In mammals, four slit-robo GTPase activating proteins (GAPs) are expressed during brain development and have been shown in tissue culture to be negative regulators of neuronal migration, via inhibition of RAC1 and/or CDC42 activity. Mutations in srGAP3 have been linked to certain types of mental retardation. The sole C. elegans homolog, which we named SRGP-1, shares a high degree of conservation (27% identity; 42% similarity) with mammalian srGAPs. The
srgp-1 promoter fused to GFP (BC10607) drives expression in the adult worm nervous system, and embryonically, it is highly expressed in neuroblasts and in the hypodermis. Knocking down
srgp-1 via RNAi in N2 worms has no overt phenotype. However,
srgp-1(RNAi) in alpha - or beta-catenin hypomorphic mutants with compromised cell-cell adhesion results in 100% embryonic lethality. The primary defect in
hmp-2(
qm39) or
hmp-1(
fe4);
srgp-1(RNAi) embryos is a failure of the final ventral gastrulation cleft to close. We carefully examined cleft closure using a GFP reporter in neuroblasts (Pkal-1::GFP). In wild-type embryos, the five-cell long gastrulation cleft closes by "zipping" of the neuroblasts from both ends inward and from bottom to top. This movement is accompanied by short-range protrusions and is completed within 30 minutes. In
srgp-1(RNAi) embryos we find an increase in the number and length of neuroblast protrusions and it takes the cleft 1.5 times longer to seal. In
hmp-2(
qm39) and
hmp-1(
fe4) embryos the zipping appears indecisive; cells adhere and then separate again. Cleft closure takes up to twice as long compared to wild-type. The combination of deregulated protrusions and weaker cell-cell adhesion in
hmp-2(
qm39) or
hmp-1(
fe4);
srpg-1(RNAi) results in ventral clefts persisting at a stage when internal pressure builds in preparation for elongation. This leads to cells popping out of the open cleft. Our genetic studies suggest
srgp-1 is downstream of
sax-3/Robo and acts in parallel to
vab-1 and
wve-1. Interestingly, SRGP-1 does not contain a C-terminal SH3 domain, which is implicated in binding to the ROBO receptor and WAVE-1 in mammals. In addition to its GAP domain SRGP-1 has a F-BAR domain, which was shown in vitro to sense and/or induce lipid membrane curvature. The in vivo role of F-BAR domains is still unknown. Our current work is aimed at elucidating the roles of the GAP and F-BAR domains of SRGP-1 via specific deletions and point mutations in conjuction with functional and localization experiments.