A PUF Hub Drives Self-Renewal in <i>Caenorhabditis elegans</i> Germline Stem Cells

Abstract

Stem cell regulation relies on extrinsic signaling from a niche plus intrinsic factors that respond and drive self-renewal within stem cells. <i>A priori</i>, loss of niche signaling and loss of the intrinsic self-renewal factors might be expected to have equivalent stem cell defects. Yet this simple prediction has not been borne out for most stem cells, including <i>Caenorhabditis elegans</i> germline stem cells (GSCs). The central regulators of <i>C. elegans</i> GSCs include extrinsically acting GLP-1/Notch signaling from the niche; intrinsically acting RNA-binding proteins in the PUF family, termed FBF-1 and FBF-2 (collectively FBF); and intrinsically acting PUF partner proteins that are direct Notch targets. Abrogation of either GLP-1/Notch signaling or its targets yields an earlier and more severe GSC defect than loss of FBF-1 and FBF-2, suggesting that additional intrinsic regulators must exist. Here, we report that those missing regulators are two additional PUF proteins, PUF-3 and PUF-11 Remarkably, an <i><i>fbf-1</i></i><i><i>fbf-2</i></i> ; <i><i>puf-3</i></i><i><i>puf-11</i></i> quadruple null mutant has a GSC defect virtually identical to that of a <i>glp-1</i><i>/</i>Notch null mutant. PUF-3 and PUF-11 both affect GSC maintenance, both are expressed in GSCs, and epistasis experiments place them at the same position as FBF within the network. Therefore, action of PUF-3 and PUF-11 explains the milder GSC defect in <i><i>fbf-1</i></i><i><i>fbf-2</i></i> mutants. We conclude that a "PUF hub," comprising four PUF proteins and two PUF partners, constitutes the intrinsic self-renewal node of the <i>C. elegans</i> GSC RNA regulatory network. Discovery of this hub underscores the significance of PUF RNA-binding proteins as key regulators of stem cell maintenance.