Worm Breeder's Gazette 14(1): 63 (October 1, 1995)
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.
Dept of Embryology, Carnegie Institution of Washington, 115 W. University Pkwy, Baltimore, MD 21210
Mutations in let-858 result in embryonic lethality with arrest
occurring at the 2-3 fold stages. We have cloned and sequenced the
minimal genomic fragments capable of rescuing this lethality and have
determined that let-858 encodes a 2.8 Kb mRNA that appears to be enriched
in the germline and gives rise to a ~90 kD protein. Interestingly, while
the genomic constructs can rescue the lethality of let-858 alleles, the
rescued animals grow to sterile adults with a variety of germline defects.
Given the apparent germline enrichment for the let-858 mRNA, it is likely
that this sterility reflects the inefficiency of expression of, and hence
poor rescue by, extrachromosomal arrays in the germline.
Database searches for proteins similar to LET-858 have identified a
human cDNA with an 85% similarity (65% identity) over a stretch of 110
amino acids. Of course, as these things usually go, the cDNA is for a
protein of unknown function. In addition, an Arabadopsis cDNA (with the
same amount of information known as the human protein) also shares
significant similarity with let-858 (66% similar, 46% identity). We feel
that this is yet another case where C. elegans. can be used as a model for
simpler systems, such as Arabadopsis and H. sapiens.
We have constructed an in-frame fusion inserting GFP coding sequence
into let-858 and have found it to be ubiquitously expressed in all somatic
cell-types, and that it is localized to the nuclei of these cells. This
construct, like the native genomic construct from which it was derived,
also rescues the lethality of mutant alleles, but not the sterility
defect. This correlates with the absence of GFP fluorescence in the
germline of rescued animals. Recent attempts to achieve efficient
germline expression of this construct using a modified transformation
protocol have met with some success (see next abstract), and we can now
strengthen the correlation between germline expression of let-858 and the
rescue of fertility: in all cases where germline expression of
let-858::gfp were observed in mutant animals, the animals were rescued for
both the lethality and sterility defects. We conclude from these results
that let-858 encodes a protein that has essential functions in both the
soma and the germline. We are currently raising antibodies to confirm
that it provides these functions in the nucleus.