A NOVEL INDUCIBLE MEIOSIN MOUSE MODEL FOR STUDYING THE REGULATION OF MEIOTIC ENTRY DURING SPERMATOGENESIS

Objective: Elucidate the regulation involved in the decision for germ cells to initiate meiosis by determining whether inducing the expression of an essential meiotic transcription factor, MEIOSIN, is sufficient to upregulate meiotic genes and cause premature meiotic initiation. Design: It is hypothesized that absence of the transcription factor, MEIOSIN, in spermatogonial progenitors prevents meiotic entry at this time. We used histological analysis and RNA-sequencing to compare spermatogenesis in animals with MEIOSIN expression constitutively induced beginning in early spermatogonial progenitors to wildtype littermates. Materials and Methods: In each study, 3 male mice with induced MEIOSIN expression were compared to 3 male wildtype littermates. All animals had spermatogenesis developmentally synchronized using a retinoic acid (RA) synthesis inhibitor, WIN 18446, daily for 7 days followed by a single injection of RA. Synchronization allows for precise timing of developmental stage when collecting tissue samples. Testes were collected 1 day after RA administration for RNA-sequencing, and 6 days after RA for histology. Hematoxylin staining and immunofluorescence staining for meiotic proteins, SYCP3 and H2AX, were used for histological staging. Differences between samples were evaluated using the Wilcoxon rank-sum test. RNA sequencing was performed to identify changes to gene expression due to induced MEIOSIN expression. Differences were evaluated by modeling count data with a Negative Binomial distribution and using Benjamini Hochberg corrections. Results: Histological analysis revealed that cells with induced MEIOSIN expression very rarely (~5%) exhibited premature entry into meiosis. RNA-sequencing failed to show robust MEIOSIN-dependent gene expression changes, although expression trends appear to be consistent with only 5% of cells entering meiosis precociously. Conclusions: This indicates that expression of the transcription factor, MEIOSIN, is not sufficient to significantly activate the meiotic cell cycle program or to promote initiation of meiosis in early progenitors, suggesting the presence of additional mechanisms responsible for regulating the specific timing of meiotic initiation. Support: Cincinnati Children’s Research Foundation Trustee Award