Trim-Cyclophilin A Fusion Proteins Can Restrict Human Immunodeficiency Virus Type 1 Infection at Two Distinct Phases in the Viral Life Cycle

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Abstract
The Trim5alpha protein from several primates restricts retroviruses in a capsid (CA)-dependent manner. In owl monkeys, the B30.2 domain of Trim5 has been replaced by cyclophilin A (CypA) following a retrotransposition. Restriction of human immunodeficiency virus type 1 (HIV-1) by the resulting Trim5-CypA fusion protein depends on CA binding to CypA, suggesting both that the B30.2 domain might be involved in CA binding and that the tripartite RING motif, B-BOX, and coiled coil (RBCC) motif domain can function independently of the B30.2 domain in restriction. To investigate the potential of RBCCs from other Trims to participate in restricting HIV-1, CypA was fused to the RBCC of Trim1, Trim18, and Trim19 and tested for restriction. Despite low identity within the RBCC domain, all fusion proteins were found to restrict HIV-1 but not the nonbinding G89V mutant, indicating that the overall structure of RBCC and not its primary sequence was important for the restriction function. The critical interaction between CA and Trim-CypA appears to take place soon after viral entry. Quantitative PCR analysis of viral reverse transcriptase products revealed that the different fusion proteins block HIV-1 at two distinct stages of its life cycle, either prior to reverse transcription or just before integration. With Trim1 and Trim18, this timing is dependent on the length of the Trim component of the fusion protein. These observations suggest that restriction factor binding can have different mechanistic consequences.