Ryanodine receptor (RyR) is a calcium release channel that is regulated by many molecules such as calcium, calmodulin, ATP and others. In C. elegans , only a single gene,
unc-68 , encodes the CeRyR. Isolated
unc-68 mutant animals are weak Unc and have a thin head even if these are null mutations;
e540 (splice-acceptor mutation),
x14 (Trp153 to stop). This is contrast to that there are three isoforms in mammals and RyR knockout mouse is embryonic lethal. The
kh30 mutant animal, isolated as a ketamine-response abnormal phenotype, causes an amino acid substitution at Ser1,444 to Asn of CeRyR. The Ser1,444-surrounding region having possible PKC phosphorylation sites and high hydrophilicity was useful to raise a specific antibody which stained CeRyR in situ (see Hamada et al., this meeting poster). This region could interact with other molecules and thus named as "KH30 domain". We aimed to isolate novel regulatory proteins of CeRyR by using the yeast two-hybrid screening. Using the KH30 domain of CeRyR as a bait, an ERM (ezrin/radixin/moesin) family gene,
erm-1 , was isolated. ERM-1 was most similar to the vertebrate moesin that crosslinks membrane and cytoskeleton.
erm-1 was mapped on chromosome I, and encoded several isoforms under the control of two promoters and an alternative splicing. By using two-hybrid analysis, we found that only the truncated form of ERM-1 could interact with the KH30 domain of CeRyR, and that the alternative form having three amino acid deletion weakened the interaction.
erm-1::gfp fusion genes were expressed in excretory canals, intestine, pharynx and possibly in body wall muscles. These results suggest that ERM-1 may be a novel regulator of CeRyR and ERM-1/CeRyR interaction could be regulated by cytoskeletal re-organization during development. We are also analyzing in vivo function of ERM-1 proteins by RNAi experiment.