Gametogenesis involves complex developmental programs executed by the integrated activities of multiple signaling networks and molecular pathways. The precise orchestration of all individual events during gametogenesis is essential for normal development. The RAS/ERK signaling pathway is a critical pathway regulating oogenesis in C. elegans. Glycogen synthase kinase 3beta (GSK-3beta), a serine/threonine kinase, was identified as a substrate of MPK-1 that regulates oocyte growth. While probing the function of gsk-3 and its impact on oocyte growth, we noticed a striking phenotype in the distal mitotic region of the gsk-3(0) germlines. To understand the nature of the phenotype in the mitotic cells of gsk-3(0) mutants, we characterized the germlines obtained from gsk-3(0) mutant adults. We found that loss of gsk-3 results in 60% reduction of proliferative cells, suggesting that GSK-3 promotes cell proliferation. Mitotic cells in a WT germline display cell cycle with both a short G1 and M phase to allow for rapid expansion of the tissue and production of gametes. Interestingly, loss of gsk-3 function, while does not alter the M phase index, results in very delayed S phase. This suggests that loss of gsk-3 alters the cell cycle structure. To test whether the G1-S transition is affected in gsk-3(0) mutants, we depleted Cye-1/Cdk2 and DNA licensing factors. Neither the loss of cye-1, cdk-2 or DNA licensing factors had an impact on mitotic cells in gsk-3(0) mutants, unlike mitotic cells in WT that underwent cell cycle arrest. This suggests that the proliferative cells in gsk-3(0) mutants skip G1, and enter an abnormal G2/M phase of the cell cycle. To determine whether mitotic cells under metabolic / nutrient stress display a similar phenotype, we assayed WT and gsk-3(0) germlines obtained from either fed/starved animals for M phase and number of proliferative cells. We find that starvation for 4-12 hours abrogates M phase in WT and gsk-3(0) germlines equally, but does not deplete the number of mitotic cells in the gsk-3(0) mutant as it does in WT. Together, our results suggest that the metabolic regulator GSK-3 regulates the adult germline mitotic cell homeostasis, such that in normal conditions, the cells are rapidly cycling with short G1 to continuously generate gametes, and do not display any adult 'stem' cell behavior (such as slow cycling / quiescent cells). However in metabolically depressed conditions / under stress, or loss of gsk-3, the cells enter dormancy/quiescence and maintain tissue output to basal levels, thus fine tuning the tissue output to metabolic conditions.

Affiliations:
- Center for genetics and genomics,
- Department of Genetics,
- Graduate School of Biomedical Science, UT MD anderson cancer center, Houston, TX, 77030