Receptor-mediated gene delivery approach demonstrates the role of 5'-proximal DNA region in conferring phenobarbitone responsiveness to CYP2B2 gene in rat liver in vivo.
The phenobarbitone (PB) responsiveness of the 5'-proximal region of the CYP2B1/B2 gene was examined in detail with plasmid DNA constructs containing G-free cassette as reporter, using in vivo targeting of the same DNA constructs into rat liver as galactosylated-polylysine complexes. The contribution of the proximal region (-1 to -179 bp) and the positive element (-69 to -98 bp) identified earlier in this laboratory to PB responsiveness was assessed. The results obtained on PB treatment of rats subjected to receptor-mediated gene delivery to liver were conclusive and dramatic, with the control (saline-treated) rats manifesting very little expression of the reporter, reflecting the in vivo picture of CYP2B1/B2 gene expression. The positive element conferred PB responsiveness to homologous and heterologous promoters. Deletion of the positive element led to elimination of PB response. The entire -179 bp region was significantly more effective in responding to PB treatment than the region up to -98 bp, both containing one copy of the positive element. Thus, the positive element and its flanking sequences in the 5'-proximal region are involved in conferring PB responsiveness to the CYP2B1/B2 gene.
Mani
S A
SA
Department of Biochemistry, Indian Institute of Science, Bangalore, 560 012, India.
Harish
S
S
Vathsala
P G
PG
Rangarajan
P N
PN
Padmanaban
G
G
D000818:Animals; D030801:Animals, Genetically Modified; D001189:Aryl Hydrocarbon Hydroxylases; D001483:Base Sequence; D019362:Cytochrome P-450 CYP2B1 / Q000235:genetics; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004247:DNA / Q000008:administration & dosage / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D017930:Genes, Reporter; D005822:Genetic Vectors; D008099:Liver / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D010634:Phenobarbital / Q000494:pharmacology*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D013250:Steroid Hydroxylases / Q000235:genetics*; D014162:Transfection
D017931:DNA Primers; D012333:RNA, Messenger; D004247:DNA; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; D019362:Cytochrome P-450 CYP2B1; C025619:steroid 16-beta-hydroxylase; D010634:Phenobarbital