Identification of a novel regulator of notch signaling in vulva development and exogenous factors affecting germline stem cell proliferation in caenorhabditis elegans
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Caenorhabditis elegans vulva development primarily requires interactions between Ras and Notch signaling pathways. Three vulval precursor cells (VPCs), P5.p, P6.p and P7.p are patterned by an epidermal growth factor (EGF) mediated ‘inductive’ signal and a LIN-12/Notch-mediated ‘lateral’ signal. The inductive signal activates the Ras–MAPK pathway in P6.p which induces 1° cell fate. The subsequent expression of Notch ligands in P6.p induces the 2° cell fate in the adjacent P5.p and P7.p cells by activating the LIN-12/Notch pathway in these cells. For proper vulval cell fate patterning, Ras inhibits Notch in the 1° cell, and Notch down-regulates Ras in the 2° cells. Excessive numbers of (or a lack of) 1° or 2° cells result in abnormal vulval phenotype. We see that inactivation of the substrate recognition subunit (SRS) LRR-1, of the cullin-RING ubiquitin ligase 2 (CRL2) complex, results in a multivulva (Muv) phenotype (~1%). We show that that LRR-1 functions in regulating vulval development by inhibiting Notch signaling via negatively regulating the transcription factor DAF-12. C. elegans germ cells proliferate in an adult stem cell niche. In this study, we developed and utilized a primary tissue culture system that can maintain cultures of C. elegans germline stem cells to identify bacterial folates as a positive regulator of germ cell proliferation. Folates are a family of B-complex vitamins. Here we show that the folate 10-formyl-THF-Glu(n) and the folate precursor compound dihydropteroate stimulates germ cell proliferation, while the majority of bacterial folate species cannot. The stimulation of germ cell proliferation by 10-formyl-THF-Glu(n) does not correlate with its role as a vitamin, as other folates that cannot stimulate germ cells are more effective in rescuing folate deficiency. Further, the folate-related compound dihydropteroate stimulates germ cell proliferation despite being incapable of participating in one-carbon metabolism. The folate receptor homolog FOLR-1 is required for the germ cell stimulatory activity, but is not essential for providing folates as vitamins. This work defines a subset of bacterial folates as exogenous signals that modulates germ cell proliferation. We also identify the steroid hormone dafachronic acid as a negative regulator of stem cell proliferation.