Because mutations in the neuronal syntaxin gene
unc-64 alter volatile anesthetic (VA) sensitivity, we have screened for
unc-64(rf) suppressors in hopes of identifying regulators of VA action. For different reasons, Owais Saifee in Mike Nonet's lab performed a similar screen. A total of 38,000 genomes were screened by the two labs, and 21 suppressor mutations were isolated. The suppressors were outcrossed from
unc-64(rf) and fell into two phenotypic classes loopy and jerky that cosegregate with
unc-64(rf) suppression. Two semidominant loopy suppressors mapped by their behavioral and VA resistant phenotypes to chromosome IL near
egl-30.
egl-30 was sequenced in the mutants, and one of the alleles
js126 was found to have a missense mutation in the coding sequence. We did not find an
egl-30 mutation in the other strain.
egl-30(
tg26gf) , kindly given to us by K. Iwasaki, was also loopy and resistant to VAs. Testing of strains transformed with a constitutively active
egl-30 array (thanks to Carol Bastiani - Sternberg lab) were also resistant to VAs confirming that
egl-30(gf) could indeed produce VA resistance. Furthermore, we treated N2 with phorbol-ester, which activates diacyl glycerol-binding proteins and effectively enhances EGL-30 signaling; treated worms were resistant to VAs. Finally, we found that
egl-30(
js126) ,
egl-30(
tg26) , the
egl-30(gf) array-containing strains, and phorbol ester treated strains were all aldicarb hypersensitive. Aldicarb hypersensitivity suggests that presynaptic release of acetylcholine is increased. Thus, consistent with our previous results for
goa-1(rf) and
unc-64(rf) that suggested a presynaptic mechanism of VA action,
egl-30 regulates VA sensitivity, presumably by controlling transmitter release. Currently, we are constructing double mutants between various VA resistant and hypersensitive strains to place
egl-30 in the VA sensitivity pathway.