sup-10 is a member of a group of interacting genes that are involved in muscle structure and function. These genes include
unc-93,
sup-9,
sup10, sup-ll, and
sup-18 (Greenwald and Horvitz, 1986, Genetics 113:63-72). We plan to exploit the dose linkage between
sup-10 and the two regulatory myosin light chain (MLC) genes, mlc-l and
mlc-2, to isolate the
sup-10 gene. In collaboration with Donna Albertson and Joshua Levin, we mapped
mlc-1,2 to the right end of the X chromosome and demonstrated that a
mlc-1,2 hybridization probe detected alterations in 6 of 40
sup-10 mutant strains tested. The 6
sup-10 mutations detected with a MLC probe include 2 deletions, a Tcl insertion, a probable tandem duplication, and two complex rearrangements. One of the deletions removes most of mlc-l (but not
mlc2) and extends at least 25 kb rightward of mlc-l. (The left-to- right gene order is
mlc-2-
mlc-1-
sup-10, defined with respect to the contig map.) The wild-type phenotype of this mutant demonstrates that the functions of
mlc2 are likely to be redundant to those of mlc-l. The second
sup-10 deletion removes only sequences to the right of mlc- l. We analyzed the gene adjacent to mlc-l with the expectation that it might correspond to sup-l O. The product of this gene is a novel integral membrane protein. The two
sup-10 deletions remove all of this gene, and the Tcl insertion maps within its 3'-untranslated region. The transcript of this gene is present in very low abundance in wild- type. The message is absent or altered in size in the strains carrying the two deletions, the Tcl insertion, and the two complex rearrangements. We have used dones containing this gene in an effort to rescue a
sup-10 mutant by germline transformation. Although multiple heritable lines (cotransformed with the
rol-6L' plasmid) have been established using plasmid, lambda, and cosmid clones, none are rescued for the
sup-10 phenotype. We now suspect that the
sup-10 gene is, in fact, located even further to the right of mlc-l and that loss- of-function mutations affecting the membrane protein gene have no phenotype. We suspect this because two
sup-10 mutants with no apparent defects in mlc-l,
mlc-2, or the membrane protein gene contain deletions that map to the right of the membrane protein gene. The sup- 10 mutant that contains a Tcl insertion within the membrane protein transcript also proves to contain a deletion of material further rightward. The two original
sup-10 deletions also extend into this region. We have extended our analysis to the end of the
mlc-1,2 cosmid contig. Unfortunately, part or all of
sup-10 may lie within or beyond the cosmid gap we have encountered. We are currently using YACs that bridge the gap in a variety of tranformation and walking experiments.