In response to a signal from the centrosome, A-P polarity is established shortly after fertilization in the one cell stage embryo. We reported previously that embryos lacking
zyg-11 function appear to use a signal from oocyte derived chromosomes to establish an inverted polarity during meiosis II. How
zyg-11 normally acts to prevent precocious polarity establishment is not understood.
ZYG-11 appears to function as a substrate recruitment subunit of a CUL-2 based E3 ligase to target substrates to proteasome destruction. This E3 ligase may have several substrates, based on the phenotypes observed after
zyg-11/cul-2 inactivation. First, such embryos have a delayed metaphase to anaphase transition and M phase exit, due in part to an increased levels in B-type cyclins. Second, such embryos establish an inverted polarity during meiosis II, a phenotype that can be dissociated from the cell cycle delays, indicating that
zyg-11 may have a distinct function and substrate for A-P polarity establishment. Third, such embryos fail to degrade OMA-1/OMA-2. Destabilization of these two proteins also requires phosphorylation by the two kinases MBK-2 and GSK-3 and is necessary for proper localization of P-granules.
To identify novel components that function together with the ZYG-11/CUL-2 E3 ligase, we conducted a genome-wide RNAi-based synthetic lethal screen with the temperature-sensitive mutant allele
zyg-11(
b2). We identified 294 candidate enhancers in this manner. Next, we sought to focus specifically on those genes that may participate in polarity establishment by assaying the synthetic lethality of these candidates with other conditional mutants. We retained those genes that also enhanced
par-2(
it5) or
mbk-2(
ne992) and excluded those genes that also enhanced two other conditional mutants not related to
zyg-11/cul-2 function. Conducting these positive and negative selections, we identified 5 enhancers of
zyg-11(
b2ts) and
par-2(
it5), 1 enhancer of
zyg-11(
b2ts) and
mbk-2(
ne992), 2 enhancers of
zyg-11(
b2ts),
par-2(
it5) and
mbk-2(
ne992). We are in the process of examining polarity markers and OMA-1 levels after inactivation of these candidates in the wild-type, as well as in sensitized background conditions, to better understand their mode of action. We expect that analysis of these genes will help us decipher how inverted polarity is established following
zyg-11/cul-2 inactivation, which may shed light on the mechanism by which A-P polarity is set up in the wild-type.