Cloning of rat liver arginase cDNA and elucidation of regulation of arginase gene expression in H4 rat hepatoma cells

Abstract

In order to study the regulation of expression of the two arginase genes in mammalian tissues, we undertook to clone cDNA specific for rat liver arginase. mRNA was isolated from rat liver polysomes enriched for the arginase message by immunopurification and was used to produce an 800-member cDNA library carried in pBR322. Four arginase clones were identified by hybrid selection, and one was used to find two others following colony hybridization. Clonal identity was verified by its enrichment in the cDNA made from immunopurified mRNA; by hybrid selection, immunoprecipitation, and competition by purified arginase; hybridization on Northern analysis with liver-derived RNA (high in arginase) and its absence with mRNA from tissues low in arginase; and independent identification by hybrid selection and colony hybridization. Northern analysis of mRNA from H4-II-E-C3 (H4) rat hepatoma cells in which arginase activity was induced by hydrocortisone demonstrated equal, eightfold augmentation of both arginase activity and arginase mRNA levels. Southern blot analysis of DNA from these cells indicated that no change in arrangement or copy number accompanied induction. Southern analysis also suggested that the gene for rat liver arginase is present in a single copy, without pseudogenes, and that a high degree of homology exists between it and its mouse counterpart.