GH3 cell-specific expression of Kv1.5 gene. Regulation by a silencer containing a dinucleotide repetitive element

Abstract

A silencer element (Kv1.5 repressor element; KRE) was characterized by deletion analyses in the promoter of Kv1.5, a voltage-gated potassium channel. The silencer element selectively decreases expression of Kv1.5- and thymidine kinase-chloramphenicol acetyl-transferase reporter gene constructs in cell lines that do not express Kv1.5 polypeptide. It contains a dinucleotide repetitive element (poly(GT)19(GA)1(CA)15(GA)16), and self-associates spontaneously in vitro to form complexes with slow electrophoretic mobility. Deletion of the repetitive element abolished self-association in vitro and the silencing activity in transient transfection experiments in vivo. Electromobility gel shift assays of KRE with GH3 cells nuclear extracts detected the formation of a unique DNA-protein complex, which was not detectable in Chinese hamster ovary and COS-7 cells. This complex does not react with an antibody against nonhistone high mobility group 1 protein, which binds KRE in gel retardation assays. These observations establish that a dinucleotide tandem repeat sequence, capable of self-association, forms part of a cell-specific silencer element in a mammalian gene.