Spermatogenesis begins in the neonatal mouse testis with the segregation of prospermatogonia into distinct undifferentiated and differentiating populations. A proportion of undifferentiated spermatogonia retain stem cell potential (as foundational spermatogonial stem cells, or SSCs), and the remainder becomes progenitor spermatogonia that proliferate and differentiate in response to retinoic acid (RA). This initial fate decision is critical, as imbalances cause spermatogenic defects that can lead to human testicular cancer or infertility. It is currently unknown how spermatogonial fate decisions are regulated. We have recently made the exciting discovery that neonatal male germ cells differ in their responsiveness to RA both in vivo and in vitro (in the absence of the niche environment). As early as postnatal day (P) 1, we can identify two distinct subpopulations of prospermatogonia: 1 ? those that cannot respond to RA (?RA-non-responsive?) either in vitro or in vivo, and 2 ? those that can respond but do not yet in vivo (?RA-responsive?). The objective of this proposal is to determine whether the foundational SSC pool forms from prospermatogonia that are intrinsically preprogrammed (predetermination) or from those that happen to occupy a limited number of stem cell niches (selection). Our data strongly suggest that some prospermatogonia are predetermined to remain undifferentiated and become SSCs, and that this fate is manifest by an inability to respond to the proliferation and differentiation signal provided by RA. In the proposed aims, we will explore whether this RA insensitivity is intrinsic (germ cell autonomous) or determined by somatic cells (germ cell non-autonomous), define when during postnatal development differential RA responsiveness appears, and determine whether the foundational SSC pool is formed solely from these RA-insensitive germ cells.
In Aim 1, we will assess gene expression heterogeneity associated with neonatal germ cell RA responsiveness. Heterogeneous expression of genes involving RA responsiveness by either the germline or soma could explain which compartment dictates the fate of SSC precursors.
In Aim 2, we will define the roles of RA reception and catabolism in RA responsiveness. We predict that RA insensitivity is intrinsically preprogrammed, long lasting, and defines the foundational SSC population.
In Aim 3, we will determine whether RA-insensitive prospermatogonia preferentially give rise to foundational SSCs. We will modulate RA levels in vivo and examine the subsequent effect on formation of the foundational SSC pool. Together, the results of these aims will define how RA responsiveness contributes to formation of the foundational SSC pool in the neonatal testis.

Public Health Relevance

Spermatogonial stem cells (SSCs) provide the foundation for the consistent production of sperm throughout the lengthy male reproductive lifespan. The proposed studies are designed to clarify how this pool of SSCs is formed based on their differential responsiveness to the proliferation and differentiation signal provided by retinoic acid (RA). The results will increase our understanding of the mechanisms that regulate cell fate decisions during steady-state spermatogenesis in the adult testis that are critical for lifelong fertility and will be applicable to understanding how other stem cell populations are formed, many of which differentiate in response to RA or signals similar to RA.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD090083-05
Application #
10112274
Study Section
Cellular, Molecular and Integrative Reproduction Study Section (CMIR)
Program Officer
Moss, Stuart B
Project Start
2017-03-26
Project End
2022-02-28
Budget Start
2021-03-01
Budget End
2022-02-28
Support Year
5
Fiscal Year
2021
Total Cost
Indirect Cost
Name
East Carolina University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
607579018
City
Greenville
State
NC
Country
United States
Zip Code
27858
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Geyer, Christopher B (2018) A historical perspective on some ""new"" discoveries on spermatogenesis from the laboratory of Enrico Sertoli in 1878. Biol Reprod 99:479-481
Hermann, Brian P; Cheng, Keren; Singh, Anukriti et al. (2018) The Mammalian Spermatogenesis Single-Cell Transcriptome, from Spermatogonial Stem Cells to Spermatids. Cell Rep 25:1650-1667.e8