Worm Breeder's Gazette 15(3): 31 (June 1, 1998)

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.

ego-6 is a member of a novel gene family

Anne Smardon, Kimberly Kelly, Eleanor Maine

Dept. of Biology, Syracuse University, Syracuse, NY 13244

        We are characterizing a number of ego (enhancer of glp-1) genes
at the molecular level.  Two genes, ego-1 (Qiao et al. 1995) and ego-6
(J. Spoerke, M. Klein, and E. Maine, unpublished), map between goa-1 and
gld-1 (most likely to the right of lrp-1) and show intergenic
non-complementation (i.e., ego-6 +/  + ego-1 animals are mutant).  The
only genetic evidence to suggest that ego-6 and ego-1 are different
genes is deficiency mapping data; two independently derived chromosomal
deficiencies, nDf25 and mnDf111, uncover ego-1 alleles (om18, om71) but
not ego-6 alleles (om54, om58, om84, om96, om97, om119).  
        ego-6 mutations cause a variety of germline defects including
enhancement of glp-1(ts) and lag-1(ts), premature onset of meiosis, slow
progression through the early meiotic "transition zone", and various
gametogenesis defects.  ego-1 mutations cause a similar, but not
identical, phenotype.  ego-6 +/  + ego-1 transheterozygotes resemble
ego-6 mutants.  
        To determine whether ego-6 and ego-1 indeed are different genes
and to better understand their role(s) in germline development, we have
begun molecular studies.  Using cosmids spanning the goa-1 to gld-1
region, we looked for DNA rearrangements associated with ego-6 alleles. 
Using cosmid F26A3 as a probe, we detected a deletion of ~300 bp
associated with the UV-induced ego-6(om84) allele.  It maps to the
vicinity of the predicted transcription unit, "F26A3.3".  No other RFLPs
were detected with any other ego-6 allele or cosmid probe.  Earlier
studies using cosmids spanning the lrp-1 to gld-1 region failed to
detect any polymorphisms associated with ego-1 mutations (S. Stacey and
E. Maine, unpublished data).  
        We isolated cDNAs spanning the F26A3.3 region and carried out
RNA blots with several subfragments of F26A3 as probes.  The GeneFinder
prediction for this region was fairly accurate, but incorrectly assigned
two separate genes to one transcription unit.  Interestingly, the two
genes are structurally related and encode "novel" proteins that are ~58%
identical at the amino acid level (as predicted from cDNA sequences). 
Based on our studies, the om84 deletion interrupts the coding region of
the upstream gene; we presume that this gene is ego-6.  In support of
this conclusion, ego-6 can be phenocopied by injection of RNA made from
a partial cDNA corresponding to the upstream gene.  C. elegans appears
to contain at least one other relative based on GeneFinder predictions
(on cosmid F10B5 on LG II); a loosely related gene is predicted by the
S. pombe genome project.  
        In-progess experiments aim to (1) determine determine whether
any existing ego-6 allele is a null; (2) investigate whether or not
ego-1 and ego-6 are in fact the same gene; (3) examine the
tissue-specificity of ego-6  expression; (4) examine the function of the
downstream, ego-6-related gene.  To examine (1), we are amplifying and
sequencing the ego-6  gene from ego-6 mutant strains.  In particular, we
are interested in knowing whether the deletion in om84  shifts the open
reading frame.  To investigate (2), we are amplifying the ego-6 gene
from ego-1 mutant strains to determine if it contains mutations
associated with the ego-1 alleles.  To investigate (3), we are examining
glp-4(bn2ts) mutants (raised at 25!C) for the presence of ego-6 RNA. 
Based on its mutant phenotype, we suspect that ego-6 might be expressed
specifically in the germ line.  To examine (4), we are carrying out RNA
interference experiments.  Also, we are amplifying and sequencing this
gene from ego-1 mutant strains to determine whether it might in fact
correspond to ego-1.