Worm Breeder's Gazette 12(3): 42 (June 15, 1992)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
sup-10 is a member of a group of interacting genes that includes unc-93 ,sup-g, sup-10 ,sup-ll and sup-18 (Greenwald and Horvitz, 1980, 1982, 1986). Rare altered function alleles of unc-93 , sup-9 ,and sup-10 confer a distinctive, muscle defective phenotype. When touched on the head, a mutant worm quickly contracts and then relaxes without moving backward as would a wild-type worm. This has been termed a "rubber band" phenotype. The regulation or coordination of muscle contraction appears to be defective in these animals. Null mutations in unc-93 , sup-9 , sup-10 and sup-18 are wild-type in phenotype on their own and act as recessive suppressors of the rubber band mutations (Greenwald and Horvitz, 1980, 1982, 1986). The unc-93 gene encodes a putative transmembrane protein (Levin and Horvitz, 1992, J. Cell Biol. 117:143). Levin and Horvitz speculate that the unc-93 protein might be a component of an ion transport system involved in excitation-contraction coupling in muscle or might be involved in the coordination of muscle contraction, perhaps by affecting the functioning of gap junctions.
We have previously reported that sup-10 and the regulatory myosin light chain (MLC) genes, mlc-1 and mlc-2 ,are linked on the X chromosome. In collaboration with Donna Albertson and Joshua Levin, we mapped mlc-1 ,2to the right end of the X chromosome and demonstrated that a mlc-1 ,2hybridization probe detected alterations in 6 of 40 sup-10 mutant strains originally tested. Initially, we predicted that sup-10 was located within a short distance of the MLC genes. Localization of the sup-10 gene by transformation rescue indicates that, in fact, sup-10 is approximately 60kb rightward of the MLC genes. (The left-to-right gene order is mlc-2 - mlc-1 - sup-10 ,defined with respect to the physical map.)
We analyzed a number of sup-10 mutant DNAs on Southern blots looking for clues as to the precise position of the sup-10 gene. We detected blot phenotypes for 18 sup-10 mutants. Fifteen contain large homozygous viable deletions. For example, sup-10 ( n184 )removes most of mlc-1 and extends rightward an estimated minimal distance of 180 kb. The remaining three sup-10 mutants for which we have detected blot phenotypes were isolated in a mut-2 background and contain Tc1 element insertions at three distinct positions within a region of approximately 350 bp. No other alterations have been observed for these 3 alleles. The leftward endpoint of the most distal of the deletions that confer a sup-10 phenotype is located a short distance to the right of these insertions. It is likely that the Tc1 insertions and this deletion endpoint are located within the sup-10 gene.
We have achieved transformation rescue of sup-10 in two ways. In both cases, we injected a cosmid, F57C11 , that flanks the sites of Tc1 insertion and the deletion endpoint to the right of the insertions. First, we rescued the suppression phenotype of sup-10 .We coinjected F57C11 cosmid DNA with the rol-6 dominant plasmid into an unc-93 ( e1500 ); sup-10 ( n184 )strain. This strain has a wild-type phenotype. e1500 is a rubber band allele of unc-93 . n184 completely deletes the sup-10 gene. We reasoned that since suppression of e1500 by n184 is recessive, expression of a wild-type sup-10 gene in a transformed animal should result in a rubber band phenotype. We have obtained a heritably transformed line of rubber band rollers that are Unc and completely egg-laying defective as are e1500 animals. Loss of the array results in the segregation of wild-type non-roller animals. Second, we rescued the recessive rubber band phenotype of sup-10 ( n983 ).We coinjected F57C11 cosmid DNA with the rol-6 dominant plasmid into a sup-10 ( n983 )strain. We have obtained a heritable line of non-rubber band rollers indicating rescue of n983 .These transformants segregate non-roller rubber band animals when the array is lost.