Worm Breeder's Gazette 11(4): 56

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.

Transformation Rescue of mei-1, a Gene that Affects Meiosis and Mitosis

Shawna Clark-Maguire and Paul E. Mains

The one-cell C.  elegans embryo must support two distinct patterns 
of cell division.  Shortly after fertilization, the embryo undergoes 
the two meiotic divisions; and about twenty minutes later, the first 
mitotic cleavage occurs.  Essentially the same cytoplasm must support 
both the meiotic and mitotic spindles, which differ in many aspects, 
including location, morphology and chromosomal alignment.  Clearly, 
the activities of the gene products that govern the features that 
distinguish meiosis from mitosis are carefully regulated during this 
time.  We have identified maternal effect mutations in four loci that 
disrupt either the meiotic or the mitotic divisions.  The dominant ts 
gain-of-function (gf) mutations mei-1(ct46) I and mel-26(ct61) I and 
recessive loss-of-function (lf) mutations of zyg-9 II show similar 
defects in the first mitotic division, and these mutations are strong 
enhancers of one another's defects.  If mutations of mei-1 and mei-2 
result in recessive meiotic abnormalities but act as dominant 
suppressors of the mitotic defects of mei-1(ct46gf and mel-26(ct61gf).
We are focusing our attention on mei-1, which can be mutated to affect 
either meiosis or mitosis.  The molecular cloning of this gene has 
been straightforward, since it maps between lin-10 and lin-28, both of 
which have been located on the physical map.  A total of 14 
overlapping cosmids (provided by Alan Coulson and John Sulston) span 
the region, and we have used these to rescue the recessive maternal-
effect lethality of mei-1(lf) by germline transformation.
The individual cosmids were co-injected with a plasmid containing 
the dominant rol-6 mutation (described by Craig Mello) into mei-1(lf)
/+ hermaphrodites (with appropriate markers in cis), and transgenic 
lines were established.  Segregants homozygous for mei-1 that rolled 
were then tested for fertility.  Since in previous mapping experiments 
recombinants between mei-1 and either lin-10 or lin-28 were easily 
obtained, we started testing cosmids in the center of the region and 
worked our way out.  Naturally the rescuing activity was very near one 
end.  Two overlapping cosmids, T01G9 (which also includes lin-10) and 
F54A4 provide mei-1+ activity.  Injecting F54A4 directly into 
homozygous mei-1 hermaphrodites does not result in even transient 
We tested F54A4 cut with various restriction enzymes for rescuing 
activity in order to identify enzymes that did not cut within the gene,
as described by Kim and Horvitz [Genes Devel.  4: 357 (90)].  The 
SalI digest was positive.  Since only one of the four fragments 
produced by this enzyme was also cut by every enzyme that abolished 
rescuing activity, we could predict which SalI fragment should rescue. 
Being dubious about taking the (negative) results of failure to 
rescue too seriously we injected the other three gel-purified 
fragments anyway (since we can inject faster than we can analyze).  
However, only the predicted 10 kb SalI fragment contained mei-1+ 
activity.  We are repeating this strategy with additional enzymes to 
whittle the clone down to a more convenient size.