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


RESULTS
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
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.
Miller
Heather B
HB
Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America.
Saunders
Kevin O
KO
Tomaras
Georgia D
GD
Garcia-Blanco
Mariano A
MA
D002470:Cell Survival; D016325:HIV Long Terminal Repeat / Q000235:genetics; D015497:HIV-1 / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D012326:RNA Splicing / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*; D015534:Trans-Activators / Q000172:deficiency / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation / Q000235:genetics*; D014779:Virus Replication; D054322:tat Gene Products, Human Immunodeficiency Virus / Q000235:genetics*
C101819:HTATSF1 protein, human; D012367:RNA, Viral; D015534:Trans-Activators; D054322:tat Gene Products, Human Immunodeficiency Virus