Structural and functional analysis of the mouse mdr1b gene promoter.
The overproduction of P-glycoprotein, an integral membrane protein thought to function as a drug efflux pump, is the hallmark of the multidrug resistance phenotype. In murine multidrug resistant J774.2 cell lines, distinct mdr genes, mdr1a and mdr1b, encode unique P-glycoprotein isoforms. To examine the transcriptional regulation of the mdr1b gene, its promoter was isolated and characterized. The transcription initiation site was mapped by primer extension, and the 5'-flanking region was sequenced. Several potential regulatory elements were identified in this region. A transient expression vector was constructed by fusion of 540 base pairs of 5'-flanking sequence and part of the first untranslated exon to the chloramphenicol acetyltransferase (CAT) gene. When transfected into monkey kidney COS-1, rat pituitary GH3 or T47D human breast cells, the mdr1b 5'-flanking sequences were capable of driving CAT expression. Transient transfection studies using deletion subclones of the mdr1b-CAT construct were done to locate potential cis-acting sequences. The studies indicate the presence of cis-acting elements in the 5'-flanking region of the mdr1b gene. The implications of these findings for expression and regulation of the mdr1b gene are discussed.
Cohen
D
D
Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, New York 10461.
Piekarz
R L
RL
Hsu
S I
SI
DePinho
R A
RA
Carrasco
N
N
Horwitz
S B
SB
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003001:Cloning, Molecular; D004351:Drug Resistance / Q000235:genetics*; D005091:Exons; D005786:Gene Expression Regulation; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D014158:Transcription, Genetic; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D008562:Membrane Glycoproteins; D015500:Chloramphenicol O-Acetyltransferase