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

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,261023), rare X-linked CNVs by 29%, (OR 1.29 [1.11–1.50], p,161023), 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 = 6.261025). 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.This work was partially funded by the Portuguese Foundation for Science and Technology FCT/MCTES (PIDDAC) and co-financed by European funds (FEDER) through the COMPETE program, research grant PTDC/SAU-GMG/101229/2008. IPATIMUP is an Associate Laboratory of the Portuguese Ministry of Science, Technology, and Higher Education and is partially supported by FCT. AML is the recipient of a postdoctoral fellowship from FCT (SFRH/BPD/73366/2010). CO is supported by a grant from the United States National Institutes of Health (R01 HD21244), JDS is supported by Damon Runyon Clinical Investigator Award, Alex's Lemonade Stand Foundation Epidemiology Award, and the Eunice Kennedy Shriver Children's Health Research Career Development Award NICHD 5K12HD001410. Support for humans studies and specimens were provided by the NIH/NIDDK George M. O'Brien Center for Kidney Disease Kidney Translational Research Core (P30DK079333) grant to Washington University. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

The genomes of two key bumblebee species with primitive eusocial organization

Background: The shift from solitary to social behavior is one of the major evolutionary transitions. Primitively eusocial bumblebees are uniquely placed to illuminate the evolution of highly eusocial insect societies. Bumblebees are also invaluable natural and agricultural pollinators, and there is widespread concern over recent population declines in some species. High-quality genomic data will inform key aspects of bumblebee biology, including susceptibility to implicated population viability threats. Results: We report the high quality draft genome sequences of Bombus terrestris and Bombus impatiens, two ecologically dominant bumblebees and widely utilized study species. Comparing these new genomes to those of the highly eusocial honeybee Apis mellifera and other Hymenoptera, we identify deeply conserved similarities, as well as novelties key to the biology of these organisms. Some honeybee genome features thought to underpin advanced eusociality are also present in bumblebees, indicating an earlier evolution in the bee lineage. Xenobiotic detoxification and immune genes are similarly depauperate in bumblebees and honeybees, and multiple categories of genes linked to social organization, including development and behavior, show high conservation. Key differences identified include a bias in bumblebee chemoreception towards gustation from olfaction, and striking differences in microRNAs, potentially responsible for gene regulation underlying social and other traits. Conclusions: These two bumblebee genomes provide a foundation for post-genomic research on these key pollinators and insect societies. Overall, gene repertoires suggest that the route to advanced eusociality in bees was mediated by many small changes in many genes and processes, and not by notable expansion or depauperation

The discovery, distribution, and evolution of viruses associated with drosophila melanogaster

Drosophila melanogaster is a valuable invertebrate model for viral infection and antiviral immunity, and is a focus for studies of insect-virus coevolution. Here we use a metagenomic approach to identify more than 20 previously undetected RNA viruses and a DNA virus associated with wild D. melanogaster. These viruses not only include distant relatives of known insect pathogens, but also novel groups of insect-infecting viruses. By sequencing virus-derived small RNAs we show that the viruses represent active infections of Drosophila. We find that the RNA viruses differ in the number and properties of their small RNAs, and we detect both siRNAs and a novel miRNA from the DNA virus. Analysis of small RNAs also allows us to identify putative viral sequences that lack detectable sequence similarity to known viruses. By surveying >2000 individually collected wild adult Drosophila we show that more than 30% of D. melanogaster carry a detectable virus, and more than 6% carry multiple viruses. However, despite a high prevalence of the Wolbachia endosymbiont—which is known to be protective against virus infections in Drosophila—we were unable to detect any relationship between the presence of Wolbachia and the presence of any virus. Using publicly available RNA-seq datasets we show that the community of viruses in Drosophila laboratories is very different from that seen in the wild, but that some of the newly discovered viruses are nevertheless widespread in laboratory lines and are ubiquitous in cell culture. By sequencing viruses from individual wild-collected flies we show that some viruses are shared between D. melanogaster and D. simulans. Our results provide an essential evolutionary and ecological context for host-virus interaction in Drosophila, and the newly reported viral sequences will help develop D. melanogaster further as a model for molecular and evolutionary virus research

Making Sense of Singletons: The N-of-One Problem in Human Genetics

The diagnosis of rare, idiopathic diseases is emerging as a primary application of medical genome sequencing. However, the application of standard tools from genetic epidemiology for many of these cases is frustrated by a combination of small sample sizes, genetic heterogeneity and the large number of singleton variants found by genome sequencing. In response, we have developed a statistical inference framework that is optimized for identifying unusual functional variation from a single genome, what we refer to as the n-of-one problem. Our statistical framework addresses the n-of-one problem in two steps, first by scoring single nucleotide variants (SNVs) according to their predicted pathogenicity, and then by calculating a test statistic based on these predictions and evaluating the significance using null distributions parameterized with population genetic data from healthy individuals. To implement this framework, we first evaluated the performance of several pathogenicity prediction methods, including a logistic regression based model developed in-house, before selecting the Combined Annotation Dependent Depletion (CADD) score as our metric for scoring pathogenicity. Next, population genetic data from over 60,000 individuals from the Exome Aggregation Consortium (ExAC) was used to parameterize population genetic models and generate gene-specific null models of the healthy population for three distinct disease inheritance models. Using this approach we assess our ability to identify the causal genotypes in over 5 million simulated cases of Mendelian disease, finding that 39% of disease genotypes are the most damaging unit in a typical exome background. We applied our approach to several cohorts of rare disease, including 129 n-of-one families from the Undiagnosed Diseases Program, nominating 60% of 30 genotype or genotype pairs determined to be diagnostic by a standard clinical workup as the most likely candidate in that family. We show that our approach provides a powerful way to include population databases in an integrative analysis by combining population sampling probabilities with gene expression data to improve the detection of UDP diagnostic genes. Currently, our method can produce well calibrated p-values when applied to single genomes, and, with further work, could become a widely used epidemiological method, like linkage analysis or GWAS. We further demonstrate the utility of population genetic data by leveraging pair genotype and gene expression data available through the Genotype-Tissue expression (GTEx) consortium to develop a framework to identify copy number variants (CNVs) that can modulate gene expression. Due to their size studies have found that an individual CNV is capable of disrupting the expression of multiple genes. Our framework leverages this finding to increase our power to detect expression modulating CNVs that are observed in only a single donor. We then characterize the expression modulating CNVs identified with our framework and discuss ways in which this model could be used to extend our n-of-one framework to CNVs and datasets that would be appropriate for analysis with this proposed framework

Recurrent de novo mutations in neurodevelopmental disorders: properties and clinical implications

Abstract Next-generation sequencing (NGS) is now more accessible to clinicians and researchers. As a result, our understanding of the genetics of neurodevelopmental disorders (NDDs) has rapidly advanced over the past few years. NGS has led to the discovery of new NDD genes with an excess of recurrent de novo mutations (DNMs) when compared to controls. Development of large-scale databases of normal and disease variation has given rise to metrics exploring the relative tolerance of individual genes to human mutation. Genetic etiology and diagnosis rates have improved, which have led to the discovery of new pathways and tissue types relevant to NDDs. In this review, we highlight several key findings based on the discovery of recurrent DNMs ranging from copy number variants to point mutations. We explore biases and patterns of DNM enrichment and the role of mosaicism and secondary mutations in variable expressivity. We discuss the benefit of whole-genome sequencing (WGS) over whole-exome sequencing (WES) to understand more complex, multifactorial cases of NDD and explain how this improved understanding aids diagnosis and management of these disorders. Comprehensive assessment of the DNM landscape across the genome using WGS and other technologies will lead to the development of novel functional and bioinformatics approaches to interpret DNMs and drive new insights into NDD biology

Differences in the number of de novo mutations between individuals are due to small family-specific effects and stochasticity

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Human Spermatogenic Failure Purges Deleterious Mutation Load from the Autosomes and Both Sex Chromosomes, including the Gene DMRT1

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

Homozygous missense mutation of <i>INHBB</i> identified in the case of UPD2.

<p>(A) We validated this candidate by Sanger sequencing in the UPD2 case and control individuals. Mutant and reference nucleotides are highlighted within the blue box, confirming the homozygous T to C nucleotide change observed at chr2:12,1107,305 bp (hg19) of the UPD2 individual. Grey boxes represent the exons of the gene and the red line indicates the location of the observed mutation within the gene. (B) <i>INHBB</i> encodes for the protein, Inhibin βB, which along with inhibin α and inhibin βA, combine combinatorially to form the inhibins and activins. Each protein expressed by <i>INHA</i>, <i>INHBA</i>, <i>INHBB</i> consists of an N-terminal signal peptide (purple), a propeptide (grey), and a subunit chain (green, red or yellow). The mutation identified here results in a M370T change of the inhibin βB subunit chain (location indicated by a vertical red line throughout the diagram). The various inhibin subunits dimerize via disulfide bonds (locations indicated by black lines between subunits). As the βB subunit participates in multiple complexes with antagonistic functions, the functional consequences of loss-of-function or gain-of-function mutations in this protein may be difficult to predict. (C) The role of inhibins and activins in the hypothalamic-pituitary testicular axis. These complexes have diverse functions in the body, but are most well known for their ability to stimulate and inhibit follicle stimulating hormone (FSH) production, a process critical for spermatogenesis. Blue arrows connect hormones to the cell or gland by which they are secreted. Green arrows indicate stimulatory interactions, and red lines indicate inhibitory interactions.</p

Summary of inbreeding coefficient estimates across cohorts, and association testing.

<p>For each case and control group we present the average the estimated inbreeding coefficient and the number of individuals with inbreeding coefficients above a specified threshold. The last column indicates the total number of individuals in each group. The bottom two rows indicate the results of an association test between inbreeding and case/control status using either a categorical variable as a definition of inbreeding status (<i>F</i>>0.5%, <i>F</i>>1.6%, and <i>F</i>>6.25%) or using the inbreeding coefficient as a continuous variable (“All <i>F</i>”).</p