GH3 cell-specific expression of Kv1.5 gene. Regulation by a silencer containing a dinucleotide repetitive element.
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.
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D006224:Cricetinae; D017931:DNA Primers; D015870:Gene Expression; D051666:Kv1.5 Potassium Channel; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D015221:Potassium Channels / Q000096:biosynthesis* / Q000235:genetics*; D024642:Potassium Channels, Voltage-Gated; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014162:Transfection
D017931:DNA Primers; D051666:Kv1.5 Potassium Channel; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011993:Recombinant Fusion Proteins; D015500:Chloramphenicol O-Acetyltransferase