Suppression in transformed avian fibroblasts of a gene (CO6) encoding a membrane protein related to mammalian potassium channel regulatory subunits.

ABSTRACT

Gene expression patterns in normal and v-myc-transformed quail embryo fibroblasts were compared by mRNA differential display. Displaying approximately 2500 mRNA species by reverse transcription/PCR, reamplification of 73 differential cDNA fragments and rescreening by Northern analysis led to the isolation of a clone, termed CO6, that hybridized to an mRNA species present only in the normal but not in the transformed fibroblasts. Further analyses revealed that the 0.95-kb CO6 mRNA was present in all normal quail and chicken embryo fibroblasts tested, but that it was undetectable in a variety of established quail cell lines transformed by the v-myc, v-myc/v-mil, v-jun/junD or v-src oncogenes or by a chemical carcinogen. Furthermore, CO6 mRNA was not detectable in fibroblasts newly transformed by retroviral constructs carrying v-myc or v-jun alleles or by the avian sarcoma virus ASV17. In fibroblasts transformed by a temperature-sensitive v-src mutant, expression of CO6 was strongly induced at the non-permissive temperature and reduced at the permissive temperature. Nucleotide sequence analysis of quail CO6 cDNA indicated that the corresponding gene encodes a 200-amino acid protein with 46 to 48% amino acid sequence identity to the regulatory beta subunits (K(VCa)beta) of the bovine, human and canine high conductance Ca2+-activated K+ channels. No sequence homology to other ion channel subunits or to any other proteins in the databases was found. Like the K(VCa)beta subunits, the CO6 protein contains two putative transmembrane segments. Based on the relationship to mammalian K(VCa)beta both in primary structure and domain topology, the CO6 protein may represent the regulatory subunit of a yet unidentified avian Ca2+-activated potassium channel or a related membrane protein possibly involved in the regulation of cell proliferation.