Spatial Pattern of Expression of Cyl Actin-β-Galactosidase Fusion Genes injected into Sea Urchin Eggs.

Abstract

The Cyl actin gene of the sea urchin Strongylocentrotus purpuratus displays a pattern of expression that is correlated with cell division; the gene is initially activated in all cells of the early blastula stage embryo, but after 18 hours Cyl actin transcripts disappear from the aboral ectoderm at a time when these cells are withdrawing from the cell cycle. As part of our investigation of the transcriptional regulation of Cyl, we tested various Cyl-β-gal fusion genes for their spatial pattern of expression by microinjection into fertilized eggs of the sea urchin, Lytechinus pictus. The plasmid Cyl-β-gal containing 2.5 kb of upstream sequence displayed a pattern of expression which reflected the endogenous Cyl actin gene. Most of the staining was seen in the gut, oral ectoderm, and mesenchyme cells, while fewer embryos had stained cells in the aboral ectoderm cells. Deletions in the upstream sequences to-195 bp resulted in a decrease in the mean number of stained cells, but the overall pattern of expression was similar to Cyl-β-gal. These results indicate that sequences required for correct spatial expression of Cyl are located in the first 195 bp upstream of the Cyl start of transcription and/or within the first intron. A mutation in the highly conserved serum response element and an adjacent protein binding site did not increase the percentage of aboral ectoderm staining nor decrease the relative level of CAT activity from a Cyl-CAT fusion gene containing the same mutation. The implications of these rsults are discussed.