HIV-1 antisense transcription is preferentially activated in primary monocyte-derived cells.

J Virol. 2012 Dec;86(24):13785-9.

Laverdure S, Gross A, Arpin-André C, Clerc I, Beaumelle B, Barbeau B, Mesnard JM.

Université Montpellier 1, Centre d’études d’agents Pathogènes et Biotechnologies pour la Santé (CPBS), Montpellier, France.

 

Abstract

In this study, an antisense luciferase-expressing human immunodeficiency virus type 1 (HIV-1) molecular clone was used to infect primary cells. We found that antisense transcription activity from the 3′ long terminal repeat (LTR) was significantly more abundant in monocyte-derived cells than in activated T lymphocytes. Moreover, by analyzing antisense transcription in infected monocyte-derived dendritic cells (MDDCs), we observed that the majority of HIV-1-infected MDDCs with significant antisense transcription activity did not produce Gag. We also confirmed that the negative-strand-encoded antisense protein (ASP) was expressed in monocyte-derived cells.

PMID: 23035216

 

Supplement:

HIV-1 is a complex retrovirus infecting human subjects. HIV-1 harbours essential genes for its replication, which include gag, pol and env but, in addition to these genes, HIV-1 harbours genes coding for regulatory proteins. Retrovirus transcription is dependent on the 5’ LTR region, which is segmented in three regions termed U3, R and U5. The U3 region harbours most of the promoter including important regions for the activation of transcription, which are highly dependent on the binding of different transcription factor. All HIV-1 genes have been thought to be transcribed through this single promoter for a long time. However, early studies described the presence of conserved open reading frames (ORF) in the complementary strand of the HIV-1 provirus, suggesting the existence of viral mRNAs of negative polarity produced from the 3’ LTR. Indeed, we demonstrated the presence of such antisense RNAs in cells infected with HIV-1 and showed that this antisense transcription is controlled from the 3’ LTR. We demonstrated that HIV-1 antisense transcription is more active in monocyte-derived cells than in activated T cells and is unaltered by Tat expression. Among the different negative sense ORFs found in HIV-1, the asp (for antisense protein) ORF, encoded by the complementary strand to the gp120/gp41 junction of the env gene, is the most conserved and the longest. Analysis of its amino acid sequence reveals in the N-terminal region of ASP the presence of two conserved cysteine triplets (with a potential palmitoylation site for the first one) and two SH3-binding motifs with a typical proline rich sequence with a PxxP minimal core. Hydropathy and immunogenicity plots suggest two trans-membrane domains extending from amino acid 63 to 84 and amino acid 146-167. We have effectively found that ASP can be associated to the cellular membrane. Our laboratory recently provided data that also argue for the production of the ASP protein from the antisense transcript in vivo. We are now studying function of ASP to determine whether ASP may serve as the basis for a vaccine.

Jean-Michel MESNARD-1

Figure. Expression of ASP in HIV-1-infected cells. Schematic representation of ASP with the two cysteine triplets, the SH3 binding motif (PxxPxxP), and two potential transmembrane regions (TM). Jurkat cells were infected with HIV-1 NL4.3ASP-Flag virions pseudotyped with VSVg. pNL4.3ASP-Flag corresponds to a molecular proviral DNA in which ASP was tagged with the Flag epitope at its C-terminal end.

Contact: jean-michel.mesnard@cpbs.cnrs.fr

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