Developmental fidelity is imposed by genetically separable ralgef activities that mediate opposing signals

Abstract

The six C. elegans vulval precursor cells (VPCs) are induced to form the 3˚-3˚-2˚-1˚-2˚-3˚ pattern of cell fates with high fidelity. In response to EGF signal, the LET-60/Ras-LIN-45/ Raf-MEK-2/MEK-MPK-1/ERK canonical MAP kinase cascade is necessary to induce 1˚ fate and synthesis of DSL ligands for the lateral Notch signal. In turn, LIN-12/Notch receptor is necessary to induce neighboring cells to become 2˚. We previously showed that, in response to graded EGF signal, the modulatory LET-60/Ras-RGL-1/RalGEF-RAL-1/Ral signal promotes 2˚ fate in support of LIN-12. In this study, we identify two key differences between RGL-1 and RAL-1. First, deletion of RGL-1 confers no overt developmental defects, while previous studies showed RAL-1 to be essential for viability and fertility. From this observation, we hypothesize that the essential functions of RAL-1 are independent of upstream activation. Second, RGL-1 plays opposing and genetically separable roles in VPC fate patterning. RGL-1 promotes 2˚ fate via canonical GEF-dependent activation of RAL-1. Conversely, RGL-1 promotes 1˚ fate via a non-canonical GEF-independent activity. Our genetic epistasis experiments are consistent with RGL-1 functioning in the modulatory 1˚-promoting AGE-1/PI3-Kinase-PDK-1-AKT-1 cascade. Additionally, animals lacking RGL-1 experience 15-fold higher rates of VPC patterning errors compared to the wild type. Yet VPC patterning in RGL-1 deletion mutants is not more sensitive to environmental perturbations. We propose that RGL-1 functions to orchestrate opposing 1˚-and 2˚-promoting modulatory cascades to decrease developmental stochasticity. We speculate that such switches are broadly conserved but mostly masked by paralog redundancy or essential functions.