Two upstream elements activate transcription of a major histocompatibility complex class I gene in vitro.
Expression of major histocompatibility complex (MHC) class I genes exhibits unique tissue and developmental specificity. In an effort to study molecular mechanisms of MHC class I gene regulation, an in vitro transcription system has been established. In B cell nuclear extracts a template DNA containing the mouse H-2Ld promoter sequence accurately directed RNA polymerase II-dependent transcription of a G-free cassette. A conserved class I regulatory complex previously shown to moderately enhance promoter activity in vivo enhanced transcription in vitro by 2-3 fold. Much of this enhancement was accounted for by a 40 bp fragment within the complex, which was capable of activating a basal H-2Ld promoter in either orientation. Farther downstream, another element called site B was identified, which independently activated MHC class I transcription in vitro by 2-4 fold. Site B bound a specific nuclear factor(s) through an NF-1 binding site but not through a neighboring CCAAT site. The functional significance of site B in vivo was demonstrated in transfection experiments in which site B enhanced MHC class I promoter activity to a degree comparable to that seen in vitro. With the identification of the two upstream activators, MHC class I genes may serve as a model to study roles of sequence-specific DNA-binding proteins in transcription in vitro.
Driggers
P H
PH
Laboratory of Molecular Growth Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892.
Elenbaas
B A
BA
An
J B
JB
Lee
I J
IJ
Ozato
K
K
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D022762:CCAAT-Enhancer-Binding Proteins; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005805:Genes, MHC Class I / Q000235:genetics*; D006801:Humans; D008745:Methylation; D051379:Mice; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics; D014158:Transcription, Genetic / Q000235:genetics*; D014162:Transfection / Q000235:genetics; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; D009687:Nuclear Proteins; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human