Leydig cells are interstitial cells of the testis and represent the major source of testosterone (T) production in men. Normal T production is critical not only for normal sperm production and male fertility, but also for normal sexual behavior and male-specific physiology in general. The molecular mechanism underlying the development and function of Leydig cells remains largely undefined. We recently demonstrated in mice that when we inhibited the microRNA (miRNA) production by inactivating either of the two key RNase III enzymes, Dicer or Drosha, which are known to be essential for miRNA biosynthesis, Leydig cells could not function normally. In the absence of miRNAs, Leydig cells in adult mice overproduced testosterone, and the hypothalamus-pituitary-testis negative feedback system malfunctioned, leading to persistently elevated levels of T, decreased levels of estradiol (E2), and slightly increased levels of both gonadotropins (FSH and LH). This abnormal hormonal profile is accompanied by sperm defects characterized by separation of sperm heads from tails, lack of motility and deformation of sperm heads, which led to complete infertility in male mutant mice. Our preliminary data suggest that normal miRNA production is essential for normal Leydig cell functions [i.e. normal T and E2 production and normal hypothalamus-pituitary- testis (HPT) axis feedback], normal sperm production and male fertility. We, therefore, propose this pilot study to answer two fundamental questions concerning Leydig cell miRNAs: 1) How many miRNAs are normally expressed in Leydig cells? To answer this question, we will define the miRNA transcriptome in control and miRNA-deficient Leydig cells using miRNA deep sequencing (miRNA-Seq). 2) How does depletion of miRNAs affect mRNA transcriptome and proteome in Leydig cells? To answer these questions, we will define the mRNA transcriptome and proteome in adult control and miRNA-deficient Leydig cells using mRNA-Seq and iTRAQ-based mass spectrometry (MS). Data from this study will allow us to establish correlations between dysregulated genes leading to abnormal steroidogenesis and the absence of critical Leydig cell miRNAs (end-point of this study), which will serve as the basis for a further study on the specific mechanisms underlying the control of Leydig cell functions by miRNAs (starting point of the future R01 proposal). Future investigation based upon this pilot study may lead to novel diagnostics and treatments for male infertility due to endocrine disorders, and miRNAs may represent a novel class of male contraceptive agents.

Public Health Relevance

This project will identify all microRNAs, mRNAs and proteins that are expressed in adult Leydig cells, and will further determine the role of microRNAs in the control of steroidogenesis in Leydig cells. Data from this study may lead to novel diagnostics and treatments for endocrine disorders that cause male infertility, as well as novel male contraceptive measures.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Small Research Grants (R03)
Project #
1R03HD074573-01A1
Application #
8583150
Study Section
Pediatrics Subcommittee (CHHD)
Program Officer
Moss, Stuart B
Project Start
2013-08-01
Project End
2015-05-31
Budget Start
2013-08-01
Budget End
2014-05-31
Support Year
1
Fiscal Year
2013
Total Cost
$70,295
Indirect Cost
$20,295
Name
University of Nevada Reno
Department
Physiology
Type
Schools of Medicine
DUNS #
146515460
City
Reno
State
NV
Country
United States
Zip Code
89557
Yuan, Shuiqiao; Schuster, Andrew; Tang, Chong et al. (2016) Sperm-borne miRNAs and endo-siRNAs are important for fertilization and preimplantation embryonic development. Development 143:635-47
Bao, Jianqiang; Tang, Chong; Yuan, Shuiqiao et al. (2015) UPF2, a nonsense-mediated mRNA decay factor, is required for prepubertal Sertoli cell development and male fertility by ensuring fidelity of the transcriptome. Development 142:352-62
Ortogero, Nicole; Hennig, Grant W; Luong, Dickson et al. (2015) Computer-assisted annotation of small RNA transcriptomes. Methods Mol Biol 1218:353-64
Yuan, Shuiqiao; Qin, Weibing; Riordan, Connor R et al. (2015) Ubqln3, a testis-specific gene, is dispensable for embryonic development and spermatogenesis in mice. Mol Reprod Dev 82:266-7
Oliver, Daniel; Yuan, Shuiqiao; McSwiggin, Hayden et al. (2015) Pervasive Genotypic Mosaicism in Founder Mice Derived from Genome Editing through Pronuclear Injection. PLoS One 10:e0129457
Yuan, Shuiqiao; Oliver, Daniel; Schuster, Andrew et al. (2015) Breeding scheme and maternal small RNAs affect the efficiency of transgenerational inheritance of a paramutation in mice. Sci Rep 5:9266
Yuan, Shuiqiao; Swiggin, Hayden M C; Zheng, Huili et al. (2015) A testis-specific gene, Ubqlnl, is dispensable for mouse embryonic development and spermatogenesis. Mol Reprod Dev 82:408-9
Wu, Jingwen; Bao, Jianqiang; Kim, Minkyung et al. (2014) Two miRNA clusters, miR-34b/c and miR-449, are essential for normal brain development, motile ciliogenesis, and spermatogenesis. Proc Natl Acad Sci U S A 111:E2851-7
Bao, J; Zhang, Y; Schuster, A S et al. (2014) Conditional inactivation of Miwi2 reveals that MIWI2 is only essential for prospermatogonial development in mice. Cell Death Differ 21:783-96
Ortogero, Nicole; Schuster, Andrew S; Oliver, Daniel K et al. (2014) A novel class of somatic small RNAs similar to germ cell pachytene PIWI-interacting small RNAs. J Biol Chem 289:32824-34

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