The mitochondrial uncoupling protein gene in brown fat: correlation between DNase I hypersensitivity and expression in transgenic mice.
The mitochondrial uncoupling protein gene is rapidly induced in mouse brown fat following cold exposure. To identify cis-regulatory elements, approximately 50 kb of chromatin surrounding the uncoupling protein gene was examined for its hypersensitivity to DNase I. Seven DNase I-hypersensitive sites were identified in the 5'-flanking DNA, and one site was identified in the 3'-flanking DNA. Transgenic mice with an uncoupling protein minigene were generated by microinjection of fertilized eggs with a transgene containing 3 kb of 5'-flanking DNA and 0.3 kb of 3'-flanking DNA. Expression of the transgene is restricted to brown fat and is cold inducible. Four additional transgenic lines were generated with a second transgene containing a 1.8-kb deletion in the 5'-flanking DNA, and expression of this minigene is absent in all tissues analyzed. A DNase I-hypersensitive site located in the 1.8-kb deletion contains a cyclic AMP response element that binds a brown fat tumor enriched nuclear factor. On the basis of these observations, we propose that a cis-acting regulatory sequence between -3 and -1.2 kb of the 5'-flanking region, possibly at a DNase I-hypersensitive site, is required for controlling uncoupling protein expression in vivo.
Boyer
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BB
Jackson Laboratory, Bar Harbor, Maine 04609.
Kozak
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LP
D002001:Adipose Tissue, Brown / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002352:Carrier Proteins; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D003850:Deoxyribonuclease I; D005796:Genes; D005813:Genes, Synthetic; D007473:Ion Channels; D008565:Membrane Proteins / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins; D008969:Molecular Sequence Data; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D000071256:Uncoupling Protein 1
D002352:Carrier Proteins; D007473:Ion Channels; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D012333:RNA, Messenger; D000071256:Uncoupling Protein 1; D012313:RNA; D004247:DNA; D003850:Deoxyribonuclease I