Worm Breeder's Gazette 14(5): 61 (February 1, 1997)

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.


Caroline A. Spike, Robert K. Herman

University of Minnesota, 250 Biological Sciences Center, 1445 Gortner Ave, St. Paul, MN 55108.

     mec-8 (ref. 1) encodes a protein with two RNA recognition motifs 
(RRMs), characteristic of RNA binding proteins, and regulates the 
accumulation of a specific subset of alternatively spliced transcripts 
of unc-52, which encodes, via alternative splicing of pre-mRNA, a set of 
basement membrane proteins that are homologs of perlecan and that are 
important for body wall muscle assembly and attachment to basement 
membrane, hypodermis and cuticle2.  mec-8 and unc-52 interact 
genetically: mec-8; unc-52(viable) double mutants exhibit a paralyzed 
arrest at the twofold stage of embryogenesis, which is characteristic of 
unc-52(null) mutants.  unc-52(viable) mutations are in a region of unc-
52 pre-mRNA that is alternatively splicing in a mec-8-dependent manner.
     mec-8 mutants exhibit additional defects, including defects in 
mechanosensation and chemosensation, that cannot be attributed to 
effects on splicing of unc-52 transcripts.  It has therefore been 
proposed that mec-8 protein regulates the processing of transcripts from 
other genes.  Indeed, two such target genes appear to be mec-8 itself 
(E. Lundquist, pers. comm.) and mec-2 (M. Huang and M. Chalfie, pers. 
comm.).  We do not know how many different targets MEC-8 may have.  We 
decided to explore the possibility that MEC-8 may overlap in function 
with one or more other proteins and designed a screen to look for 
mutations that would be synthetic lethal with a mec-8 loss-of-function 
     We established a mec-8(u74) I; unc-36 III; mnEx52[mec-8(+) unc-
36(+)] stock.  Progeny carrying the extrachromosomal array mnEx52 are 
rescued for mec-8 mutant phenes as well as for the uncoordination 
conferred by unc-36.  Many progeny do not inherit mnEx52 and are, as a 
consequence, (Mec) Unc-36.  We treated the array-bearing stock with EMS 
and looked among the broods of F2 hermaphrodites for the absence of Unc-
36 animals.  We found one mutant in a small-scale screen (equivalent to 
less than 300 haploid genomes screened) and confirmed that it carries a 
mutation, mn601, that is synthetic lethal with mec-8, by replacing 
mnEx52 in the mutant line with mnEx2[mec-8(+) rol-6(d)] to give a stock 
that is mec-8; unc-36; mn601; mnEx2.  These animals, which are Unc-36 
non-Mec rollers, are incapable of segregating Unc-36 Mec non-rollers 
because mec-8; mn601 is lethal.
     We mapped mn601 by mating mec-8 males with mec-8 I; dpy-8 unc-
3/mn601 X hermaphrodites and scoring the male progeny.  Males hemizygous 
for mn601 die.  Indeed, among 1907 male progeny, only one was non-Dpy 
non-Unc (and probably arose as a consequence of nondisjunction in the 
mother).  We conclude that the penetrance of the synthetic lethality is 
close to 100%.  We mapped mn601 1.0 map unit left of unc-3 (and 21.8 map 
units right of dpy-8) and have balanced it with mnDp1(X;V) in a mec-8; 
mnDp1/+; mn601 strain.  The synthetic lethal arrest is primarily at 
about the twofold stage of elongation, so it is possible that the gene 
defined by mn601 can affect the alternative splicing of unc-52 or some 
other muscle-affecting gene.
     The phenotype of animals homozygous for mn601 by itself is not 
dramatically different from wild-type.  Perhaps additional genes that 
interact with mec-8 can be identified through the identification of more 
synthetic lethal mutations.  Previous work3 identified mutations in smu-
1 I and smu-2 II that suppress mec-8 mutations -- as well as certain 
unc-52 mutations.  We are working to clone one or more of these mec-8-
interacting genes.

1Lundquist, E. A., et al., 1996 Development 122: 1601-1610.
2Rogalski, T. M., et al., 1993 Genes & Development 7: 1471-1484.
3Lundquist, E. A., and R. K. Herman, 1994 Genetics 138: 83-101