Cell polarity and asymmetric cell divisions are essential for the generation of cellular diversity. The importance of these processes during development has been emphasized by recent findings, which show that defects lead to the formation of tumors. Towards identifying most of the factors involved in cell polarity and asymmetric cell division, we carried out 17 large scale RNAi suppressor screens of temperature sensitive mutants of genes involved in actomyosin contraction, par polarity and microtubule pulling forces governing spindle positioning. This allowed us to generate a polarity genetic interaction network of 186 genes with 229 genetic interactions (see Fievet et al., abstract). Of these genes, 23 have been previously shown to have a role in the asymmetric first cell division, validating our approach. For the rest of the candidates (87% of our network), no early polarity phenotype has been reported for knock-down in a wt background.
To identify functions of novel candidates, we have been studying effects of their knockdown in sensitized backgrounds. This strategy has successfully identified specific functions for genes in the network. For example, out of the 24 identified suppressors of
nmy-2(ts) (non-muscle myosin II), we found that eight enhance embryonic lethality when knocked-down in a gain of function mutant of
act-2 (actin) and seven out of these eight enhancers show defects in the actomyosin cytoskeleton during the first cell division. In collaboration with Stephan Grill we are using biophysical methods to characterize actomyosin dynamics upon loss of function of these candidates. We are using a similar approach to characterize the suppressors of
par-2(ts) and
pkc-3(ts). Functional antagonism between PAR-2 and PKC-3 is critical for C. elegans asymmetric first cell division and robustly found in our screens. Therefore, we are studying the enhancement phenotype of
par-2 suppressors in
pkc-3(ts) and vice versa. In another strategy, we have been using patterns of genetic interactions to predict functions for novel genes in the network. This analysis identified several potential new components and/or regulators of the anaphase-promoting complex (APC). In addition, the network functionally implicates different signalling pathways in polarity. For example, we have identified a role for PKA signalling in the regulation of the asymmetric first cell division, which we are currently studying.
Due to the conservation of cell polarity mechanisms, we expect that the C. elegans polarity network we have built will be relevant for cell polarity events in other model organisms.