characterize normal testicular function and distinguish clinically distinct forms of male infertility. Unexpectedly, we identified a subpopulation of Sertoli cells within multiple individuals with KS that lack transcription from the XIST locus, and the consequence of this is increased X-linked gene expression compared to all other KS cell populations. By systematic assessment of known cell signaling pathways, we identify 72 pathways potentially active in testis, dozens of which appear upregulated in KS. Altogether our data support a model of pathogenic changes in interstitial cells cascading from loss of X inactivation in pubertal Sertoli cells and nominate dosage-sensitive factors secreted by Sertoli cells that may contribute to the process. Our findings demonstrate the value of comparative patient analysis in mapping genetic mechanisms of disease and identify an epigenetic phenomenon in KS Sertoli cells that may prove important for understanding causes of infertility and sex chromosome evolution.},

keywords = {Human Reproduction, Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid34347949,

title = {Variant , Defective piRNA Processing, and Azoospermia},

author = {Liina Nagirnaja and Nina Mørup and John E Nielsen and Rytis Stakaitis and Ieva Golubickaite and Manon S Oud and Sofia B Winge and Filipa Carvalho and Kenneth I Aston and Francesca Khani and Godfried W van der Heijden and C Joana Marques and Niels E Skakkebaek and Ewa Rajpert-De Meyts and Peter N Schlegel and Niels Jørgensen and Joris A Veltman and Alexandra M Lopes and Donald F Conrad and Kristian Almstrup},

doi = {10.1056/NEJMoa2028973},

issn = {1533-4406},

year  = {2021},

date = {2021-08-01},

urldate = {2021-08-01},

journal = {N Engl J Med},

volume = {385},

number = {8},

pages = {707--719},

abstract = {BACKGROUND: P-element-induced wimpy testis (PIWI)-interacting RNAs (piRNAs) are short (21 to 35 nucleotides in length) and noncoding and are found almost exclusively in germ cells, where they regulate aberrant expression of transposable elements and postmeiotic gene expression. Critical to the processing of piRNAs is the protein poly(A)-specific RNase-like domain containing 1 (PNLDC1), which trims their 3' ends and, when disrupted in mice, causes azoospermia and male infertility.nnMETHODS: We performed exome sequencing on DNA samples from 924 men who had received a diagnosis of nonobstructive azoospermia. Testicular-biopsy samples were analyzed by means of histologic and immunohistochemical tests, in situ hybridization, reverse-transcriptase-quantitative-polymerase-chain-reaction assay, and small-RNA sequencing.nnRESULTS: Four unrelated men of Middle Eastern descent who had nonobstructive azoospermia were found to carry mutations in : the first patient had a biallelic stop-gain mutation, p.R452Ter (rs200629089; minor allele frequency, 0.00004); the second, a novel biallelic missense variant, p.P84S; the third, two compound heterozygous mutations consisting of p.M259T (rs141903829; minor allele frequency, 0.0007) and p.L35PfsTer3 (rs754159168; minor allele frequency, 0.00004); and the fourth, a novel biallelic canonical splice acceptor site variant, c.607-2A→T. Testicular histologic findings consistently showed error-prone meiosis and spermatogenic arrest with round spermatids of type Sa as the most advanced population of germ cells. Gene and protein expression of PNLDC1, as well as the piRNA-processing proteins PIWIL1, PIWIL4, MYBL1, and TDRKH, were greatly diminished in cells of the testes. Furthermore, the length distribution of piRNAs and the number of pachytene piRNAs was significantly altered in men carrying  mutations.nnCONCLUSIONS: Our results suggest a direct mechanistic effect of faulty piRNA processing on meiosis and spermatogenesis in men, ultimately leading to male infertility. (Funded by Innovation Fund Denmark and others.).},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid33963445,

title = {Variants in GCNA, X-linked germ-cell genome integrity gene, identified in men with primary spermatogenic failure},

author = {Jimmaline J Hardy and Margot J Wyrwoll and William Mcfadden and Agnieszka Malcher and Nadja Rotte and Nijole C Pollock and Sarah Munyoki and Maria V Veroli and Brendan J Houston and Miguel J Xavier and Laura Kasak and Margus Punab and Maris Laan and Sabine Kliesch and Peter Schlegel and Thomas Jaffe and Kathleen Hwang and Josip Vukina and Miguel A Brieño-Enríquez and Kyle Orwig and Judith Yanowitz and Michael Buszczak and Joris A Veltman and Manon Oud and Liina Nagirnaja and Marta Olszewska and Moira K O'Bryan and Donald F Conrad and Maciej Kurpisz and Frank Tüttelmann and Alexander N Yatsenko and },

doi = {10.1007/s00439-021-02287-y},

issn = {1432-1203},

year  = {2021},

date = {2021-08-01},

journal = {Hum Genet},

volume = {140},

number = {8},

pages = {1169--1182},

abstract = {Male infertility impacts millions of couples yet, the etiology of primary infertility remains largely unknown. A critical element of successful spermatogenesis is maintenance of genome integrity. Here, we present a genomic study of spermatogenic failure (SPGF). Our initial analysis (n = 176) did not reveal known gene-candidates but identified a potentially significant single-nucleotide variant (SNV) in X-linked germ-cell nuclear antigen (GCNA). Together with a larger follow-up study (n = 2049), 7 likely clinically relevant GCNA variants were identified. GCNA is critical for genome integrity in male meiosis and knockout models exhibit impaired spermatogenesis and infertility. Single-cell RNA-seq and immunohistochemistry confirm human GCNA expression from spermatogonia to elongated spermatids. Five identified SNVs were located in key functional regions, including N-terminal SUMO-interacting motif and C-terminal Spartan-like protease domain. Notably, variant p.Ala115ProfsTer7 results in an early frameshift, while Spartan-like domain missense variants p.Ser659Trp and p.Arg664Cys change conserved residues, likely affecting 3D structure. For variants within GCNA's intrinsically disordered region, we performed computational modeling for consensus motifs. Two SNVs were predicted to impact the structure of these consensus motifs. All identified variants have an extremely low minor allele frequency in the general population and 6 of 7 were not detected in > 5000 biological fathers. Considering evidence from animal models, germ-cell-specific expression, 3D modeling, and computational predictions for SNVs, we propose that identified GCNA variants disrupt structure and function of the respective protein domains, ultimately arresting germ-cell division. To our knowledge, this is the first study implicating GCNA, a key genome integrity factor, in human male infertility.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid33442887,

title = {The Sertoli cell expressed gene secernin-1 (Scrn1) is dispensable for male fertility in the mouse},

author = {Brendan J Houston and Liina Nagirnaja and D Jo Merriner and Anne E O'Connor and Hidenobu Okuda and Kenan Omurtag and Craig Smith and Kenneth I Aston and Donald F Conrad and Moira K O'Bryan},

doi = {10.1002/dvdy.299},

issn = {1097-0177},

year  = {2021},

date = {2021-07-01},

journal = {Dev Dyn},

volume = {250},

number = {7},

pages = {922--931},

abstract = {BACKGROUND: Male infertility is a prevalent clinical presentation for which there is likely a strong genetic component due to the thousands of genes required for spermatogenesis. Within this study we investigated the role of the gene Scrn1 in male fertility. Scrn1 is preferentially expressed in XY gonads during the period of sex determination and in adult Sertoli cells based on single cell RNA sequencing. We investigated the expression of Scrn1 in juvenile and adult tissues and generated a knockout mouse model to test its role in male fertility.nnRESULTS: Scrn1 was expressed at all ages examined in the post-natal testis; however, its expression peaked at postnatal days 7-14 and SCRN1 protein was clearly localized to Sertoli cells. Scrn1 deletion was achieved via removal of exon 3, and its loss had no effect on male fertility or sex determination. Knockout mice were capable of siring litters of equal size to wild type counterparts and generated equal numbers of sperm with comparable motility and morphology characteristics.nnCONCLUSIONS: Scrn1 was found to be dispensable for male fertility, but this study identifies SCRN1 as a novel marker of the Sertoli cell cytoplasm.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid33864768,

title = {Population-scale tissue transcriptomics maps long non-coding RNAs to complex disease},

author = {Olivia M de Goede and Daniel C Nachun and Nicole M Ferraro and Michael J Gloudemans and Abhiram S Rao and Craig Smail and Tiffany Y Eulalio and François Aguet and Bernard Ng and Jishu Xu and Alvaro N Barbeira and Stephane E Castel and Sarah Kim-Hellmuth and YoSon Park and Alexandra J Scott and Benjamin J Strober and and Christopher D Brown and Xiaoquan Wen and Ira M Hall and Alexis Battle and Tuuli Lappalainen and Hae Kyung Im and Kristin G Ardlie and Sara Mostafavi and Thomas Quertermous and Karla Kirkegaard and Stephen B Montgomery},

doi = {10.1016/j.cell.2021.03.050},

issn = {1097-4172},

year  = {2021},

date = {2021-05-01},

urldate = {2021-05-01},

journal = {Cell},

volume = {184},

number = {10},

pages = {2633--2648.e19},

abstract = {Long non-coding RNA (lncRNA) genes have well-established and important impacts on molecular and cellular functions. However, among the thousands of lncRNA genes, it is still a major challenge to identify the subset with disease or trait relevance. To systematically characterize these lncRNA genes, we used Genotype Tissue Expression (GTEx) project v8 genetic and multi-tissue transcriptomic data to profile the expression, genetic regulation, cellular contexts, and trait associations of 14,100 lncRNA genes across 49 tissues for 101 distinct complex genetic traits. Using these approaches, we identified 1,432 lncRNA gene-trait associations, 800 of which were not explained by stronger effects of neighboring protein-coding genes. This included associations between lncRNA quantitative trait loci and inflammatory bowel disease, type 1 and type 2 diabetes, and coronary artery disease, as well as rare variant associations to body mass index.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid33211200,

title = {Disruption of human meiotic telomere complex genes TERB1, TERB2 and MAJIN in men with non-obstructive azoospermia},

author = {Albert Salas-Huetos and Frank Tüttelmann and Margot J Wyrwoll and Sabine Kliesch and Alexandra M Lopes and João Goncalves and Steven E Boyden and Marius Wöste and James M Hotaling and  and Liina Nagirnaja and Donald F Conrad and Douglas T Carrell and Kenneth I Aston},

doi = {10.1007/s00439-020-02236-1},

issn = {1432-1203},

year  = {2021},

date = {2021-01-01},

journal = {Hum Genet},

volume = {140},

number = {1},

pages = {217--227},

abstract = {Non-obstructive azoospermia (NOA), the lack of spermatozoa in semen due to impaired spermatogenesis affects nearly 1% of men. In about half of cases, an underlying cause for NOA cannot be identified. This study aimed to identify novel variants associated with idiopathic NOA. We identified a nonconsanguineous family in which multiple sons displayed the NOA phenotype. We performed whole-exome sequencing in three affected brothers with NOA, their two unaffected brothers and their father, and identified compound heterozygous frameshift variants (one novel and one extremely rare) in Telomere Repeat Binding Bouquet Formation Protein 2 (TERB2) that segregated perfectly with NOA. TERB2 interacts with TERB1 and Membrane Anchored Junction Protein (MAJIN) to form the tripartite meiotic telomere complex (MTC), which has been shown in mouse models to be necessary for the completion of meiosis and both male and female fertility. Given our novel findings of TERB2 variants in NOA men, along with the integral role of the three MTC proteins in spermatogenesis, we subsequently explored exome sequence data from 1495 NOA men to investigate the role of MTC gene variants in spermatogenic impairment. Remarkably, we identified two NOA patients with likely damaging rare homozygous stop and missense variants in TERB1 and one NOA patient with a rare homozygous missense variant in MAJIN. Available testis histology data from three of the NOA patients indicate germ cell maturation arrest, consistent with mouse phenotypes. These findings suggest that variants in MTC genes may be an important cause of NOA in both consanguineous and outbred populations.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32248361,

title = {A framework for high-resolution phenotyping of candidate male infertility mutants: from human to mouse},

author = {Brendan J Houston and Donald F Conrad and Moira K O'Bryan},

doi = {10.1007/s00439-020-02159-x},

issn = {1432-1203},

year  = {2021},

date = {2021-01-01},

journal = {Hum Genet},

volume = {140},

number = {1},

pages = {155--182},

abstract = {Male infertility is a heterogeneous condition of largely unknown etiology that affects at least 7% of men worldwide. Classical genetic approaches and emerging next-generation sequencing studies support genetic variants as a frequent cause of male infertility. Meanwhile, the barriers to transmission of this disease mean that most individual genetic cases will be rare, but because of the large percentage of the genome required for spermatogenesis, the number of distinct causal mutations is potentially large. Identifying bona fide causes of male infertility thus requires advanced filtering techniques to select for high-probability candidates, including the ability to test causality in animal models. The mouse remains the gold standard for defining the genotype-phenotype connection in male fertility. Here, we present a best practice guide consisting of (a) major points to consider when interpreting next-generation sequencing data performed on infertile men, and, (b) a systematic strategy to categorize infertility types and how they relate to human male infertility. Phenotyping infertility in mice can involve investigating the function of multiple cell types across the testis and epididymis, as well as sperm function. These findings will feed into the diagnosis and treatment of male infertility as well as male health broadly.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32741963,

title = {Genetic dissection of spermatogenic arrest through exome analysis: clinical implications for the management of azoospermic men},

author = {Csilla Krausz and Antoni Riera-Escamilla and Daniel Moreno-Mendoza and Kaylee Holleman and Francesca Cioppi and Ferran Algaba and Marc Pybus and Corinna Friedrich and Margot J Wyrwoll and Elena Casamonti and Sara Pietroforte and Liina Nagirnaja and Alexandra M Lopes and Sabine Kliesch and Adrian Pilatz and Douglas T Carrell and Donald F Conrad and Elisabet Ars and Eduard Ruiz-Castañé and Kenneth I Aston and Willy M Baarends and Frank Tüttelmann},

doi = {10.1038/s41436-020-0907-1},

issn = {1530-0366},

year  = {2020},

date = {2020-12-01},

journal = {Genet Med},

volume = {22},

number = {12},

pages = {1956--1966},

abstract = {PURPOSE: Azoospermia affects 1% of men and it can be the consequence of spermatogenic maturation arrest (MA). Although the etiology of MA is likely to be of genetic origin, only 13 genes have been reported as recurrent potential causes of MA.nnMETHODS: Exome sequencing in 147 selected MA patients (discovery cohort and two validation cohorts).nnRESULTS: We found strong evidence for five novel genes likely responsible for MA (ADAD2, TERB1, SHOC1, MSH4, and RAD21L1), for which mouse knockout (KO) models are concordant with the human phenotype. Four of them were validated in the two independent MA cohorts. In addition, nine patients carried pathogenic variants in seven previously reported genes-TEX14, DMRT1, TEX11, SYCE1, MEIOB, MEI1, and STAG3-allowing to upgrade the clinical significance of these genes for diagnostic purposes. Our meiotic studies provide novel insight into the functional consequences of the variants, supporting their pathogenic role.nnCONCLUSION: Our findings contribute substantially to the development of a pre-testicular sperm extraction (TESE) prognostic gene panel. If properly validated, the genetic diagnosis of complete MA prior to surgical interventions is clinically relevant. Wider implications include the understanding of potential genetic links between nonobstructive azoospermia (NOA) and cancer predisposition, and between NOA and premature ovarian failure.},

keywords = {GEMINI},

pubstate = {published},

tppubtype = {article}

}

@article{pmid33239672,

title = {The genetic architecture of sporadic and multiple consecutive miscarriage},

author = {Triin Laisk and Ana Luiza G Soares and Teresa Ferreira and Jodie N Painter and Jenny C Censin and Samantha Laber and Jonas Bacelis and Chia-Yen Chen and Maarja Lepamets and Kuang Lin and Siyang Liu and Iona Y Millwood and Avinash Ramu and Jennifer Southcombe and Marianne S Andersen and Ling Yang and Christian M Becker and Anders D Børglum and Scott D Gordon and Jonas Bybjerg-Grauholm and Øyvind Helgeland and David M Hougaard and Xin Jin and Stefan Johansson and Julius Juodakis and Christiana Kartsonaki and Viktorija Kukushkina and Penelope A Lind and Andres Metspalu and Grant W Montgomery and Andrew P Morris and Ole Mors and Preben B Mortensen and Pål R Njølstad and Merete Nordentoft and Dale R Nyholt and Margaret Lippincott and Stephanie Seminara and Andres Salumets and Harold Snieder and Krina Zondervan and Thomas Werge and Zhengming Chen and Donald F Conrad and Bo Jacobsson and Liming Li and Nicholas G Martin and Benjamin M Neale and Rasmus Nielsen and Robin G Walters and Ingrid Granne and Sarah E Medland and Reedik Mägi and Deborah A Lawlor and Cecilia M Lindgren},

doi = {10.1038/s41467-020-19742-5},

issn = {2041-1723},

year  = {2020},

date = {2020-11-01},

urldate = {2020-11-01},

journal = {Nat Commun},

volume = {11},

number = {1},

pages = {5980},

abstract = {Miscarriage is a common, complex trait affecting ~15% of clinically confirmed pregnancies. Here we present the results of large-scale genetic association analyses with 69,054 cases from five different ancestries for sporadic miscarriage, 750 cases of European ancestry for multiple (≥3) consecutive miscarriage, and up to 359,469 female controls. We identify one genome-wide significant association (rs146350366, minor allele frequency (MAF) 1.2%, P = 3.2 × 10, odds ratio (OR) = 1.4) for sporadic miscarriage in our European ancestry meta-analysis and three genome-wide significant associations for multiple consecutive miscarriage (rs7859844, MAF = 6.4%, P = 1.3 × 10, OR = 1.7; rs143445068, MAF = 0.8%, P = 5.2 × 10, OR = 3.4; rs183453668, MAF = 0.5%, P = 2.8 × 10, OR = 3.8). We further investigate the genetic architecture of miscarriage with biobank-scale Mendelian randomization, heritability, and genetic correlation analyses. Our results show that miscarriage etiopathogenesis is partly driven by genetic variation potentially related to placental biology, and illustrate the utility of large-scale biobank data for understanding this pregnancy complication.},

keywords = {Genomic Variation, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32913098,

title = {The GTEx Consortium atlas of genetic regulatory effects across human tissues},

author = { },

doi = {10.1126/science.aaz1776},

issn = {1095-9203},

year  = {2020},

date = {2020-09-01},

journal = {Science},

volume = {369},

number = {6509},

pages = {1318--1330},

abstract = {The Genotype-Tissue Expression (GTEx) project was established to characterize genetic effects on the transcriptome across human tissues and to link these regulatory mechanisms to trait and disease associations. Here, we present analyses of the version 8 data, examining 15,201 RNA-sequencing samples from 49 tissues of 838 postmortem donors. We comprehensively characterize genetic associations for gene expression and splicing in cis and trans, showing that regulatory associations are found for almost all genes, and describe the underlying molecular mechanisms and their contribution to allelic heterogeneity and pleiotropy of complex traits. Leveraging the large diversity of tissues, we provide insights into the tissue specificity of genetic effects and show that cell type composition is a key factor in understanding gene regulatory mechanisms in human tissues.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32913075,

title = {Cell type-specific genetic regulation of gene expression across human tissues},

author = {Sarah Kim-Hellmuth and François Aguet and Meritxell Oliva and Manuel Muñoz-Aguirre and Silva Kasela and Valentin Wucher and Stephane E Castel and Andrew R Hamel and Ana Viñuela and Amy L Roberts and Serghei Mangul and Xiaoquan Wen and Gao Wang and Alvaro N Barbeira and Diego Garrido-Martín and Brian B Nadel and Yuxin Zou and Rodrigo Bonazzola and Jie Quan and Andrew Brown and Angel Martinez-Perez and José Manuel Soria and  and Gad Getz and Emmanouil T Dermitzakis and Kerrin S Small and Matthew Stephens and Hualin S Xi and Hae Kyung Im and Roderic Guigó and Ayellet V Segrè and Barbara E Stranger and Kristin G Ardlie and Tuuli Lappalainen},

doi = {10.1126/science.aaz8528},

issn = {1095-9203},

year  = {2020},

date = {2020-09-01},

journal = {Science},

volume = {369},

number = {6509},

abstract = {The Genotype-Tissue Expression (GTEx) project has identified expression and splicing quantitative trait loci in cis (QTLs) for the majority of genes across a wide range of human tissues. However, the functional characterization of these QTLs has been limited by the heterogeneous cellular composition of GTEx tissue samples. We mapped interactions between computational estimates of cell type abundance and genotype to identify cell type-interaction QTLs for seven cell types and show that cell type-interaction expression QTLs (eQTLs) provide finer resolution to tissue specificity than bulk tissue cis-eQTLs. Analyses of genetic associations with 87 complex traits show a contribution from cell type-interaction QTLs and enables the discovery of hundreds of previously unidentified colocalized loci that are masked in bulk tissue.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32913074,

title = {Determinants of telomere length across human tissues},

author = {Kathryn Demanelis and Farzana Jasmine and Lin S Chen and Meytal Chernoff and Lin Tong and Dayana Delgado and Chenan Zhang and Justin Shinkle and Mekala Sabarinathan and Hannah Lin and Eduardo Ramirez and Meritxell Oliva and Sarah Kim-Hellmuth and Barbara E Stranger and Tsung-Po Lai and Abraham Aviv and Kristin G Ardlie and François Aguet and Habibul Ahsan and  and Jennifer A Doherty and Muhammad G Kibriya and Brandon L Pierce},

doi = {10.1126/science.aaz6876},

issn = {1095-9203},

year  = {2020},

date = {2020-09-01},

journal = {Science},

volume = {369},

number = {6509},

abstract = {Telomere shortening is a hallmark of aging. Telomere length (TL) in blood cells has been studied extensively as a biomarker of human aging and disease; however, little is known regarding variability in TL in nonblood, disease-relevant tissue types. Here, we characterize variability in TLs from 6391 tissue samples, representing >20 tissue types and 952 individuals from the Genotype-Tissue Expression (GTEx) project. We describe differences across tissue types, positive correlation among tissue types, and associations with age and ancestry. We show that genetic variation affects TL in multiple tissue types and that TL may mediate the effect of age on gene expression. Our results provide the foundational knowledge regarding TL in healthy tissues that is needed to interpret epidemiological studies of TL and human health.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32913073,

title = {Transcriptomic signatures across human tissues identify functional rare genetic variation},

author = {Nicole M Ferraro and Benjamin J Strober and Jonah Einson and Nathan S Abell and Francois Aguet and Alvaro N Barbeira and Margot Brandt and Maja Bucan and Stephane E Castel and Joe R Davis and Emily Greenwald and Gaelen T Hess and Austin T Hilliard and Rachel L Kember and Bence Kotis and YoSon Park and Gina Peloso and Shweta Ramdas and Alexandra J Scott and Craig Smail and Emily K Tsang and Seyedeh M Zekavat and Marcello Ziosi and Aradhana and  and Kristin G Ardlie and Themistocles L Assimes and Michael C Bassik and Christopher D Brown and Adolfo Correa and Ira Hall and Hae Kyung Im and Xin Li and Pradeep Natarajan and  and Tuuli Lappalainen and Pejman Mohammadi and Stephen B Montgomery and Alexis Battle},

doi = {10.1126/science.aaz5900},

issn = {1095-9203},

year  = {2020},

date = {2020-09-01},

journal = {Science},

volume = {369},

number = {6509},

abstract = {Rare genetic variants are abundant across the human genome, and identifying their function and phenotypic impact is a major challenge. Measuring aberrant gene expression has aided in identifying functional, large-effect rare variants (RVs). Here, we expanded detection of genetically driven transcriptome abnormalities by analyzing gene expression, allele-specific expression, and alternative splicing from multitissue RNA-sequencing data, and demonstrate that each signal informs unique classes of RVs. We developed Watershed, a probabilistic model that integrates multiple genomic and transcriptomic signals to predict variant function, validated these predictions in additional cohorts and through experimental assays, and used them to assess RVs in the UK Biobank, the Million Veterans Program, and the Jackson Heart Study. Our results link thousands of RVs to diverse molecular effects and provide evidence to associate RVs affecting the transcriptome with human traits.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}

@article{pmid32913072,

title = {The impact of sex on gene expression across human tissues},

author = {Meritxell Oliva and Manuel Muñoz-Aguirre and Sarah Kim-Hellmuth and Valentin Wucher and Ariel D H Gewirtz and Daniel J Cotter and Princy Parsana and Silva Kasela and Brunilda Balliu and Ana Viñuela and Stephane E Castel and Pejman Mohammadi and François Aguet and Yuxin Zou and Ekaterina A Khramtsova and Andrew D Skol and Diego Garrido-Martín and Ferran Reverter and Andrew Brown and Patrick Evans and Eric R Gamazon and Anthony Payne and Rodrigo Bonazzola and Alvaro N Barbeira and Andrew R Hamel and Angel Martinez-Perez and José Manuel Soria and  and Brandon L Pierce and Matthew Stephens and Eleazar Eskin and Emmanouil T Dermitzakis and Ayellet V Segrè and Hae Kyung Im and Barbara E Engelhardt and Kristin G Ardlie and Stephen B Montgomery and Alexis J Battle and Tuuli Lappalainen and Roderic Guigó and Barbara E Stranger},

doi = {10.1126/science.aba3066},

issn = {1095-9203},

year  = {2020},

date = {2020-09-01},

journal = {Science},

volume = {369},

number = {6509},

abstract = {Many complex human phenotypes exhibit sex-differentiated characteristics. However, the molecular mechanisms underlying these differences remain largely unknown. We generated a catalog of sex differences in gene expression and in the genetic regulation of gene expression across 44 human tissue sources surveyed by the Genotype-Tissue Expression project (GTEx, v8 release). We demonstrate that sex influences gene expression levels and cellular composition of tissue samples across the human body. A total of 37% of all genes exhibit sex-biased expression in at least one tissue. We identify cis expression quantitative trait loci (eQTLs) with sex-differentiated effects and characterize their cellular origin. By integrating sex-biased eQTLs with genome-wide association study data, we identify 58 gene-trait associations that are driven by genetic regulation of gene expression in a single sex. These findings provide an extensive characterization of sex differences in the human transcriptome and its genetic regulation.},

keywords = {Genomic Variation},

pubstate = {published},

tppubtype = {article}

}