An in vitro transcription system that recapitulates equine infectious anemia virus Tat-mediated inhibition of human immunodeficiency virus type 1 Tat activity demonstrates a role for positive transcription elongation factor b and associated proteins in the mechanism of Tat activation

C. Sune, A. C. Goldstrohm, J. Peng, D. H. Price, Mariano Garcia-Blanco

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

Equine infectious anemia virus (EIAV) activates transcription via a Tat protein, a TAR element, and the equine elongation factor positive transcription elongation factor b (P-TEFb). In human cells, EIAV Tat (eTat) can inhibit the ability of human immunodeficiency virus type 1 (HIV-1) Tat (hTat) to activate transcription from the HIV-1 long terminal repeat, demonstrating that EIAV Tat can interact nonproductively with human P-TEFb. To study the mechanism of EIAV Tat and HIV-1 Tat activation, we developed an in vitro elongation assay that recapitulates EIAV Tat-mediated inhibition of HIV-1 Tat trans-activation. We found that eTat specifically inhibits activation of elongation by HIV-1 Tat while having no effect on basal transcription elongation. The competitive inhibition of hTat activation was reversed by an activity present in HeLa cell nuclear extracts, most likely a form of P-TEFb. Recombinant P-TEFb (cyclin T1 and CDK9) overcame the inhibition of transcription by eTat but in a nonspecific manner. EIAV Tat affinity chromatography was used to purify the activity present in nuclear extract that was capable of reversing eTat inhibition. We characterized the protein components of this activity, which include cyclin T1, CDK9, Tat-SF1, and at least three unidentified proteins. These data suggest that additional factors are involved in the mechanism of Tat activation. (C) 2000 Academic Press.

Original languageEnglish (US)
Pages (from-to)356-366
Number of pages11
JournalVirology
Volume274
Issue number2
DOIs
StatePublished - Sep 1 2000
Externally publishedYes

Fingerprint

Equine infectious anemia virus
Peptide Elongation Factors
HIV-1
Transcription Factors
Proteins
Cyclin T
HIV Long Terminal Repeat
tat Gene Products
In Vitro Techniques
Cell Extracts
Affinity Chromatography
HeLa Cells
Horses

ASJC Scopus subject areas

  • Virology
  • Infectious Diseases

Cite this

@article{43a9f037c30b4dd78500728f22238327,
title = "An in vitro transcription system that recapitulates equine infectious anemia virus Tat-mediated inhibition of human immunodeficiency virus type 1 Tat activity demonstrates a role for positive transcription elongation factor b and associated proteins in the mechanism of Tat activation",
abstract = "Equine infectious anemia virus (EIAV) activates transcription via a Tat protein, a TAR element, and the equine elongation factor positive transcription elongation factor b (P-TEFb). In human cells, EIAV Tat (eTat) can inhibit the ability of human immunodeficiency virus type 1 (HIV-1) Tat (hTat) to activate transcription from the HIV-1 long terminal repeat, demonstrating that EIAV Tat can interact nonproductively with human P-TEFb. To study the mechanism of EIAV Tat and HIV-1 Tat activation, we developed an in vitro elongation assay that recapitulates EIAV Tat-mediated inhibition of HIV-1 Tat trans-activation. We found that eTat specifically inhibits activation of elongation by HIV-1 Tat while having no effect on basal transcription elongation. The competitive inhibition of hTat activation was reversed by an activity present in HeLa cell nuclear extracts, most likely a form of P-TEFb. Recombinant P-TEFb (cyclin T1 and CDK9) overcame the inhibition of transcription by eTat but in a nonspecific manner. EIAV Tat affinity chromatography was used to purify the activity present in nuclear extract that was capable of reversing eTat inhibition. We characterized the protein components of this activity, which include cyclin T1, CDK9, Tat-SF1, and at least three unidentified proteins. These data suggest that additional factors are involved in the mechanism of Tat activation. (C) 2000 Academic Press.",
author = "C. Sune and Goldstrohm, {A. C.} and J. Peng and Price, {D. H.} and Mariano Garcia-Blanco",
year = "2000",
month = "9",
day = "1",
doi = "10.1006/viro.2000.0480",
language = "English (US)",
volume = "274",
pages = "356--366",
journal = "Virology",
issn = "0042-6822",
publisher = "Academic Press Inc.",
number = "2",

}

TY - JOUR

T1 - An in vitro transcription system that recapitulates equine infectious anemia virus Tat-mediated inhibition of human immunodeficiency virus type 1 Tat activity demonstrates a role for positive transcription elongation factor b and associated proteins in the mechanism of Tat activation

AU - Sune, C.

AU - Goldstrohm, A. C.

AU - Peng, J.

AU - Price, D. H.

AU - Garcia-Blanco, Mariano

PY - 2000/9/1

Y1 - 2000/9/1

N2 - Equine infectious anemia virus (EIAV) activates transcription via a Tat protein, a TAR element, and the equine elongation factor positive transcription elongation factor b (P-TEFb). In human cells, EIAV Tat (eTat) can inhibit the ability of human immunodeficiency virus type 1 (HIV-1) Tat (hTat) to activate transcription from the HIV-1 long terminal repeat, demonstrating that EIAV Tat can interact nonproductively with human P-TEFb. To study the mechanism of EIAV Tat and HIV-1 Tat activation, we developed an in vitro elongation assay that recapitulates EIAV Tat-mediated inhibition of HIV-1 Tat trans-activation. We found that eTat specifically inhibits activation of elongation by HIV-1 Tat while having no effect on basal transcription elongation. The competitive inhibition of hTat activation was reversed by an activity present in HeLa cell nuclear extracts, most likely a form of P-TEFb. Recombinant P-TEFb (cyclin T1 and CDK9) overcame the inhibition of transcription by eTat but in a nonspecific manner. EIAV Tat affinity chromatography was used to purify the activity present in nuclear extract that was capable of reversing eTat inhibition. We characterized the protein components of this activity, which include cyclin T1, CDK9, Tat-SF1, and at least three unidentified proteins. These data suggest that additional factors are involved in the mechanism of Tat activation. (C) 2000 Academic Press.

AB - Equine infectious anemia virus (EIAV) activates transcription via a Tat protein, a TAR element, and the equine elongation factor positive transcription elongation factor b (P-TEFb). In human cells, EIAV Tat (eTat) can inhibit the ability of human immunodeficiency virus type 1 (HIV-1) Tat (hTat) to activate transcription from the HIV-1 long terminal repeat, demonstrating that EIAV Tat can interact nonproductively with human P-TEFb. To study the mechanism of EIAV Tat and HIV-1 Tat activation, we developed an in vitro elongation assay that recapitulates EIAV Tat-mediated inhibition of HIV-1 Tat trans-activation. We found that eTat specifically inhibits activation of elongation by HIV-1 Tat while having no effect on basal transcription elongation. The competitive inhibition of hTat activation was reversed by an activity present in HeLa cell nuclear extracts, most likely a form of P-TEFb. Recombinant P-TEFb (cyclin T1 and CDK9) overcame the inhibition of transcription by eTat but in a nonspecific manner. EIAV Tat affinity chromatography was used to purify the activity present in nuclear extract that was capable of reversing eTat inhibition. We characterized the protein components of this activity, which include cyclin T1, CDK9, Tat-SF1, and at least three unidentified proteins. These data suggest that additional factors are involved in the mechanism of Tat activation. (C) 2000 Academic Press.

UR - http://www.scopus.com/inward/record.url?scp=0034284591&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034284591&partnerID=8YFLogxK

U2 - 10.1006/viro.2000.0480

DO - 10.1006/viro.2000.0480

M3 - Article

C2 - 10964778

AN - SCOPUS:0034284591

VL - 274

SP - 356

EP - 366

JO - Virology

JF - Virology

SN - 0042-6822

IS - 2

ER -