ISLET EXPRESSION OF RHOMBOTIN AND ISL-1 SUGGESTS CELL-TYPE-SPECIFIC EXPOSURE OF LIM-DOMAIN EPITOPES

The homeodomain protein Isl-1 and the proto-oncogene Rhombotin (a LIM- only protein), share a double zinc-binding LIM domain and have both be en implicated in neural and possibly endocrine development. Isl-1 is e xpressed in all endocrine cell-types of the islet of Langerhans while Rhombotin mRNA expression was reported in rat insulinoma cells. We hav e cloned and sequenced Rhombotin cDNA from rat insulinoma (99.4% ident ical to human and mouse sequences) and demonstrate that it is expresse d in normal islets, intestinal tissue, and testis, in addition to the brain; but absent in all other organs tested. Rhombotin mRNA is expres sed in phenotypically distinct islet tumours (alpha-, beta, and delta- tumours) at levels comparable to that of normal islets. Antisera raise d against two distinct epitopes contained within a short synthetic pep tide representing part of the N-terminal LIM domain of Rhombotin surpr isingly stain alpha- and delta-cells, respectively, on sections of rat pancreas. Rhombotin is undetectable by immunocytochemistry using LIM- domain antisera on intact monolayer islet tumor cells or transfected f ibroblasts while readily detectable when equipped with a FLAG epitope, as detected with FLAG antiserum. In contrast, recombinant FLAG-Rhombo tin is efficiently recognised by Western blotting or immunoprecipitati on with all LIM-specific antisera. Almost identical results were obtai ned with LIM-specific versus homeodomain/C-terminal Isl-1 antisera sta ining alpha-cell cytoplasm or all islet nuclei, respectively. We concl ude that Rhombotin in addition to Isl-1 is expressed in the islet of L angerhans and propose that the differential staining patterns obtained with antisera towards the LIM domains versus flanking epitopes of bot h proteins reflect (1) cell-specific protein-protein interactions of t hese domains or alternatively, (2) islet cell type specific expression of novel homologous LIM domain proteins.