We are studying asymmetric cell division (ACD), the process by which a mother cell divides to produce daughter cells that adopt different fates. In a screen for mutations that disrupt ACD in C. elegans neuroblast lineages, we isolated an allele of
grp-1. GRP-1 is the C. elegans ortholog of the cytohesin family of ARF guanine nucleotide exchange factors (GEFs) that regulate cell adhesion, actin cytoskeletal dynamics and membrane trafficking. Cytohesins have an N-terminal coiled-coil domain, a SEC7 domain that contains ARF GEF activity, and a C-terminal pleckstrin-homology domain that binds to phosphatidylinositides. Cytohesins are recruited to the plasma membrane in response to receptor tyrosine kinase activation of phosphatidylinositol-3-kinase (PI3K) where they activate ARF6. ARF6 stimulates molecules that control actin dynamics and phospholipid metabolism.
We have found that mutations in C. elegans
grp-1 disrupt ACD in certain neuroblasts, resulting in the transformation of the apoptotic daughters of these neuroblasts into their sisters and the production of extra neurons. GRP-1 acts cell non-autonomously in at least one of these divisions, suggesting that it controls a signal. To determine whether GRP-1 functions with other components of the cytohesin pathway, we used RNAi to screen candidate genes for Grp-1 phenotypes. This approach identified
arf-1.2,
arf-3,
age-1,
cnt-2, and
rng-1.
arf-1.2 and
arf-3 encode the C. elegans orthologs of Class I and II ARFs, respectively.
age-1 encodes the C. elegans ortholog of Class I PI3K that is activated by receptor tyrosine kinases.
cnt-2 encodes the worm ortholog of centaurin gamma, a protein that contains an ARF GTPase activating domain and functions in protein trafficking and the regulation of nuclear PI3K activity.
rng-1 encodes a conserved protein containing a ring zinc finger motif. Mutational analysis verified the results obtained by RNAi for
arf-1.2,
arf-3 and
age-1. Analysis of an
arf-6 deletion allele has also implicated this Class III ARF in ACD. Our results suggest that C. elegans GRP-1 functions in a cytohesin-like signaling pathway. Current efforts are directed towards characterizing the
cnt-2 and
rng-1 phenotypes, and determining whether these genes function in the GRP-1 pathway.