Discovery and in vivo experimental validation of a novel, non-meiotic pathway governing production of spermatozoa and oocytes in human. A. S. Lee1, N. Huang1, Y. Yin2, R. A. Hess3, L. Ma2, P. N. Schlegel4, A. M. Lopes5, D. T. Carrell6, Z. Hu7, D. F. Conrad1 1) Department of Genetics, Washington University School of Medicine, St. Louis, MO; 2) Department of Dermatology, Washington University School of Medicine, St. Louis, MO; 3) Department of Comparative Biosciences, College of Veterinary Medicine, University of Illinois at Urbana-Champaign, Urbana, IL; 4) Department of Urology, Weill Cornell Medical College, New York-Presbyterian Hospital, New York, NY; 5) Institute of Molecular Pathology and Immunology, University of Porto, Portugal; 6) Department of Physiology, University of Utah School of Medicine, Salt Lake City, UT; 7) Department of Epidemiology and Biostatistics, School of Public Health, Nanjing Medical University, China.

   Here we report the first genome-wide association study (GWAS) for human gonadal function across males and females, and experimental validation of an exceptional result using mouse models. Although male and female infertility are typically researched as distinct diseases, some of the molecular machinery for gametogenesis is shared between both sexes. To search for a common genetic basis for sperm and egg production, we generated genome-wide CNV calls across 18,931 females (841 cases of primary ovarian insufficiency vs. 18,090 controls) and 5,197 males (1,705 cases of spermatogenic impairment vs. 3,492 controls), and used the resulting callsets to perform rare variant GWAS separately in males and females. Remarkably, the strongest association signal in both sexes was found at the same locus (CSMD1). CSMD1 is located on 8p23, within the region of highest sequence divergence between human and chimpanzee outside the Y chromosome (on average 0.03 substitutions per site across 1Mb).
   CSMD1 is expressed in both the testes and ovaries and shares primary sequence homology with complement-interacting proteins, but little else is known of its biological function. Thus we turned to the mouse model for an in vivo reproductive model. We took two parallel approaches to perturb Csmd1 in the mouse: (i) we delivered an shRNA targeting Csmd1 directly into the testes via lentiviral vector, and (ii) we generated a colony of Csmd1 gene knockout mice. We then performed extensive cellular and molecular phenotyping of the gonads, including histopathology; computer-assisted semen analysis; purification of germ cells via FACS; and RNAseq of whole testis, whole ovary, and FACS-purified male germ cells in wildtype and knockout.
   Csmd1-deficient males show near-complete histological degeneration of the testes as early as 34 days of age (approx. onset of sexual maturity), with concomitant reduction in sperm count and motility. Remarkably, in both the post-meiotic (haploid) male germ cells and female ovaries, Csmd1-knockouts show deranged expression of ciliary and flagellar components of the axoneme (GO:0005930, GO:0044447, GO:0035082) which are typically associated with sperm motility. Taken together, these results suggest that at least a subset of overlapping processes govern both male and female germ cell development, and that CSMD1 plays a crucial role in their maintenance.

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