Worm Breeder's Gazette 10(1): 38
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.
Two independent approaches to the study of muscle genes have
converged to demonstrate that the sup-10 gene encodes a regulatory
myosin light chain (MLC) protein. Sup-10, described by I. Greenwald
and R. Horvitz (1986, Genetics 113: 63-72), is a member of a group of
interacting genes involved in muscle structure and function. These
genes include unc-93, sup-10,
500) and sup-10(n983) are
rare alleles that confer a characteristic uncoordinated ('rubber band')
muscle defective phenotype. Null alleles of unc-93,
sup-10 and sup-18 are phenotypically wild-type and
act as recessive suppressors of unc-93(e1500) and sup-10(n983). The
wild-type null phenotype for four of the five genes and the patterns
of cross-suppression suggest that these genes are members of a multi-
gene family.
We have been using transposon tagging to clone four of these genes.
Alleles of sup-10, sup-9 and sup-18 were isolated
from strains with unc-93(e1500) or sup-10(n983) in a mut-2(r459)
background (see Levin and Horvitz, 1987 C. elegans meeting abstracts).
We have identified 27 suppressor mutations: eight alleles of sup-9,
13 alleles of unc-93, one sup-18 allele and five alleles of sup-10.
We have shown that one of the unc-93 alleles (e1500n1415) has a very
closely linked Tc1 insertion. Of recombinants in the interval between
daf-2 and dpy-17 (7mu), the extra Tc1 is present in all nine unc-93 (0)
recombinants and absent in all 17 unc-93 (+) recombinants (p<0.7mu to
the left and p<0.8mu to the right within 95% confidence limits; note
that this gene is on an arm). We have cloned a 7 Kb EcoRI restriction
fragment containing this Tc1 into a phage vector. We have examined
one sup-9 allele and found no linked Tc1, Tc3 or Tc4. Other
backcrossed sup-9 alleles are being examined.
The C. elegans 'regulatory' MLC gene family consists of two gene
copies mlc-1 and mlc-2, that are separated by 2.7 Kb. (see Cummins
and Anderson, 1987 C. elegans meeting abstracts). Cosmid clones that
cover mlc-1 and mlc-2 were identified by J. Sulston and A. Coulson.
In situ hybridization of these clones to metaphase chromosomes
indicated that these genes are located near the right end of the X
chromosome. To define more precisely the position of the mlc-1 and
mlc-2 genes, a mlc-1/mlc-2 plasmid clone was used to probe Southern
blots of total genomic DNA from strains heterozygous for deficiencies
in this region. Hybridizations were quantitated and the results
indicated that mlc-1 and mlc-2 are located within the deletion
interval defined by mnDf8 and mnDf42. sup-10 and nine other genes
have been mapped within this interval. These results suggested the
possibility that sup-20 encodes a myosin light chain protein.
To test whether sup-10 encodes a myosin light chain protein, we have
screened for MLC gene alterations in sup-10 mutants. A mlc-1/mlc-2
clone was used to probe Southern blots of total genomic DNA from 13
sup-10 alleles. Two of five sup-10 alleles isolated in a mut-2
background and three of eight gamma ray-induced alleles (seven
isolated by I. Greenwald) exhibit altered Southern blot patterns (two
insertions, one deletion, one apparent tandem duplication and one
complex rearrangement). Each of the rearrangements affects the mlc-1
gene but not the mlc-2 gene.
The C. elegans 'essential' MLC gene family consists of at least two
members, only one of which has been cloned. The cloned gene, mlc-3,
was mapped by in situ hybridization to linkage group III, 34-41% from
one end. Both unc-93 and sup-18 are located in this region. We have
examined 11 unc-93 alleles (nine isolated in a mut-2 background,
including unc-93(e1500n1415) and two gamma ray-induced alleles). All
of these have a wild-type hybridization pattern when probed with a
cosmid that includes the mlc-3 gene. Thus, we think it unlikely that
unc-93 encodes the mlc-3 gene product, but we will examine additional
unc-93 alleles. We are currently investigating whether sup-18 is mlc-
3 and whether unc-93, nd to other
members of the C. elegans 'essential' MLC gene family.