To further understand signaling via the heterotrimeric G protein alpha subunit Gas in C. elegans, a constitutively active Gas mutant was constructed (Korswagen et al., 1997). Expression of this constitutively active form of Gas results in hypercontraction of body-wall muscle cells and in induction of neuronal degeneration. Extragenic suppressors that inhibit the neuronal degeneration were identified in a screen using mutagenesis. All the suppressors tested so far fall into two complementation groups,
sgs-1 or
sgs-2 (suppressor of activated Gs).
sgs-1 has been cloned, and encodes an adenylyl cyclase that is most similar to mammalian adenylyl cyclase type IX (Korswagen et al., 1998). Currently, 23
sgs-1 alleles have been sequenced. The frequency and spectrum of the
sgs-1 mutations suggest that none of them represent a complete null allele of
sgs-1. Therefore, we isolated a Tc1 insertion derived deletion allele (
pk1279te) of
sgs-1. In this allele, a 2.9 kb region of the
sgs-1 sequence is deleted, removing the second transmembrane region and the second catalytic domain of
sgs-1. Animals homozygous for
sgs-1(
pk1279te) arrest during early larval development. This phenotype was rescued by introduction of an
sgs-1(+) containing plasmid. When constitutively active Gas is expressed, no neuronal degeneration is observed in homozygous
sgs-1(
pk1279te) animals. Our results indicate that
sgs-1 is an essential gene. Presumably, the
sgs-1 alleles that were found in the suppressor screen are reduction-of-function alleles, and not complete loss-of-function alleles. Currently, we are placing all
sgs-1 alleles in trans to the
sgs-1(
pk1279te) allele. Preliminary results indicate that most, but not all,
sgs-1 alleles are haplo-sufficient. Our second suppressor of the activated Gas induced neuronal degeneration,
sgs-2, has been mapped to chromosome V. Using visible markers and SNPs,
sgs-2 can now be placed in a 110 kb interval. The region is covered by 4 overlapping cosmids, which are currently being introduced in
sgs-2 animals to test for rescue. In addition, SNP mapping will be continued to refine the location of
sgs-2. References: H.C. Korswagen, J.-H. Park, Y. Ohshima, and R.H.A. Plasterk. (1997) An activating mutation in a Caenorhabditis elegans Gs protein induces neural degeneration. Genes Dev. 11, 1493-1503 H.C. Korswagen, A.M. van der Linden, and R.H.A. Plasterk. (1998) G protein hyperactivation of the Caenorhabditis elegansadenylyl cyclase SGS-1 induces neuronal degeneration. Embo J. 17, 5059-5065