Effects of heterologous downstream sequences on the activity of the HIV-1 promoter and its response to Tat

Michael E. Greenberg, Michael B. Mathews

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

In HIV-1 infection, Tat acts at least in part to control transcriptional elongation by overcoming premature transcriptional termination. In some other genes this process is governed by DNA elements called attenuators in concert with cellular transcription factors. To understand the action of Tat more fully and explore its role as an anti-attenuator, we examined the ability of several natural and synthetic attenuation sequences to modulate transcription initiated at the HIV LTR. Fragments containing these signals were inserted downstream of the TAR element in an HIV-CAT chimera and their effects on transcription were assessed both in vitro and in vivo. Runoff transcription assays in HeLa cell extracts demonstrated that the attenuators give rise to premature termination of transcripts initiated from the heterologous HIV-LTR promoter in vitro. When transiently expressed following transfection into Cos cells, however, premature transcript termination at the attenuation site was not observed. Nevertheless, many of the inserted sequences exerted marked effects on CAT gene expression and on transactivation by Tat at both the RNA and protein levels. The nature and magnitude of the effects depended upon the identity of the attenuator and its orientation but only one of 16 sequences tested met the criteria for a Tat-suppressible attenuator in vivo. One other sequence, in contrast, severely reduced Tat-activated transcription without inhibiting basal transcription These results indicate that sequences downstream of the HIV LTR can influence its function as a promoter and its response to Tat transactivation, but lend little support to their role as attenuators in vivo.

Original languageEnglish (US)
Pages (from-to)5017-5024
Number of pages8
JournalNucleic acids research
Volume25
Issue number24
DOIs
StatePublished - Dec 15 1997

All Science Journal Classification (ASJC) codes

  • Genetics

Fingerprint Dive into the research topics of 'Effects of heterologous downstream sequences on the activity of the HIV-1 promoter and its response to Tat'. Together they form a unique fingerprint.

Cite this