Mutations in the
unc-1 gene suppress the altered sensitivities of
unc-79 and
unc-80 animals to all anesthetics except ether. The various
unc-1 alleles are to different extents, themselves sensitive to ether. The
unc-1 alleles differ from each other phenotypically, (dominant coilers and recessive kinkers); and also in their ability to suppress mutations in
unc-79 and
unc-80 (coilers cannot suppress, while kinkers can). Moreover, the
unc-1 protein is required for the hypersensitivity of
unc-79 and
unc-80 animals to 1% halothane (a clinically relevant concentration) . Therefore, in an effort to understand the mechanism of action of volatile anesthetics we have characterized the
unc-1 gene. We previously reported the physical localization of the
unc-1 locus to the cosmid K03E6 which has been sequenced by the sequencing consortium. We now know that the
unc-1 gene encodes stomatinan integral membrane phosphoprotein. It has 3 domains: a short extracellular N-terminal head, a single trans-membrane domain and a large cytoplasmic C-terminal domain. Though its exact function is unknown, stomatin may act to support, activate and/or regulate an associated cation channel in a ball and chain! fashion. The coding region of
unc-1 consists of 6 exons. These define an ORF of 855bp encoding a protein of 285 amino acids with a M Wt. of about 31 kD. Northern blots of RNA from a mixed stage population of worms when probed with a cDNA fragment containing exons 2-6 revealed a single transcript of 1.2 kb (including a 3!UTR of 343 bp). RT-PCR experiments showed that the
unc-1 message is trans-spliced to an SL1 leader. We have sequenced a representative of each class of
unc-1 alleles, and identified mutations in all but one of them. All of the mutations identified lie in the putative regulatory (cytoplasmic) domain of the
unc-1 protein. We have also characterized the allele
fc51, isolated during a screen for transposon induced alleles of
unc-1. This allele in addition to having the same change seen in the
unc-1 allele
e114, also has a second lesion. It has a 2.3 kb deletion in the neighboring neurocalcin gene which is also present (though not entirely) on the cosmid K03E6 and transcribed in the opposite direction from
unc-1. Northern blots revealed that not only is the neurocalcin message absent in
fc51, but the level of the stomatin message is also drastically reduced. Moreover, while
e114 genetically behaves like a partial loss of function mutation in
unc-1,
fc51 behaves like the
unc-1 null allele
e580. So, we may have identified an enhancer like element which lies in this 2.3 kb region and controls the transcription of the
unc-1 gene. Expression studies of the
unc-1 gene are currently underway