tat regulates binding of the human immunodeficiency virus trans-activating region RNA loop-binding protein TRP-185

Foon Wu, Joseph Garcia, David Sigman, Richard Gaynor

Research output: Contribution to journalArticle

95 Citations (Scopus)

Abstract

The TAR element extending from -17 to +80 in the human immunodeficiency virus long terminal repeat (HIV LTR) is required for activation of gene expression by the tat trans-activator protein. TAR RNA forms a stable stem-loop structure, and mutagenesis studies indicate that the stem structure, the primary sequence of the loop, and the bulge element are the major determinants for tat activation. RNA gel retardation analysis demonstrates that both tat and cellular proteins bind to TAR RNA, but the mechanism by which these proteins increase HIV gene expression is unknown. We have fractionated HeLa cell nuclear extracts in an attempt to identify cellular proteins that bind to TAR RNA and are involved in regulating HIV gene expression. RNA gel retardation and UV cross-linking reveal that a cellular protein of 185 kD, which we designate TAR RNA-binding protein 185 (TRP-185), binds with both high affinity and marked specificity to TAR RNA. RNA gel retardation and competition analyses indicate that TRP-185 binding is strongly dependent on the TAR RNA loop sequences. The binding of TRP-185 is modulated by both a set of cellular cofactors and the tat protein. Highly purified preparations of TRP-185 are capable of activating in vitro transcription of wild-type, but not mutated, HIV LTR chloramphenicol acetyltransferase (CAT) constructs. These results characterize a positively acting cellular RNA-binding factor, TRP-185, which is involved in the regulation of HIV gene expression.

Original languageEnglish (US)
Pages (from-to)2128-2140
Number of pages13
JournalGenes and Development
Volume5
Issue number11
StatePublished - Nov 1991

Fingerprint

RNA-Binding Proteins
HIV
RNA
HIV Long Terminal Repeat
tat Gene Products
Gels
Gene Expression
Human Immunodeficiency Virus Proteins
Proteins
Chloramphenicol O-Acetyltransferase
Trans-Activators
trans-activation responsive RNA-binding protein
Gene Expression Regulation
Cell Extracts
HeLa Cells
Protein Binding
Mutagenesis

Keywords

  • Cellular factors
  • Gene expression
  • HIV
  • TAR RNA
  • tat

ASJC Scopus subject areas

  • Developmental Biology
  • Genetics

Cite this

tat regulates binding of the human immunodeficiency virus trans-activating region RNA loop-binding protein TRP-185. / Wu, Foon; Garcia, Joseph; Sigman, David; Gaynor, Richard.

In: Genes and Development, Vol. 5, No. 11, 11.1991, p. 2128-2140.

Research output: Contribution to journalArticle

Wu, Foon ; Garcia, Joseph ; Sigman, David ; Gaynor, Richard. / tat regulates binding of the human immunodeficiency virus trans-activating region RNA loop-binding protein TRP-185. In: Genes and Development. 1991 ; Vol. 5, No. 11. pp. 2128-2140.
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