The Trans-Omics for Precision Medicine (TOPMed) programme seeks to elucidate the genetic architecture and biology of heart, lung, blood and sleep disorders, with the ultimate goal of improving diagnosis, treatment and prevention of these diseases. The initial phases of the programme focused on whole-genome sequencing of individuals with rich phenotypic data and diverse backgrounds. Here we describe the TOPMed goals and design as well as the available resources and early insights obtained from the sequence data. The resources include a variant browser, a genotype imputation server, and genomic and phenotypic data that are available through dbGaP (Database of Genotypes and Phenotypes)¹. In the first 53,831 TOPMed samples, we detected more than 400 million single-nucleotide and insertion or deletion variants after alignment with the reference genome. Additional previously undescribed variants were detected through assembly of unmapped reads and customized analysis in highly variable loci. Among the more than 400 million detected variants, 97% have frequencies of less than 1% and 46% are singletons that are present in only one individual (53% among unrelated individuals). These rare variants provide insights into mutational processes and recent human evolutionary history. The extensive catalogue of genetic variation in TOPMed studies provides unique opportunities for exploring the contributions of rare and noncoding sequence variants to phenotypic variation. Furthermore, combining TOPMed haplotypes with modern imputation methods improves the power and reach of genome-wide association studies to include variants down to a frequency of approximately 0.01%.

Hormone-induced brain masculinization occurs during a perinatal sensitive period but endures into adulthood. Researchers explored DNA methylation as a candidate mechanism. Methylation is higher in female brain and suppresses masculinization genes, which are liberated by hormone-induced reductions in DNMT activity in males. Pharmacological inhibition of DNMTs reduces methylation, masculinizes female brain and behavior and reopens the sensitive period. The developing mammalian brain is destined for a female phenotype unless exposed to gonadal hormones during a perinatal sensitive period. It has been assumed that the undifferentiated brain is masculinized by direct induction of transcription by ligand-activated nuclear steroid receptors. We found that a primary effect of gonadal steroids in the highly sexually dimorphic preoptic area (POA) is to reduce activity of DNA methyltransferase (Dnmt) enzymes, thereby decreasing DNA methylation and releasing masculinizing genes from epigenetic repression. Pharmacological inhibition of Dnmts mimicked gonadal steroids, resulting in masculinized neuronal markers and male sexual behavior in female rats. Conditional knockout of the de novo Dnmt isoform, Dnmt3a, also masculinized sexual behavior in female mice. RNA sequencing revealed gene and isoform variants modulated by methylation that may underlie the divergent reproductive behaviors of males versus females. Our data show that brain feminization is maintained by the active suppression of masculinization via DNA methylation.

Extracellular vesicles (EVs) are a unique mode of intercellular communication capable of incredible specificity in transmitting signals involved in cellular function, including germ cell maturation. Spermatogenesis occurs in the testes, behind a protective barrier to ensure safeguarding of germline DNA from environmental insults. Following DNA compaction, further sperm maturation occurs in the epididymis. Here, we report reproductive tract EVs transmit information regarding stress in the paternal environment to sperm, potentially altering fetal development. Using intracytoplasmic sperm injection, we found that sperm incubated with EVs collected from stress-treated epididymal epithelial cells produced offspring with altered neurodevelopment and adult stress reactivity. Proteomic and transcriptomic assessment of these EVs showed dramatic changes in protein and miRNA content long after stress treatment had ended, supporting a lasting programmatic change in response to chronic stress. Thus, EVs as a normal process in sperm maturation, can also perform roles in intergenerational transmission of paternal environmental experience.

Significance: Through the Peruvian Genome Project we generate and analyze the genomes of 280 individuals where the majority have >90% Native American ancestry and explore questions at the interface of evolutionary genetics, history, anthropology, and medicine. This is the most extensive sampling of high-coverage Native American and mestizo whole genomes to date. We estimate an initial peopling of Peru was rapid and began by 12,000 y ago. In addition, the mestizo populations exhibit admixture between Native American groups prior to their Spanish admixture and was likely influenced by the Inca Empire and Spanish conquest. Our results address important Native American population history questions and establish a dataset beneficial to address the underrepresentation of Native American ancestry in sequencing studies.

Treatment of vulvovaginal candidiasis (VVC), caused most frequently by Candida albicans , represents a significant unmet clinical need. C. albicans , as both a commensal and a pathogenic organism, has a complex and poorly understood interaction with the vaginal environment. Understanding the complex nature of this relationship is necessary for the development of desperately needed therapies to treat symptomatic infection. Using transcriptome sequencing (RNA-seq), we characterized the early murine vaginal and fungal transcriptomes of the organism during VVC. Network analysis of host genes that were differentially expressed between infected and naive mice predicted the activation or repression of several signaling pathways that have not been previously associated with VVC, including NLRP3 inflammasome activation. Intravaginal challenge of Nlrp3 ^−/− mice with C. albicans demonstrated severely reduced levels of polymorphonuclear leukocytes (PMNs), alarmins, and inflammatory cytokines, including interleukin-1β (IL-1β) (the hallmarks of VVC immunopathogenesis) in vaginal lavage fluid. Intravaginal administration of wild-type (WT) mice with glyburide, a potent inhibitor of the NLRP3 inflammasome, reduced PMN infiltration and IL-1β to levels comparable to those observed in Nlrp3 ^−/− mice. Furthermore, RNA-seq analysis of C. albicans genes indicated robust expression of hypha-associated secreted aspartyl proteinases 4, 5, and 6 (SAP4–6), which are known inflammasome activators. Despite colonization similar to that of the WT strain, Δ SAP4–6 triple and Δ SAP5 single mutants induced significantly less PMN influx and IL-1β during intravaginal challenge. Our findings demonstrate a novel role for the inflammasome in the immunopathogenesis of VVC and implicate the hypha-associated SAPs as major C. albicans virulence determinants during vulvovaginal candidiasis. IMPORTANCE Vaginitis, most commonly caused by the fungus Candida albicans , results in significant quality-of-life issues for all women of reproductive age. Recent efforts have suggested that vaginitis results from an immunopathological response governed by host innate immunity, although an explanatory mechanism has remained undefined. Using comprehensive genomic, immunological, and pharmacological approaches, we have elucidated the NLRP3 inflammasome as a crucial molecular mechanism contributing to host immunopathology. We have also demonstrated that C. albicans hypha-associated secreted aspartyl proteinases (SAP4–6 and SAP5, more specifically) contribute to disease immunopathology. Ultimately, this study enhances our understanding of the complex interplay between host and fungus at the vaginal mucosa and provides proof-of-principle evidence for therapeutic targeting of inflammasomes for symptomatic vulvovaginal candidiasis.