Territorial expression of three different trans-genes in early sea urchin embryos detected by a whole-mount fluorescence procedure

Abstract

We have developed a new procedure for detection of the protein product of chloramphenicol acetyltransferase (CAT) reporter genes in whole mounted sea urchin embryos. The position of a commercially available anti-CAT antibody is visualized by video or confocal microscopy, and thus the spatial domains of exogenous reporter gene expression can be determined with regard to the intact three-dimensional structures of the embryo. We show that in pluteus stage embryos CAT protein expression patterns for SM50·CAT or CyIIIa·CAT reporter genes are similar to those previously obtained by in situ hybridizations with radioactive probes. Taking advantage of the superior resolution of cellular CAT expression patterns using the antibody visualization method, we found for the first time that, in addition to the expression in aboral ectoderm, some cells in the ciliated band of the pluteus express CyIIIa·CAT. The expression of a new fusion construct, CyIIa·CAT, was also examined. As expected from the localization of endogenous CyIIa mRNA, CAT protein was expressed under control of the CyIIa promoter in gut and skeletogenic mesenchyme cells.