Background:  Post-entry replication of HIV-1 and N-tropic murine leukemia virus (MLV) is inhibited in cell lines derived from Old World monkeys. These restriction activities are termed Ref1 and Lv1, respectively. Viral capsid (CA) is the main determinant of this restriction. The host factor responsible for this restriction in Old World monkeys was identified as TRIM5a, a putative E3 ligase. The cellular protein cyclophilin A (CypA), an HIV-1 CA interactor, has been suggested to modulate TRIM5a-mediated restriction of HIV-1. It was found in our lab that the Ref1 restriction activity against N-MLV in human cells can be counteracted by the mitochondria-targeting drug As₂O₃. As₂O₃ is known to affect the functions of the TRIM-family member PML. Here, we asked whether As₂O₃ and similarly acting drugs counteracted TRIM5a-mediated restriction in non-human primate cell lines, and how CypA depletion would affect these restriction activities.

Methods:  We used pharmacological agents that affect mitochondrial membrane integrity, such as As₂O₃. CypA was depleted by RNA interference. Viral cDNA synthesis was monitored by Southern blot and real-time PCR. Infections were analyzed over a single cycle by the use of GFP-expressing viral constructs.

Results:  We analyzed the effect of the mitochondria-targeting drugs As₂O₃ , PK11195, and m-Cl-CCP on retroviral replication in African green monkey Vero cells, which are known to show both Ref1 and Lv1 activities, and in rhesus macaque FRhK4 cells, which show only Lv1 activity. As₂O₃ and PK11195 counteracted both Ref1 and Lv1 in African green monkey cells. PK11195 suppressed Lv1 activity in FRhK4 cells. In the absence of cellular CypA (RNAi-treated cells), infectivity of HIV-1 was enhanced in both Vero and FRhK4 cells. Disruption of the CypA:CA interaction increased viral cDNA synthesis in Vero cells, whereas in FRhK4 cells, transport to the nucleus was increased.

Conclusions:  CypA was found to be a necessary co-factor for TRIM5a-mediated restriction of HIV-1 in Old World monkeys. CypA effects on HIV-1 replication are cell-type specific. Restriction of HIV-1 and N-MLV in Old World monkeys was shown to be counteracted by mitochondria-targeting agents.

Keywords: restriction; TRIM5alpha; arsenic trioxide