Characterization of the promoter region of the cystic fibrosis transmembrane conductance regulator gene.
To identify the transcription regulatory elements of the cystic fibrosis transmembrane conductance regulator (CFTR) gene, DNA fragments located in the 5'-upstream region were fused with the bacterial chloramphenicol acetyltransferase (CAT) reporter gene and transfected into various cell lines to test for promoter activity. The results of these studies suggested that there were at least two positive and one negative cisacting elements involved in CFTR transcription initiation. One of them was a proximal, positive element delimited by the 5' deletion constructs -226 base parts upstream of the transcription start site. This minimal promoter sequence (-226 to +98) alone seemed to be sufficient to direct cell-specific CAT expression. The sequences immediately upstream of -227, on the other hand, appeared to contain a negative regulatory element; inclusion of this sequence with the proximal element (e.g. a construct containing sequences -345 to +98) rendered the CFTR promoter inactive. This negative regulatory element could also suppress the activity of a heterologous promoter. In addition, the DNA transfection study suggested the existence of another positive regulatory element outside the CFTR promoter region examined, as the inability of this region (e.g. -658 to +98) to function in a CAT assay could be overcome by the presence of a viral enhancer element.
Chou
J L
JL
Department of Genetics, Hospital for Sick Children, Toronto, Ontario, Canada.
Rozmahel
R
R
Tsui
L C
LC
D000595:Amino Acid Sequence; D001483:Base Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D003550:Cystic Fibrosis / Q000235:genetics*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004247:DNA / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
C505032:CFTR protein, human; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D012313:RNA; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase