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 mutation. 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