Initiation of reverse transcription during cell-to-cell transmission of human immunodeficiency virus infection uses pre-existing reverse transcriptase.
H3B cells, a laboratory clone of H9 cells persistently infected with the HTLV-IIIB strain of human immunodeficiency virus (HIV), contained significant levels of cell-associated reverse transcriptase (RT) activity measured by in vitro assays using either exogenous or endogenous templates. The cell-associated RT activity detected using exogenous template was almost wholly in a soluble (non-sedimentable) form whereas endogenous activity sedimented as a particulate structure associated with viral RNA. Despite this, H3B cells did not contain episomal HIV DNA detectable by Southern blot, indicating that in vivo reverse transcription was not occurring to any significant extent in these cells. However, when susceptible HUT 78 cells were infected by co-cultivation with H3B cells, dramatic synthesis of episomal HIV DNA occurred. Concurrently with this de novo initiation of reverse transcription, however, we found no detectable change in intracellular levels or cleavage profiles of immunoprecipitable RT polypeptides. Finally, actinomycin D pre-treatment of H3B cells to prevent de novo transcription from donor cell proviral DNA after co-cultivation did not affect the initiation of in vivo reverse transcription following cell-to-cell HIV infection. These results demonstrated that cells persistently infected with HIV contained significant fully cleaved cell-associated RT in a form that was active in vitro but not in vivo and that following cell-to-cell transmission of HIV infection to susceptible cells, de novo reverse transcription was initiated without detectable evidence of further synthesis or proteolytic processing of HIV RT. The nature of this initiation process requires further study.
Li
P
P
National Centre for HIV Virology Research, Institute of Medical and Veterinary Science, Adelaide, Australia.
Stephenson
A J
AJ
Brennan
P A
PA
Karageorgos
L
L
Kok
T
T
Kuiper
L J
LJ
Swift
J
J
Burrell
C J
CJ
D001692:Biological Transport; D015153:Blotting, Western; D002478:Cells, Cultured; D006678:HIV / Q000201:enzymology / Q000235:genetics / Q000254:growth & development* / Q000648:ultrastructure; D006801:Humans; D008854:Microscopy, Electron; D011233:Precipitin Tests; D012367:RNA, Viral / Q000302:isolation & purification; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D014158:Transcription, Genetic; D016662:Virus Integration
D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase