Cloning of the sequences expressed abundantly in established cell lines: identification of a cDNA clone highly homologous to S-100, a calcium binding protein

Abstract

To identify and characterize mRNAs which are abundant in established cell lines, but less so in the corresponding parental counterparts, a cDNA library was constructed from poly(A)+RNA of Balb/c3T3 cells. We screened this cDNA library by differential colony hybridization using single-stranded cDNAs from Balb/c3T3 cells and mouse embryo fibroblasts (MEF), and obtained four cDNA clones, out of 8,300 clones. One such clone, named pEL98, corresponding to mRNA of approximately 600 nucleotides, was further characterized. The entire nucleotide sequence of the cDNA insert in the pEL98 revealed a single open reading frame that encodes a 101 amino acid of putative protein (hereafter referred to as the pEL98 protein). pEL98 protein is highly homologous to both alpha and beta subunits of bovine S-100 protein (49 and 47% homology, respectively), a protein belonging to the family of calcium binding proteins. pEL98 protein is also homologous to several S-100 related proteins such as "calcyclin," a product of growth-regulated human gene 2A9, porcine p11 (or p10), the small subunit of a protein complex which serves as a major substrate of tyrosine kinase, and human cystic fibrosis antigen (CFAg). The amounts of transcript of pEL98 were increased in murine cells transformed with both activated oncogene and chemical carcinogen.