Worm Breeder's Gazette 13(1): 53 (October 1, 1993)

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.

Investigating the vets

Jonathan Pettitt, William B. Wood

Department of MCD Biology, University of Colorado, Boulder, CO 80309

We have given the name vet (very early transcript) to a set of genes originally identified by Irene Schauer(1) in molecular screens for genes that were preferentially transcribed in pre-gastrulation embryos. Irene isolated sixteen vet genes, eight of which have been placed on the physical map. Sequence analysis of the six vet genes for which cDNA clones exist ( vet-1 to -6) revealed that only one gene ( vet-1 ) had any similarity to other sequences in the Genbank databases. We are extending the sequence analysis of these genes and have determined the entire coding sequences for vet-1 and vet-6 ,both of which are trans-spliced with SL1 .This work has established that vet-1 encodes a product which is likely to be almost entirely coiled-coil in structure, whereas vet-6 appears to encode a novel protein.

We have begun to characterize the spatial expression patterns of selected vet genes, in order to establish which ones, if any, show lineage specific expression patterns. We are beginning by making lacZ constructs of the six vet genes which we know most about. The results with our first lacZ fusion construct showed that at least one vet gene has a spatially restricted expression pattern. This fusion construct (termed pVET6 B)consist of a 6 kb genomic DNA fragment, derived from the vet-6 gene, fused to the lacZ gene in Andy Fire's pPD22 .11vector. We have used four lines, each carrying an integrated array of pVET6 B,to study its expression pattern.

The original screen in which vet-6 was isolated was biased for highly transcribed genes, and pVET6 Bappears to be highly expressed as well, producing patterns that are apparent after only 15 minutes of staining with X-Gal. The fusion is expressed only relatively briefly and staining first appears at the 28 cell stage and disappears by the end of gastrulation. We have not yet established which cells express the fusion; however, the expression begins in a few cells and spreads to include nearly all the cells of 60-100 cell embryos. The staining pattern is reduced to a subset of these cells in subsequent divisions prior to its disappearance. The fusion protein appears to be cytoplasmic in all the cells that express it.

We are currently using Geraldine Seydoux's in situ hybridization protocol to try to confirm that the expression of pVET6 Bcorrelates with the presence of vet-6 transcripts. We have also made a lacZ fusion using a 3.5 kb genomic fragment derived from vet-1 ,but we have not been able to obtain any expression from this construct. Further lacZ fusion constructs using larger fragments from vet-1 ,as well as constructs for other vet genes are underway.

(1) Schauer and Wood (1990) Development 110,1303.