Expression of gastrin, a gut hormone and growth factor, has tissue-specific transcriptional regulation and can be induced in some tumors. Previous studies have shown that a CACC cis-regulatory element is important for transcriptional activation in pancreatic insulinoma cells. To identify CACC-binding proteins, a lambda phage cDNA library derived from a rat insulinoma cell line, RIN 38A, was screened by a Southwestern method. A novel member of the Cys2-His2 zinc finger gene family was cloned and designated RIN ZF, having a cDNA sequence of 3.8 kilobases. One full-length and a shorter splice variant were sequenced and had predicted protein masses of 91.6 and 88.7 kDa. Expression of both splice forms were ubiquitous in fetal and adult rat tissues. Recombinant RIN ZF protein exhibited sequence-specific binding to the gastrin CACC element in a gel mobility shift assay. In transient transfections, both splice variants appeared to have only weak activating effects on gastrin-luciferase reporter gene transcription. Furthermore, RIN ZF coexpression with Sp1 appeared to block the strongly activating effects of Sp1 mediated through the CACC element. These findings suggest that a novel set of zinc finger proteins may help regulate gastrin gene expression by interfering with Sp1 transactivation.