Worm Breeder's Gazette 10(1): 26
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.
The mec-7 gene is required for the normal response to gentle touch in C. elegans. Mutations in mec-7 result in the loss of the large 15- protofilament microtubules normally present in and specific to the six touch receptor neurons (Chalfie and Thomson, 1982). Thus, study of the mec-7 gene should provide some insight into the processes by which microtubule structure is defined. Thirty-eight alleles of mec-7 X have been isolated; 36 by EMS mutagenesis and 2 from TR679. Each allele produces a characteristic phenotype at 25 C that can be classified as strong (completely touch insensitive) or weak (touch insensitive at the head but not at the tail) and that allows the mutations to be grouped in 5 classes: (a) strong recessive (8 alleles, including the 2 from TR679); (b) weak recessive (2); (c) weak semi-dominant (3; i.e. both the homozygotes and heterozygotes show the weak phenotype); (d) strong semidominant ( 20; i.e. the homozygotes have a strong phenotype, the heterozygotes a weak one); (e) strong dominant (5). The phenotypes of semi-dominant and dominant alleles show a temperature dependence: all of the heterozygotes (except n434/+, but see below) are more completely touch insensitive at 25 C than at 15 C. These results suggest that strong recessive alleles of mec-7 eliminate functional gene product, weak alleles encode abnormal products which partially interfere with touch cell function, and dominant alleles encode products which more strongly interfere with touch cell function. We have examined the effects of manipulating mec-7 gene dosage to further characterize the null phenotype and the dominant alleles. A gamma-ray induced deficiency of the region, uDf1, was generated as a non-complementing mutation of mec-7. This deficiency uncovers unc-6, unc-18 and mec-7 (as well as xol-1, L. Miller, personal communication), and does not uncover lon-2, dpy-6 and mec-10. Since mec-7[e1506(r)]/uDf1 animals are Mec, the deficiency does not increase the severity of the phenotype of a recessive mutation. This supports the hypothesis that touch insensitivity is the null phenotype of mec-7. In addition, uDf1/+ animals are phenotypically wild type, indicating that dominant mec-7 alleles are not the result of haplo insufficiency of this locus. To determine whether the dominant phenotypes are due to a gain of normal function or a gain of novel function, we constructed mec-7/+/+ strains using stDp2 and 4 different dominant alleles (e1527, n434, u129 and u162). In all cases, partial rescue of touch sensitivity was seen. The rescue of n434 was stronger at 15 C than at 25 C. These results suggest the dominant phenotypes of these alleles are due to expression of an abnormal gene product. Finally, to be sure that the recessive alleles of mec-7 thus far isolated represent null alleles, we have begun a screen for non- complementing mec-7 alleles. Five such alleles have been isolated from 3750 EMS mutagenized chromosomes. All five are X-linked, viable as homozygotes, and show no phenotypes other than touch insensitivity. These are likely to be mec-7 alleles (although the possibility that they are X-linked dominant mutations in other genes has not yet been eliminated). Several approaches to the cloning of mec-7 have been explored. Two alleles of mec-7 have been isolated from TR679 (by Shai Shaham and Peggy Brickman in our lab). Both alleles revert in the mut-2 background using Mike Finney's HH*10 strain. Southern analysis of DNA from these strains and from recombinants suggests that neither of these alleles is due to the insertion of Tc1, Tc3, Tc4, or the new element isolated in P. Anderson's lab (Tc5?). This analysis did reveal a Tc1-containing restriction fragment in one of the two mutant mec-7 strains which maps within 0.02 mu to the left of mec-7. This fragment could be used to initiate a chromosomal walk or to identify a contig containing mec-7.As we were doing these experiments, Donna Albertson told us about a -tubulin clone isolated by Linda Gremke and Joe Culotti ( 4) that mapped by in situ hybridization to the general region around mec-7. Using a lambda clone containing the 4 sequences, we found restriction fragment length differences in DNA from two EMS induced mec-7 mutants. The size difference for one of these mutations was mapped and found to be inseparable from the mec-7 defect. The 4 probe, however, did not reveal any differences in the TR679-derived strains. Thus, the mec-7 gene may be very close to or at this - tubulin site. We are analyzing the contig containing tubB4 to determine whether it contains the mec-7 gene.