Abstract
BackgroundThe development of a safe, effective, reversible, non-hormonal contraceptive method for men has been an ongoing effort for the past few decades. However, despite significant progress on elucidating the function of key proteins involved in reproduction, understanding male reproductive physiology is limited by incomplete information on the genes expressed in reproductive tissues, and no contraceptive targets have so far reached clinical trials. To advance product development, further identification of novel reproductive tract-specific genes leading to potentially druggable protein targets is imperative.ResultsIn this study, we expand on previous single tissue, single species studies by integrating analysis of publicly available human and mouse RNA-seq datasets whose initial published purpose was not focused on identifying male reproductive tract-specific targets. We also incorporate analysis of additional newly acquired human and mouse testis and epididymis samples to increase the number of targets identified. We detected a combined total of 1178 genes for which no previous evidence of male reproductive tract-specific expression was annotated, many of which are potentially druggable targets. Through RT-PCR, we confirmed the reproductive tract-specific expression of 51 novel orthologous human and mouse genes without a reported mouse model. Of these, we ablated four epididymis-specific genes (Spint3, Spint4, Spint5, and Ces5a) and two testis-specific genes (Pp2d1 and Saxo1) in individual or double knockout mice generated through the CRISPR/Cas9 system. Our results validate a functional requirement for Spint4/5 and Ces5a in male mouse fertility, while demonstrating that Spint3, Pp2d1, and Saxo1 are each individually dispensable for male mouse fertility.ConclusionsOur work provides a plethora of novel testis- and epididymis-specific genes and elucidates the functional requirement of several of these genes, which is essential towards understanding the etiology of male infertility and the development of male contraceptives.
Highlights
The development of a safe, effective, reversible, non-hormonal contraceptive method for men has been an ongoing effort for the past few decades
Study approaches and data Despite significant advances in our understanding of the human and rodent testis and epididymis transcriptome, mostly through microarray-based studies, no prior studies have utilized purified human testis cells for the identification of human testis-specific transcripts, no prior studies have utilized the more state-of-the-art RNA-seqbased transcriptomics methodology for analysis of human epididymis-specific transcripts, and no prior studies have utilized RNA-seq analysis of rodent reproductive tissues or cells to identify rodent reproductive tractspecific transcripts. To address these gaps in knowledge, and to increase the number of identified reproductive tract-specific genes in both species using the most relevant high-throughput transcriptomics methodology, we analyzed in parallel on a custom bioinformatics pipeline a large number of published and newly acquired human and mouse RNA-seq datasets
Of the seventy-three genes that our study identifies as reproductive tract-specific in humans and for which a published mouse model shows male infertility phenotype (Additional file 14: Fig. S6) [28, 29, 31], it is worth noting that 21 genes—CNBD2, DEFB110, FAM170A, FBXO47, MEIG1, MEIOB, MEIOC, ODF1, ODF4, REC114, RNF17, SPACA1, SPATA22, SPEM1, SPO11, SYCP1, TERB1, TEX19, TEX38, TNP2, and TOPAZ—do not have any associated paralogs and, thereby, may be considered most suitable for further drug development
Summary
The development of a safe, effective, reversible, non-hormonal contraceptive method for men has been an ongoing effort for the past few decades. Despite significant progress on elucidating the function of key proteins involved in reproduction, understanding male reproductive physiology is limited by incomplete information on the genes expressed in reproductive tissues, and no contraceptive targets have so far reached clinical trials. There are numerous contraceptive options available to women; identification of a safe, non-hormonal contraceptive option for men is still an ongoing challenge. Our understanding of the mechanisms underlying male reproductive physiology is still at an early stage as the identification and elucidation of the function of key reproductive proteins is still an ongoing effort. Identifying druggable protein targets expressed in the male reproductive tract has been the focus of numerous studies dedicated to the development of male contraception
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