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PMID:7933101

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Citation

Ou, SH, Garcia-Martínez, LF, Paulssen, EJ and Gaynor, RB (1994) Role of flanking E box motifs in human immunodeficiency virus type 1 TATA element function. J. Virol. 68:7188-99

Abstract

Human immunodeficiency virus type 1 (HIV-1) gene expression is dependent on a number of cis-acting DNA elements present in the HIV-1 long terminal repeat. Previous studies have demonstrated that the TATA element is critical for basal and Tat-induced HIV-1 gene expression. The HIV-1 TATA region has an unusual structure in that the TATA sequence is flanked by two palindromic sequence motifs (CANNTG) known as E boxes which can serve as binding sites for the basic helix-loop-helix (bHLH) class of DNA-binding proteins. In this study, we performed site-directed mutagenesis of both the TATA and the flanking E box sequences of HIV-1. We also substituted the sequences flanking the adenovirus E3 promoter TATA sequence for those flanking the HIV-1 TATA sequence. Constructs were assayed for their levels of basal and Tat-induced gene expression by both in vitro transcription and transient expression assays. Both the TATA box and flanking sequences including the E box motifs were found to be important in modulating both basal gene expression and Tat-induced HIV-1 gene expression. Gel retardation analysis demonstrated that binding of both the recombinant TATA-binding protein (TBP) and the TFIID fraction which contains both TBP and TBP-associated factors was dependent primarily on the TATA element. However, competition analysis suggested that the E boxes may play a role in stabilizing the binding of TFIID but not recombinant TBP. Two proteins representing different classes of bHLH proteins, E47 and AP-4, were assayed for their ability to bind to the flanking E box motifs. We isolated a cDNA clone encoding the complete AP-4 protein and demonstrated that both AP-4 and E47 bound specifically to the 3' E box motif, which contains sequences that correspond to the consensus binding site (CAGCTG). Gel retardation analysis indicated that the binding of AP-4 to the E boxes excluded the binding of TBP to the TATA box. These studies are consistent with a model in which different classes of cellular bHLH proteins may be involved in regulating HIV-1 TATA element function by either inhibiting or promoting the assembly of different preinitiation transcriptional complexes.

Links

PubMed PMC237158

Keywords

Amino Acid Sequence; Base Sequence; Cell Line; DNA-Binding Proteins/metabolism; Gene Expression Regulation, Viral; Genes, Viral; HIV Long Terminal Repeat; HIV-1/genetics; Humans; Molecular Sequence Data; TATA Box/physiology; TATA-Box Binding Protein; TCF Transcription Factors; Transcription Factor 7-Like 1 Protein; Transcription Factor TFIID; Transcription Factors/metabolism; Transcription, Genetic

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

HUMAN:TBP

enables

GO:0000976: transcription cis-regulatory region binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:TFAP4

located_in

GO:0005634: nucleus

ECO:0000314: direct assay evidence used in manual assertion

C

Seeded From UniProt

complete

HUMAN:TFAP4

involved_in

GO:0043922: negative regulation by host of viral transcription

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

HUMAN:TFAP4

involved_in

GO:0043392: negative regulation of DNA binding

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

HUMAN:TFAP4

enables

GO:0070888: E-box binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:TFE2

enables

GO:0070888: E-box binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

HUMAN:TBP

enables

GO:0044212: transcription regulatory region DNA binding

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete


See also

References

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