Mutation of the gene
spe-4 disrupts C.elegans sperm meiosis, and cells that appear by Nomarski optics to be spermatocytes actually contain four haploid nuclei. This sterile phenotype is also observed when either of two
spe-4 EMS alleles is placed in trans to the noncomplementing deficiency sDf6 (L'Hernault, S. et al 1988 Genetics 120:435). Two factor mapping places
spe-4 very close to
unc-13. Of 2, 951 F2 from
spe-4(
q347)
unc13(e1091)/++ F1 animals, only one nonSpe Unc recombinant was obtained. We probed cosmids from the continuous contig covering this region (extending right from
unc-13; cosmids courtesy of J. Sulston and A. Coulson) to Southern blots of genomic DNA prepared from each of the three
spe-4 nonconditional alleles balanced over sDf5. The cosmid C44E1 was discovered to identify an allele specific deletion of approximately 200 b.p. associated with the spontaneous
spe-4 allele,
q347 (provided by Tim Schedl), which arose on a Bristol chromosome 1. Southern anlaysis of the single nonSpe Unc recombinant obtained in the two factor mapping studies described above shows that the recombinant does not contain the 200 b.p. deletion, further supporting the association between this deletion and
spe-4. Labeled restriction fragments of the cosmid C44E1 further localize the
q347 deletion to a 2.5 kb Eco Rl fragment. This same EcoRI fragment hybridizes to an abundant 1.3 kb male specific RNA on total RNA male vs female Northerns (male= RNA from Mog
fem-3 (
q23gf); female= RNA from
fem-1(
hc17 If). No other RNA's have been detected in this 2.5 kb region. Three cDNAs have been isolated for
spe-4. Two of these cDNA's, which come from a developmentally staged library ( courtesy of J. Ahringer and J. Kimble), hybridize to the 1.3 kb male specific RNA. One cDNA hybridizes to the 200 b.p. deletion of
q347, suggesting that this allele is likely to be a
spe-4 null. The third cDNA has recently been obtained from a lambda-zap library (courtesy of R. Barstead and R. Waterston), and it appears nearly full length. The three
spe-4 cDNAs and genomic clones are being sequenced, (we are about half finished at this time) and transgenic rescue esperiments are currently being pursued using C44E1.