PUBLICATIONS

@article{pmid41664592,

title = {Mapping whole-organism genetic comorbidities across model Species using unified ontologies},

author = {Caitlin Peaslee and Madison Held and Eloise Fadial and Donald F Conrad},

doi = {10.1093/genetics/iyag038},

issn = {1943-2631},

year  = {2026},

date = {2026-04-01},

urldate = {2026-04-01},

journal = {Genetics},

volume = {232},

number = {4},

abstract = {Understanding how genetic variation contributes to organism-wide phenotypes is critical for identifying mechanisms of disease. Here, we present a computational approach to analyze whole-organism comorbidities associated with genes underlying non-obstructive azoospermia (NOA), the most severe form of male infertility. We curated 204 mouse genes with experimentally validated spermatogenic failure and mapped their orthologs and phenotype annotations across humans, M. musculus, D. rerio, D. melanogaster, and C. elegans using a unified cross-species phenotype structure. This framework integrates a newly developed reproductive phenotype ontology with standardized whole-body phenotype categories to enable direct cross-species comparisons, using thousands of genotype-phenotype associations stored in model organism databases. Our analysis shows that most NOA genes have conserved orthologs across models, and that perturbation of these genes is frequently associated with non-reproductive phenotypes. In particular, integumentary defects and neoplastic phenotypes recur across species, supporting a shared genetic basis for epidemiological links between male infertility and systemic disease. Gene-level comorbidity patterns are partially predictable from single-cell RNA-seq, whole-body gene expression, and Gene Ontology annotations, suggesting that fundamental biological constraints shape the systemic consequences of reproductive gene dysfunction. Clustering genes by comorbidity profiles further distinguished genes with isolated reproductive effects from those with broad organismal consequences. This work demonstrates the power of ontology-based cross-species analysis for identifying pleiotropic effects of Mendelian disease mutations, and provides a resource, CoMorbidity DataBase Mapper (CoMo DBM), for joint analysis of genotype-phenotype associations in model organism databases.},

keywords = {GEMINI, Genomic Variation, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40815353,

title = {Identification of missense DMC1 variants in males with non-obstructive azoospermia},

author = {Noor Ullah and Christopher Pombar and Rachel Hvasta-Gloria and Andrea J Berman and Michelle Malizio and Muhammad Jaseem Khan and Rubina Nazli and Sadia Fatima and Antoni Riera-Escamilla and Helen Castillo-Madeen and Agnieszka Malcher and Marta Olszewska and Miguel J Xavier and Kristiina Lillepea and Avirup Dutta and Carlos A Castro and Anu Valkna and Rain Inno and Kyle E Orwig and Donald F Conrad and Maciej Kurpisz and Joris A Veltman and Maris Laan and Alexander N Yatsenko},

doi = {10.1007/s10815-025-03591-6},

issn = {1573-7330},

year  = {2025},

date = {2025-11-01},

urldate = {2025-11-01},

journal = {J Assist Reprod Genet},

volume = {42},

number = {11},

pages = {3723--3735},

abstract = {PURPOSE: Human male infertility is a significant reproductive condition, with non-obstructive azoospermia (NOA) being the most severe form, resulting from impaired spermatogenesis. Many genetic variants have been identified as negatively impacting sperm development and maturation at multiple stages, leading to spermatogenic failure (SPGF). Here, we aim to study such variants, particularly those in the critical, highly conserved, meiosis-specific DMC1 (DNA meiotic recombinase 1) gene, to identify genetic candidates for male infertility and to strengthen DMC1's existing genotype-phenotype relationships.nnMETHODS: We used whole exome sequencing (WES) and in silico analysis to investigate select DMC1 variants in a large cohort of infertile sporadic and familial cases (n = 3150).nnRESULTS: Our familial analyses identified a homozygous DMC1 missense variant, p.Thr55Ile, in two NOA-affected male siblings. We also report additional homozygous missense variants, p.Thr164Ala and p.Tyr194Cys, and one notable, rare single heterozygous variant, p.Asp160Gly, in unrelated sporadic patients. Our 3D protein modeling indicates that each of our identified variants would significantly impact the structure and functional activity of DMC1 protein.nnCONCLUSION: Our extensive genomic study identified three rare, recessive DMC1 variants in human NOA patients. Further, we report an alternative maturation arrest phenotype than previously observed in DMC1-related NOA. We also provide preliminary support for the possible exploration of select single heterozygous variants in the DMC1 gene, potentially expanding the male infertility field's understanding of the disease states and inheritance patterns associated with variants in DMC1.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40846510,

title = {Genetic and epigenetic landscape of male infertility},

author = {Emma R James and Maliha Tasnim and Antoni Riera-Escamilla and Katinka Vigh-Conrad and Benjamin R Emery and Donald F Conrad and Kenneth I Aston},

doi = {10.1016/j.tig.2025.07.007},

issn = {0168-9525},

year  = {2025},

date = {2025-10-01},

urldate = {2025-10-01},

journal = {Trends Genet},

volume = {41},

number = {10},

pages = {883--898},

abstract = {Male infertility is a global health problem, affecting up to 6% of reproductive age men worldwide. It has an enormous personal and societal burden, along with public health implications beyond the inability to reproduce, including reduced future health and longevity. While the impact of infertility has long been appreciated, the molecular architecture of the disease is largely unknown. Nevertheless, the past decade has witnessed significant advances in our understanding of the molecular basis of male infertility. Here, we describe the contributions of genetic and epigenetic mechanisms to infertility-associated phenotypes and their impact beyond reproduction. This review focuses on progress in understanding defects in sperm production and function, and the potential impact of these advances on diagnosis, treatments, and improved health.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40545437,

title = {The rising role of genetics in andrology research and clinical practice},

author = {Maris Laan and Kenneth I Aston and Donald F Conrad},

doi = {10.1111/andr.70073},

issn = {2047-2927},

year  = {2025},

date = {2025-07-01},

urldate = {2025-07-01},

journal = {Andrology},

volume = {13},

number = {5},

pages = {983--985},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40598224,

title = {A novel mechanism of sperm midpiece epididymal maturation and the role of CCDC112 in sperm midpiece formation and establishing an optimal flagella waveform},

author = {Maddison L Graffeo and Joseph Nguyen and Farin Yazdan Parast and Jessica E M Dunleavy and Denis Korneev and Hongyi Yang and Hidenobu Okuda and Anne E O'Connor and Donald F Conrad and Reza Nosrati and Brendan J Houston and Moira K O'Bryan},

doi = {10.1186/s12964-025-02320-x},

issn = {1478-811X},

year  = {2025},

date = {2025-07-01},

urldate = {2025-07-01},

journal = {Cell Commun Signal},

volume = {23},

number = {1},

pages = {319},

abstract = {BACKGROUND: The mitochondrial sheath is a defining feature of mammalian sperm with proposed functions in structural support and energy production for flagella movement. Recently, coiled coil domain containing (CCDC) protein 112 (CCDC112) was suggested to play a role in the regulation of ciliogenesis. CCDC112 is a poorly characterised protein and there is virtually no knowledge of its in vivo function.nnMETHODS: Here, we define CCDC112 as crucial for male fertility using a Ccdc112 loss-of-function mouse line. To characterize and analyze male fertility, and to identify a novel process of epididymal midpiece maturation, we utilized a range of assays including fertility testing, scanning electron microscopy, high-resolution sperm motility and power output analysis, in vitro fertilization, intracytoplasmic sperm injection, mitochondria stress test assays and glycolytic flux assays. Localization of CCDC112 in cilia was assessed via the transfection of IMCD-3 cells with a CCDC112-eGFP vector and subsequent immunofluorescent staining.nnRESULTS: Results reveal CCDC112 as a requirement for male fertility in the mouse with an essential role in mitochondrial sheath formation. Our data reveal the critical role of CCDC112 in mitochondrial morphogenesis during midpiece formation, with the lack of CCDC112 leading to significantly reduced respiration capacity, irregular flagellar waveforms, diminished progressive motility and ultimately male sterility. In the absence of CCDC112, sperm are unable to traverse the female reproductive tract to the site of fertilization and in vitro have a poor capacity to penetrate the zonae pellucidae of oocytes or fuse with the oocyte. We further unveil a previously unrecognized process of epididymal mitochondrial sheath maturation. We show the sperm midpiece is structurally immature upon exiting the testis and maturation continues during transit from the caput to the cauda epididymis. Finally, we identify CCDC112 as a component of the distal appendages of the mother centriole in IMCD-3 cells suggestive of a facilitative role for CCDC112 in protein entry into the ciliary compartment within germ cells.nnCONCLUSION: Collectively, we establish CCDC112 as a key regulator of sperm midpiece assembly and function while further expanding our understanding on functional sperm production, energy generation and flagella kinematics.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40591933,

title = {CCDC89 is required for optimal sperm motility and male fertility in mammals},

author = {Moira K O'Bryan and Gülizar Saritas and Joseph Nguyen and Sofia B Winge and Anne E O'Connor and Helen Castillo-Madeen and Donald F Conrad and Maddison Graffeo and Reza Nosrati and Jessica E M Dunleavy and Kristian Almstrup and Brendan J Houston},

doi = {10.1093/humrep/deaf126},

issn = {1460-2350},

year  = {2025},

date = {2025-07-01},

urldate = {2025-07-01},

journal = {Hum Reprod},

abstract = {STUDY QUESTION: What is the role of coiled-coil domain-containing protein 89 (CCDC89) in mammalian male fertility?nnSUMMARY ANSWER: The presence of CCDC89 is required for normal sperm motility and therefore optimal male fertility in mice, while CCDC89 variants affected spermatogenesis in both mice and humans.nnWHAT IS KNOWN ALREADY: Coiled-coiled domain-containing proteins play a variety of roles in biological processes, including cell division, the production of motile sperm, and the regulation of their motility.nnSTUDY DESIGN, SIZE, DURATION: DNA from infertile men with azoospermia was sequenced to identify genetic variants as per the Genetics of Male Infertility Initiative (GEMINI) study. Genetic variants were identified in CCDC89 in three men, by whole exome sequencing. A testis biopsy from infertile Patient 1 (CCDC89 variant c. G903T) was available and used to inspect tissue pathology. Ccdc89 knockout (Ccdc89-/-) and Ccdc89E297D/E297D mutant mouse models were generated to define the role of CCDC89 in male fertility and the role of the specific CCDC89 genetic variant, c. G903T, in the pathogenesis of infertility.nnPARTICIPANTS/MATERIALS, SETTING, METHODS: CCDC89 RNA expression and protein localization were investigated in a testis biopsy from a control male with normal spermatogenesis. Male fertility of the mutant mouse lines was assessed via breeding, histology, daily sperm production, electron microscopy, computer-assisted and high-speed sperm motility analysis, and in vitro fertilization.nnMAIN RESULTS AND THE ROLE OF CHANCE: Ccdc89-/- male mice were sub-fertile, with impaired progressive sperm motility and curvilinear velocity due to a rigid sperm tail midpiece without any overt structural defects. While Ccdc89E297D/E297D males were fertile, their testis weights and germ cell content were reduced, suggesting a potential role of the c. G903T variant, observed in each of the two men, in the pathogenesis of their spermatogenic impairment. We also identified a new genetic variant in CCDC89 (c.G1024A) in another infertile man, that was in trans with the c. G903T genetic variant.nnLARGE SCALE DATA: N/A.nnLIMITATIONS, REASONS FOR CAUTION: The identification of additional infertile men with genetic variants in CCDC89, and quality clinical data, are required to determine prognostic reliability regarding CCDC89 variants. There are likely to be species-specific differences in gene function.nnWIDER IMPLICATIONS OF THE FINDINGS: Our data highlight a role for CCDC89, in regulating the sperm tail waveform, that is required for optimal sperm fertilization capacity and male fertility. We highlight CCDC89 as a regulator of male fertility in mammals, where variants in CCDC89 can affect spermatogenesis and may be a risk factor for human male infertility. This study underscores the importance of validating clinical genetic findings.nnSTUDY FUNDING/COMPETING INTEREST(S): This work was supported by a National Health and Medical Research Council grant (APP1120356), National Institutes of Health grants (R01HD078641 and P50HD096723), the Novo Nordisk Foundation (grant numbers NNF210C0069913 and NNF21C0069969), the Capital Region of Denmark, the Independent Research Fund (grant number: 1030-00381B), and the Svend Andersen Foundation (grant number: 84-A.08), and the Christian and Ottilia Brorsons (No. 12038-1) and Frimodt-Heineke Foundation travel grants for research exchange. The authors have no conflicts of interest.},

keywords = {GEMINI, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid40174856,

title = {Genetic and genomic insights into male reproductive tract development},

author = {Madison Held and Helen Castillo-Madeen and Katinka A Vigh-Conrad and Kenneth I Aston and Donald F Conrad},

doi = {10.1016/j.fertnstert.2025.03.024},

issn = {1556-5653},

year  = {2025},

date = {2025-03-01},

urldate = {2025-03-01},

journal = {Fertil Steril},

abstract = {Genetic and genomic analysis continues to drive important insights into male reproductive tract (MRT) development. Here, we briefly review normal MRT development, highlighting recent discoveries of cell types and cellular processes delivered by single-cell sequencing. We report a systematic review of phenotype terms and genes linked to MRT development, identifying 35 terms from the Human Phenotype Ontology associated with 269 unique genes. A parallel review of mouse data revealed differences in the phenotype terms available and the number and identity of genes linked to MRT defects, indicating opportunities for harmonization of knowledge. We used a published single-cell atlas of the developing testis to characterize the regulation of MRT genes across cell types and stages of fetal testis development. Single-cell RNA sequencing data support the conclusion that Leydig cells and Sertoli cells are the primary testicular cell types expressing MRT genes. Furthermore, we find post-conception weeks 6, 8, and 16 to be the key points of upregulation of testicular MRT genes. New advances, especially in imaging and spatially resolved molecular measurements, provide exciting prospects for MRT research and diagnosis, and we expect rapid progress in the coming years. Continued investigation in this space is essential to understand the genetic basis of MRT development and how MRT defects are related to medical outcomes in adult life.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid39961022,

title = {SATINN v2: automated image analysis for mouse testis histology with multi-laboratory data integration},

author = {Ran Yang and Fritzie T Celino-Brady and Jessica E M Dunleavy and Katinka A Vigh-Conrad and Georgia Rae Atkins and Rachel L Hvasta and Christopher R X Pombar and Alexander N Yatsenko and Kyle E Orwig and Moira K O'Bryan and Ana C Lima and Donald F Conrad},

doi = {10.1093/biolre/ioaf033},

issn = {1529-7268},

year  = {2025},

date = {2025-02-01},

urldate = {2025-02-01},

journal = {Biol Reprod},

abstract = {Analysis of testis histology is fundamental to the study of male fertility, but it is a slow task with a high skill threshold. Here, we describe new neural network models for the automated classification of cell types and tubule stages from whole-slide brightfield images of mouse testis. The cell type classifier recognizes 14 cell types, including multiple steps of meiosis I prophase, with an external validation accuracy of 96%. The tubule stage classifier distinguishes all 12 canonical tubule stages with external validation accuracy of 63%, which increases to 96% when allowing for ±1 stage tolerance. We addressed generalizability of SATINN, through extensive training diversification and testing on external (non-training population) wildtype and mutant datasets. This allowed us to use SATINN to successfully process data generated in multiple laboratories. We used SATINN to analyze testis images from 8 different mutant lines, generated from 3 different labs with a range of tissue processing protocols. Finally, we show that it is possible to use SATINN output to cluster histology images in latent space, which, when applied to the 8 mutant lines, reveals known relationships in their pathology. This work represents significant progress towards a tool for robust, automated testis histopathology that can be used by multiple labs.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid39903672,

title = {Deletion of an evolutionarily conserved TAD boundary impacts spermatogenesis in mice†},

author = {Ana C Lima and Mariam Okhovat and Alexandra M Stendahl and Ran Yang and Jake VanCampen and Kimberly A Nevonen and Jarod Herrera and Weiyu Li and Lana Harshman and Lev M Fedorov and Katinka A Vigh-Conrad and Nadav Ahituv and Donald F Conrad and Lucia Carbone},

doi = {10.1093/biolre/ioaf017},

issn = {1529-7268},

year  = {2025},

date = {2025-02-01},

urldate = {2025-02-01},

journal = {Biol Reprod},

abstract = {Spermatogenesis is a complex process that can be disrupted by genetic and epigenetic changes, potentially leading to male infertility. Recent research has rapidly increased the number of protein coding mutations causally linked to impaired spermatogenesis in humans and mice. However, the role of non-coding mutations remains largely unexplored. As a case study to evaluate the effects of non-coding mutations on spermatogenesis, we first identified an evolutionarily conserved topologically associated domain (TAD) boundary near two genes with important roles in mammalian testis function: Dmrtb1 and Lrp8. We then used CRISPR-Cas9 to generate a mouse line where 26 kb of the boundary was removed including a strong and evolutionarily conserved CTCF binding site. ChIP-seq and Hi-C experiments confirmed the removal of the CTCF site and a resulting mild increase in the DNA-DNA interactions across the domain boundary. Mutant mice displayed significant changes in testis gene expression, higher frequency of histological abnormalities, a drop of 47-52% in efficiency of meiosis, a 15-18% reduction in efficiency of spermatogenesis, and consistently, a 12-28% decrease in daily sperm production compared to littermate controls. Despite these quantitative changes in testis function, mutant mice show no significant changes in fertility. This suggests that non-coding deletions affecting testis gene regulation may have smaller effects on fertility compared to coding mutations of the same genes. Our results demonstrate that disruption of a TAD boundary can have a negative impact on sperm production and highlight the importance of considering non-coding mutations in the analysis of patients with male infertility.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid39780503,

title = {Introduction to androgenetics: terminology, approaches, and impactful studies across 60 years},

author = {Arvand Akbari and Laura Kasak and Maris Laan},

doi = {10.1111/andr.13835},

issn = {2047-2927},

year  = {2025},

date = {2025-01-01},

urldate = {2025-01-01},

journal = {Andrology},

abstract = {Across six decades, androgenetics has consistently concentrated on discovering genetic causes and enhancing the molecular diagnostics of male infertility, disorders of sex development, and their broader implications on health, such as cancer and other comorbidities. Despite vast clinical knowledge, the training of andrologists often lacks fundamental basics in medical genetics. This work, as part of the Special Issue of Andrology "Genetics in Andrology", provides the core terminology in medical genetics and technological advancements in genomics, required to understand the ever-progressing research in the field. It also gives an overview of study designs and approaches that have frequently led to discoveries in androgenetics. The rapid progress in the methodological toolbox in human genetics is illustrated by numerous examples of impactful androgenetic studies over 60 years, and their clinical implications.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid39300832,

title = {Utility of exome sequencing in primary spermatogenic disorders: From research to diagnostics},

author = {Antoni Riera-Escamilla and Liina Nagirnaja},

doi = {10.1111/andr.13753},

issn = {2047-2927},

year  = {2024},

date = {2024-09-01},

urldate = {2024-09-01},

journal = {Andrology},

abstract = {BACKGROUND: Primary spermatogenic disorders represent a severe form of male infertility whereby sperm production is impaired due to testicular dysfunction, leading to reduced quality or quantity of spermatozoa. Gene-centered research has certainly demonstrated the importance of the genetic factor in the etiology of both poor sperm morphology or motility and reduced sperm count. In the last decade, next-generation sequencing has expanded the research to whole exome which has transformed our understanding of male infertility genetics, but uncertainty persists in its diagnostic yield, especially in large unrelated populations.nnOBJECTIVE: To evaluate the utility of exome sequencing in detecting genetic factors contributing to various traits of primary spermatogenic disorders, which is a crucial step before interpreting the diagnostic yield of the platform.nnMATERIALS AND METHODS: We manually curated 415 manuscripts and included 19 research studies that predominantly performed whole exome sequencing in cohorts of unrelated cases with primary spermatogenic defects.nnRESULTS: The detection rate, defined as the fraction of cases with an identifiable genetic cause, typically remained below 25% for quantitative defects of spermatozoa, whereas improved rates were observed for traits of abnormal sperm morphology/motility and in populations enriched with consanguineous families. Unlike the quantitative defects, the genetic architecture of the qualitative issues of spermatozoa featured a small number of recurrent genes describing a large fraction of studied cases. These observations were also in line with the lower biological complexity of the pathways affected by the reported genes.nnDISCUSSION AND CONCLUSIONS: This review demonstrates the variability in detection rates of exome sequencing across semen phenotypes, which may have an impact on the expectations of the diagnostic yield in the clinical setting.},

keywords = {Human Reproduction},

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{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{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{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{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{pmid31604923,

title = {Rare mutations in the complement regulatory gene CSMD1 are associated with male and female infertility},

author = {Arthur S Lee and Jannette Rusch and Ana C Lima and Abul Usmani and Ni Huang and Maarja Lepamets and Katinka A Vigh-Conrad and Ronald E Worthington and Reedik Mägi and Xiaobo Wu and Kenneth I Aston and John P Atkinson and Douglas T Carrell and Rex A Hess and Moira K O'Bryan and Donald F Conrad},

doi = {10.1038/s41467-019-12522-w},

issn = {2041-1723},

year  = {2019},

date = {2019-10-01},

journal = {Nat Commun},

volume = {10},

number = {1},

pages = {4626},

abstract = {Infertility in men and women is a complex genetic trait with shared biological bases between the sexes. Here, we perform a series of rare variant analyses across 73,185 women and men to identify genes that contribute to primary gonadal dysfunction. We report CSMD1, a complement regulatory protein on chromosome 8p23, as a strong candidate locus in both sexes. We show that CSMD1 is enriched at the germ-cell/somatic-cell interface in both male and female gonads. Csmd1-knockout males show increased rates of infertility with significantly increased complement C3 protein deposition in the testes, accompanied by severe histological degeneration. Knockout females show significant reduction in ovarian quality and breeding success, as well as mammary branching impairment. Double knockout of Csmd1 and C3 causes non-additive reduction in breeding success, suggesting that CSMD1 and the complement pathway play an important role in the normal postnatal development of the gonads in both sexes.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid31237565,

title = {Unified single-cell analysis of testis gene regulation and pathology in five mouse strains},

author = {Min Jung and Daniel Wells and Jannette Rusch and Suhaira Ahmad and Jonathan Marchini and Simon R Myers and Donald F Conrad},

doi = {10.7554/eLife.43966},

issn = {2050-084X},

year  = {2019},

date = {2019-06-01},

journal = {Elife},

volume = {8},

abstract = {To fully exploit the potential of single-cell functional genomics in the study of development and disease, robust methods are needed to simplify the analysis of data across samples, time-points and individuals. Here we introduce a model-based factor analysis method, SDA, to analyze a novel 57,600 cell dataset from the testes of wild-type mice and mice with gonadal defects due to disruption of the genes , ,  or . By jointly analyzing mutant and wild-type cells we decomposed our data into 46 components that identify novel meiotic gene-regulatory programs, mutant-specific pathological processes, and technical effects, and provide a framework for imputation. We identify, de novo, DNA sequence motifs associated with individual components that define temporally varying modes of gene expression control. Analysis of SDA components also led us to identify a rare population of macrophages within the seminiferous tubules of  and  mice, an area typically associated with immune privilege.},

keywords = {Human Reproduction, Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid29307395,

title = {Genetic intersection of male infertility and cancer},

author = {Liina Nagirnaja and Kenneth I Aston and Donald F Conrad},

doi = {10.1016/j.fertnstert.2017.10.028},

issn = {1556-5653},

year  = {2018},

date = {2018-01-01},

urldate = {2018-01-01},

journal = {Fertil Steril},

volume = {109},

number = {1},

pages = {20--26},

abstract = {Recent epidemiological studies have identified an association between male factor infertility and increased cancer risk, however, the underlying etiology for the shared risk has not been investigated. It is likely that much of the association between the two disease states can be attributed to underlying genetic lesions. In this article we review the reported associations between cancer and spermatogenic defects, and through database searches we identify candidate genes and gene classes that could explain some of the observed shared genetic risk. We discuss the importance of fully characterizing the genetic basis for the relationship between cancer and male factor infertility and propose future studies to that end.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid28745623,

title = {A Standardized Approach for Multispecies Purification of Mammalian Male Germ Cells by Mechanical Tissue Dissociation and Flow Cytometry},

author = {Ana C Lima and Min Jung and Jannette Rusch and Abul Usmani and Alexandra M Lopes and Donald F Conrad},

doi = {10.3791/55913},

issn = {1940-087X},

year  = {2017},

date = {2017-07-01},

urldate = {2017-07-01},

journal = {J Vis Exp},

number = {125},

abstract = {Fluorescence-activated cell sorting (FACS) has been one of the methods of choice to isolate enriched populations of mammalian testicular germ cells. Currently, it allows the discrimination of up to 9 murine germ cell populations with high yield and purity. This high-resolution in discrimination and purification is possible due to unique changes in chromatin structure and quantity throughout spermatogenesis. These patterns can be captured by flow cytometry of male germ cells stained with fluorescent DNA-binding dyes such as Hoechst-33342 (Hoechst). Herein is a detailed description of a recently developed protocol to isolate mammalian testicular germ cells. Briefly, single cell suspensions are generated from testicular tissue by mechanical dissociation, double stained with Hoechst and propidium iodide (PI) and processed by flow cytometry. A serial gating strategy, including the selection of live cells (PI negative) with different DNA content (Hoechst intensity), is used during FACS sorting to discriminate up to 5 germ cell types. These include, with corresponding average purities (determined by microscopy evaluation): spermatogonia (66%), primary (71%) and secondary (85%) spermatocytes, and spermatids (90%), further separated into round (93%) and elongating (87%) subpopulations. Execution of the entire workflow is straightforward, allows the isolation of 4 cell types simultaneously with the appropriate FACS machine, and can be performed in less than 2 h. As reduced processing time is crucial to preserve the physiology of ex vivo cells, this method is ideal for downstream high-throughput studies of male germ cell biology. Moreover, a standardized protocol for multispecies purification of mammalian germ cells eliminates methodological sources of variables and allows a single set of reagents to be used for different animal models.},

keywords = {Human Reproduction, Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid28859288,

title = {The long and short of translational control in male germ cells},

author = {Ana C Lima and Donald F Conrad},

doi = {10.1093/biolre/iox075},

issn = {1529-7268},

year  = {2017},

date = {2017-07-01},

urldate = {2017-07-01},

journal = {Biol Reprod},

volume = {97},

number = {1},

pages = {2--4},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid28658705,

title = {How to Map the Genetic Basis for Conditions that are Comorbid with Male Infertility},

author = {Liina Nagirnaja and Katinka Vigh-Conrad and Donald F Conrad},

doi = {10.1055/s-0037-1603567},

issn = {1526-4564},

year  = {2017},

date = {2017-05-01},

urldate = {2017-05-01},

journal = {Semin Reprod Med},

volume = {35},

number = {3},

pages = {225--230},

keywords = {Genomic Variation, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid27856695,

title = {Multiplex shRNA Screening of Germ Cell Development by in Vivo Transfection of Mouse Testis},

author = {Nicholas R Y Ho and Abul R Usmani and Yan Yin and Liang Ma and Donald F Conrad},

doi = {10.1534/g3.116.036087},

issn = {2160-1836},

year  = {2017},

date = {2017-01-01},

urldate = {2017-01-01},

journal = {G3 (Bethesda)},

volume = {7},

number = {1},

pages = {247--255},

abstract = {Spermatozoa are one of the few mammalian cell types that cannot be fully derived in vitro, severely limiting the application of modern genomic techniques to study germ cell biology. The current gold standard approach of characterizing single-gene knockout mice is slow as generation of each mutant line can take 6-9 months. Here, we describe an in vivo approach to rapid functional screening of germline genes based on a new nonsurgical, nonviral in vivo transfection method to deliver nucleic acids into testicular germ cells. By coupling multiplex transfection of short hairpin RNA (shRNA) constructs with pooled amplicon sequencing as a readout, we were able to screen many genes for spermatogenesis function in a quick and inexpensive experiment. We transfected nine mouse testes with a pilot pool of RNA interference (RNAi) against well-characterized genes to show that this system is highly reproducible and accurate. With a false negative rate of 18% and a false positive rate of 12%, this method has similar performance as other RNAi screens in the well-described Drosophila model system. In a separate experiment, we screened 26 uncharacterized genes computationally predicted to be essential for spermatogenesis and found numerous candidates for follow-up studies. Finally, as a control experiment, we performed a long-term selection screen in neuronal N2a cells, sampling shRNA frequencies at five sequential time points. By characterizing the effect of both libraries on N2a cells, we show that our screening results from testis are tissue-specific. Our calculations indicate that the current implementation of this approach could be used to screen thousands of protein-coding genes simultaneously in a single mouse testis. The experimental protocols and analysis scripts provided will enable other groups to use this procedure to study diverse aspects of germ cell biology ranging from epigenetics to cell physiology. This approach also has great promise as an applied tool for validating diagnoses made from medical genome sequencing, or designing synthetic biological sequences that can act as potent and highly specific male contraceptives.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid27557646,

title = {Multispecies Purification of Testicular Germ Cells},

author = {Ana C Lima and Min Jung and Jannette Rusch and Abul Usmani and Alexandra Lopes and Donald F Conrad},

doi = {10.1095/biolreprod.116.140566},

issn = {1529-7268},

year  = {2016},

date = {2016-10-01},

urldate = {2016-10-01},

journal = {Biol Reprod},

volume = {95},

number = {4},

pages = {85},

abstract = {Advanced methods of cellular purification are required to apply genome technology to the study of spermatogenesis. One approach, based on flow cytometry of murine testicular cells stained with Hoechst-33342 (Ho-FACS), has been extensively optimized and currently allows the isolation of 9 germ cell types. This staining technique is straightforward to implement, highly effective at purifying specific germ cell types and yields sufficient cell numbers for high throughput studies. Ho-FACS is a technique that does not require species-specific markers, but whose applicability to other species is largely unexplored. We hypothesized that, due to the similar cell physiology of spermatogenesis across mammals, Ho-FACS could be used to produce highly purified subpopulations of germ cells in mammals other than mouse. To test this hypothesis, we applied Ho-FACS to 4 mammalian species that are widely used in testis research - Rattus norvegicus, Cavia porcellus, Canis familiaris and Sus scrofa domesticus We successfully isolated 4 germ cell populations from these species with average purity of 79% for spermatocytes, and 90% for spermatids and 66% for spermatogonia. Additionally, we compare the performance of mechanical and chemical dissociation for each species, and propose an optimized gating strategy to better discriminate round and elongating spermatids in the mouse, which can potentially be applied to other species. Our work indicates that spermatogenesis may be uniquely accessible among mammalian developmental systems, as a single set of reagents may be sufficient to isolate germ cell populations from many different mammalian species, opening new avenues in the fields of development and male reproductive biology.},

keywords = {Human Reproduction, Single Cell Genomics},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26473511,

title = {Improved detection of disease-associated variation by sex-specific characterization and prediction of genes required for fertility},

author = {N R Y Ho and N Huang and D F Conrad},

doi = {10.1111/andr.12109},

issn = {2047-2927},

year  = {2015},

date = {2015-11-01},

urldate = {2015-11-01},

journal = {Andrology},

volume = {3},

number = {6},

pages = {1140--1149},

abstract = {Despite its great potential, high-throughput functional genomic data are rarely integrated and applied to characterizing the genomic basis of fertility. We obtained and reprocessed over 30 functional genomics datasets from human and mouse germ cells to perform genome-wide prediction of genes underlying various reproductive phenotypes in both species. Genes involved in male fertility are easier to predict than their female analogs. Of the multiple genomic data types examined, protein-protein interactions are by far the most informative for gene prediction, followed by gene expression, and then epigenetic marks. As an application of our predictions, we show that copy number variants (CNVs) disrupting predicted fertility genes are more strongly associated with gonadal dysfunction in male and female case-control cohorts when compared to all gene-disrupting CNVs (OR = 1.64, p < 1.64 × 10(-8) vs. OR = 1.25, p < 4 × 10(-6)). Using gender-specific fertility gene annotations further increased the observed associations (OR = 2.31, p < 2.2 × 10(-16)). We provide our gene predictions as a resource with this article.},

keywords = {Genomic Variation, Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26199320,

title = {Low-frequency germline variants across 6p22.2-6p21.33 are associated with non-obstructive azoospermia in Han Chinese men},

author = {Bixian Ni and Yuan Lin and Liangdan Sun and Meng Zhu and Zheng Li and Hui Wang and Jun Yu and Xuejiang Guo and Xianbo Zuo and Jing Dong and Yankai Xia and Yang Wen and Hao Wu and Honggang Li and Yong Zhu and Ping Ping and Xiangfeng Chen and Juncheng Dai and Yue Jiang and Peng Xu and Qiang Du and Bing Yao and Ning Weng and Hui Lu and Zhuqing Wang and Xiaobin Zhu and Xiaoyu Yang and Chenliang Xiong and Hongxia Ma and Guangfu Jin and Jianfeng Xu and Xinru Wang and Zuomin Zhou and Jiayin Liu and Xuejun Zhang and Donald F Conrad and Zhibin Hu and Jiahao Sha},

doi = {10.1093/hmg/ddv257},

issn = {1460-2083},

year  = {2015},

date = {2015-10-01},

journal = {Hum Mol Genet},

volume = {24},

number = {19},

pages = {5628--5636},

abstract = {Genome-wide association studies (GWAS) have identified several common loci contributing to non-obstructive azoospermia (NOA). However, a substantial fraction of NOA heritability remains undefined, especially those low-frequency [defined here as having a minor allele frequency (MAF) between 0.5 and 5%] and rare (MAF below 0.5%) variants. Here, we performed a 3-stage exome-wide association study in Han Chinese men to evaluate the role of low-frequency or rare germline variants in NOA development. The discovery stage included 962 NOA cases and 1348 healthy male controls genotyped by exome chips and was followed by a 2-stage replication with an additional 2168 cases and 5248 controls. We identified three low-frequency variants located at 6p22.2 (rs2298090 in HIST1H1E encoding p.Lys152Arg: OR = 0.30, P = 2.40 × 10(-16)) and 6p21.33 (rs200847762 in FKBPL encoding p.Pro137Leu: OR = 0.11, P = 3.77 × 10(-16); rs11754464 in MSH5: OR = 1.78, P = 3.71 × 10(-7)) associated with NOA risk after Bonferroni correction. In summary, we report an instance of newly identified signals for NOA risk in genes previously undetected through GWAS on 6p22.2-6p21.33 in a Chinese population and highlight the role of low-frequency variants with a large effect in the process of spermatogenesis.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26203179,

title = {A Screen for Genomic Disorders of Infertility Identifies MAST2 Duplications Associated with Nonobstructive Azoospermia in Humans},

author = {Ni Huang and Yang Wen and Xuejiang Guo and Zheng Li and Juncheng Dai and Bixian Ni and Jun Yu and Yuan Lin and Wen Zhou and Bing Yao and Yue Jiang and Jiahao Sha and Donald F Conrad and Zhibin Hu},

doi = {10.1095/biolreprod.115.131185},

issn = {1529-7268},

year  = {2015},

date = {2015-09-01},

journal = {Biol Reprod},

volume = {93},

number = {3},

pages = {61},

abstract = {Since the cytogenetic identification of azoospermia factor regions 40 years ago, the Y chromosome has dominated research on the genetics of male infertility. We hypothesized that hotspots of structural rearrangement, which are dispersed across the genome, may mediate rare, recurrent copy number variations (CNVs), leading to severe infertility. We tested this hypothesis by contrasting patterns of rare CNVs in 970 Han Chinese men with idiopathic nonobstructive azoospermia and 1661 ethnicity-matched controls. Our results strongly support our previous claim that sperm production is modulated by genetic variation across the entire genome. The X chromosome in particular was enriched for loci modulating spermatogenesis--rare X-linked deletions larger than 100 kb were twice as common in patients compared with controls (odds ratio [OR] = 2.05, P = 0.01). At rearrangement hotspots across the genome, we observed a 2.4-fold enrichment of singleton CNVs in patients (P < 0.02), and we identified 117 testis genes, such as SYCE1, contained within 47 hotspots that may plausibly mediate genomic disorders of fertility. In our discovery sample we observed 3 case-specific duplications of the autosomal gene MAST2, and in a replication phase we found another 11 duplications in 1457 patients and 1 duplication in 1590 controls (P < 5 × 10(-5), combined data). With a large, polygenic genetic basis, new ways of establishing the pathogenicity of rare, large-effect mutations will be needed to fully reap the benefit of genome data in the management of azoospermia.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid26139570,

title = {Rare double sex and mab-3-related transcription factor 1 regulatory variants in severe spermatogenic failure},

author = {A C Lima and F Carvalho and J Gonçalves and S Fernandes and P I Marques and M Sousa and A Barros and S Seixas and A Amorim and D F Conrad and A M Lopes},

doi = {10.1111/andr.12063},

issn = {2047-2927},

year  = {2015},

date = {2015-09-01},

journal = {Andrology},

volume = {3},

number = {5},

pages = {825--833},

abstract = {The double sex and mab-3-related transcription factor 1 (DMRT1) gene has long been linked to sex-determining pathways across vertebrates and is known to play an essential role in gonadal development and maintenance of spermatogenesis in mice. In humans, the genomic region harboring the DMRT gene cluster has been implicated in disorders of sex development and recently DMRT1 deletions were shown to be associated with non-obstructive azoospermia (NOA). In this work, we have employed different methods to screen a cohort of Portuguese NOA patients for DMRT1 exonic insertions and deletions [by multiplex ligation probe assay (MLPA); n = 68] and point mutations (by Sanger sequencing; n = 155). We have found three novel patient-specific non-coding variants in heterozygosity that were absent from 357 geographically matched controls. One of these is a complex variant with a putative regulatory role (c.-223_-219CGAAA>T), located in the promoter region within a conserved sequence involved in Dmrt1 repression. Moreover, while DMRT1 domains are highly conserved across vertebrates and show reduced levels of diversity in human populations, two rare synonymous substitutions (rs376518776 and rs34946058) and two rare non-coding variants that potentially affect DMRT1 expression and splicing (rs144122237 and rs200423545) were overrepresented in patients when compared with 376 Portuguese controls (301 fertile and 75 normozoospermic). Overall our previous and present results suggest a role of changes in DMRT1 dosage in NOA potentially also through a process of gene misregulation, even though DMRT1 deleterious variants seem to be rare.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid23555275,

title = {Human spermatogenic failure purges deleterious mutation load from the autosomes and both sex chromosomes, including the gene DMRT1},

author = {Alexandra M Lopes and Kenneth I Aston and Emma Thompson and Filipa Carvalho and João Gonçalves and Ni Huang and Rune Matthiesen and Michiel J Noordam and Inés Quintela and Avinash Ramu and Catarina Seabra and Amy B Wilfert and Juncheng Dai and Jonathan M Downie and Susana Fernandes and Xuejiang Guo and Jiahao Sha and António Amorim and Alberto Barros and Angel Carracedo and Zhibin Hu and Matthew E Hurles and Sergey Moskovtsev and Carole Ober and Darius A Paduch and Joshua D Schiffman and Peter N Schlegel and Mário Sousa and Douglas T Carrell and Donald F Conrad},

doi = {10.1371/journal.pgen.1003349},

issn = {1553-7404},

year  = {2013},

date = {2013-03-01},

urldate = {2013-03-01},

journal = {PLoS Genet},

volume = {9},

number = {3},

pages = {e1003349},

abstract = {Gonadal failure, along with early pregnancy loss and perinatal death, may be an important filter that limits the propagation of harmful mutations in the human population. We hypothesized that men with spermatogenic impairment, a disease with unknown genetic architecture and a common cause of male infertility, are enriched for rare deleterious mutations compared to men with normal spermatogenesis. After assaying genomewide SNPs and CNVs in 323 Caucasian men with idiopathic spermatogenic impairment and more than 1,100 controls, we estimate that each rare autosomal deletion detected in our study multiplicatively changes a man's risk of disease by 10% (OR 1.10 [1.04-1.16], p<2 × 10(-3)), rare X-linked CNVs by 29%, (OR 1.29 [1.11-1.50], p<1 × 10(-3)), and rare Y-linked duplications by 88% (OR 1.88 [1.13-3.13], p<0.03). By contrasting the properties of our case-specific CNVs with those of CNV callsets from cases of autism, schizophrenia, bipolar disorder, and intellectual disability, we propose that the CNV burden in spermatogenic impairment is distinct from the burden of large, dominant mutations described for neurodevelopmental disorders. We identified two patients with deletions of DMRT1, a gene on chromosome 9p24.3 orthologous to the putative sex determination locus of the avian ZW chromosome system. In an independent sample of Han Chinese men, we identified 3 more DMRT1 deletions in 979 cases of idiopathic azoospermia and none in 1,734 controls, and found none in an additional 4,519 controls from public databases. The combined results indicate that DMRT1 loss-of-function mutations are a risk factor and potential genetic cause of human spermatogenic failure (frequency of 0.38% in 1306 cases and 0% in 7,754 controls, p = 6.2 × 10(-5)). Our study identifies other recurrent CNVs as potential causes of idiopathic azoospermia and generates hypotheses for directing future studies on the genetic basis of male infertility and IVF outcomes.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}

@article{pmid22992931,

title = {A review of genome-wide approaches to study the genetic basis for spermatogenic defects},

author = {Kenneth I Aston and Donald F Conrad},

doi = {10.1007/978-1-62703-038-0_34},

issn = {1940-6029},

year  = {2013},

date = {2013-01-01},

urldate = {2013-01-01},

journal = {Methods Mol Biol},

volume = {927},

pages = {397--410},

abstract = {Rapidly advancing tools for genetic analysis on a genome-wide scale have been instrumental in identifying the genetic bases for many complex diseases. About half of male infertility cases are of unknown etiology in spite of tremendous efforts to characterize the genetic basis for the disorder. Advancing our understanding of the genetic basis for male infertility will require the application of established and emerging genomic tools. This chapter introduces many of the tools available for genetic studies on a genome-wide scale along with principles of study design and data analysis.},

keywords = {Human Reproduction},

pubstate = {published},

tppubtype = {article}

}