Abstract
Dysregulated Wnt signalling is associated with human infertility and testicular cancer. However, the role of Wnt signalling in male germ cells remains poorly understood. In this study, we first confirmed the activity of Wnt signalling in mouse, dog and human testes. To determine the physiological importance of the Wnt pathway, we developed a mouse model with germ cell-specific constitutive activation of βcatenin. In young mutants, similar to controls, germ cell development was normal. However, with age, mutant testes showed defective spermatogenesis, progressive germ cell loss, and flawed meiotic entry of spermatogonial cells. Flow sorting confirmed reduced germ cell populations at the leptotene/zygotene stages of meiosis in mutant group. Using thymidine analogues-based DNA double labelling technique, we further established decline in germ cell proliferation and differentiation. Overactivation of Wnt/βcatenin signalling in a spermatogonial cell line resulted in reduced cell proliferation, viability and colony formation. RNA sequencing analysis of testes revealed significant alterations in the non-coding regions of mutant mouse genome. One of the novel non-coding RNAs was switched on in mutant testes compared to controls. QPCR analysis confirmed upregulation of this unique non-coding RNA in mutant testis. In summary, our results highlight the significance of Wnt signalling in male germ cells.
Highlights
Spermatogonial stem cells (SSCs) sustain spermatogenesis throughout the life of a male by selfrenewal and differentiation to committed progenitors [1]
We found that across the species, testicular germ cells express TCF1 and LEF1 (Figure 1A-1F; N=5/ each), suggesting that Wnt signalling is active during spermatogenesis in different mammalian species
These findings suggested that constitutive activation of Wnt/βcatenin signalling in germ cells adversely affects meiotically committed progenitor cells and meiotic cells without significantly influencing spermatogonial stem/progenitor cell population
Summary
Spermatogonial stem cells (SSCs) sustain spermatogenesis throughout the life of a male by selfrenewal and differentiation to committed progenitors [1]. SSCs comprise only 0.02-0.03% of the testis and differentiate in a highly synchronized manner, consisting of mitotic expansion, meiotic divisions, and spermiogenesis. These stages are difficult to distinguish from committed progenitors via morphological analysis [1, 3]. Detailed analysis of SSC division is difficult due to lack of stage-specific markers These factors make it hard to find cues deciding the fate of SSCs
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