In C. elegans, all 302 neurons are generated by asymmetric cell division. To investigate the mechanisms underlying asymmetric neuroblast division we have focused on the lineage that generates PHA, a sensory neuron in the tail. Asymmetric division in this lineage requires
dsh-2 and
mom-5, which encode Dishevelled (Dsh) and Frizzled (Fz) homologs respectively. We have shown that MOM-5 regulates the distribution of DSH-2 protein to the cell cortex, suggesting that MOM-5 regulates asymmetric cell division by controlling DSH-2 localization. DSH-2 and MOM-5 are components of both canonical and non-canonical Wnt signaling pathways. To elucidate which pathway is controlling asymmetric division, we performed a domain analysis of DSH-2. All Dshs contain three highly conserved domains: a N-terminal DIX domain, a central PDZ domain and a C-terminal DEP domain. The DIX domain is essential for canonical Wnt signaling while the DEP domain is required for non-canonical signaling. Deletion of the DIX domain did not disrupt DSH-2 localization to the cell cortex. Deletion of the DEP domain, however, delocalized DSH-2 from the cell cortex to the cytoplasm. We have shown that a DSH-2 construct lacking the DIX domain is able to rescue
dsh-2 asymmetric division defects, while no rescue is observed with DSH-2 lacking the DEP domain. Thus, we propose that DSH-2 and MOM-5 may operate through a non-canonical Wnt pathway. We have evidence that CWN-1 likely functions upstream of MOM-5 and DSH-2. We have generated a CWN-1::GFP fusion construct to analyze the pattern of CWN-1 expression. In addition, we are currently determining if CWN-1 misexpression disrupts asymmetric neuroblast division. Because our attempts to identify genes that function downstream of
dsh-2 by RNAi were unsuccessful, we took a genetic approach. In addition to division defects, loss of
dsh-2 function also results in maternal effect embryonic lethality. Therefore, we performed a genetic screen to isolate suppressors of the
dsh-2 lethality. We isolated over 60 suppressors, the majority of which suppressed the division defects in the PHA lineage. While the majority of the suppressors were dominant, we have isolated 4 X-linked suppressors and 3 autosomal recessive suppressors. We have mapped one of our X-linked recessive to the right of
lin-15 on the far right end of the X-chromosome. There are no known Wnt of PCP pathway genes in this region indicating that we have likely identified a novel gene that functions to modulate
dsh-2 signaling. We are also in the process of mapping a dominant suppressor on the X chromosome.