Tat-SF1 is not required for Tat transactivation but does regulate the relative levels of unspliced and spliced HIV-1 RNAs

PLoS One. 2009 May 27;4(5):e5710. doi: 10.1371/journal.pone.0005710.

Abstract

Background: HIV-1 relies on several host proteins for productive viral transcription. HIV-1 Tat-specific factor 1 (Tat-SF1) is among these cofactors that were identified by in vitro reconstituted transcription reactions with immunodepleted nuclear extracts. At the onset of this work, the prevailing hypothesis was that Tat-SF1 was a required cofactor for the viral regulatory protein, Tat; however, this had not previously been formally tested in vivo.

Methodology/principal findings: To directly address the involvement of Tat-SF1 in HIV-1 gene expression, we depleted Tat-SF1 in HeLa cells by conventional expression of shRNAs and in T- Rex -293 cells containing tetracycline-inducible shRNAs targeting Tat-SF1. We achieved efficient depletion of Tat-SF1 and demonstrated that this did not affect cell viability. HIV-1 infectivity decreased in Tat-SF1-depleted cells, but only when multiple rounds of infection occurred. Neither Tat-dependent nor basal transcription from the HIV-1 LTR was affected by Tat-SF1 depletion, suggesting that the decrease in infectivity was due to a deficiency at a later step in the viral lifecycle. Finally, Tat-SF1 depletion resulted in an increase in the ratio of unspliced to spliced viral transcripts.

Conclusions/significance: Tat-SF1 is not required for regulating HIV-1 transcription, but is required for maintaining the ratios of different classes of HIV-1 transcripts. These new findings highlight a novel, post-transcriptional role for Tat-SF1 in the HIV-1 life cycle.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Cell Survival
  • HIV Long Terminal Repeat / genetics
  • HIV-1 / genetics*
  • HIV-1 / pathogenicity
  • HIV-1 / physiology
  • HeLa Cells
  • Humans
  • RNA Splicing / genetics*
  • RNA, Viral / genetics*
  • Trans-Activators / deficiency
  • Trans-Activators / metabolism*
  • Transcription, Genetic
  • Transcriptional Activation / genetics*
  • Virus Replication
  • tat Gene Products, Human Immunodeficiency Virus / genetics*

Substances

  • HTATSF1 protein, human
  • RNA, Viral
  • Trans-Activators
  • tat Gene Products, Human Immunodeficiency Virus