Proteasome Inhibition Reveals that a Functional Preintegration Complex Intermediate Can Be Generated during Restriction by Diverse TRIM5 Proteins

Abstract

The primate TRIM5 proteins constitute a class of restriction factors that prevent host cell infection by retroviruses from different species. The TRIM5 proteins act early after virion entry and prevent viral reverse transcription products from accumulating. We recently found that proteasome inhibitors altered the rhesus monkey TRIM5␣ restriction of human immunodeficiency virus type 1 (HIV-1), allowing reverse transcription products to accumulate even though viral infection remained blocked. To assess whether sensitivity to proteasome inhibitors was a common feature of primate TRIM5 proteins, we conducted a similar analysis of restriction mediated by owl monkey TRIM-cyclophilin A (CypA) or human TRIM5␣. Similar to rhesus monkey TRIM5␣ restriction, proteasome inhibition prevented owl monkey TRIM-CypA restriction of HIV-1 reverse transcription, even though HIV-1 infection and the output of 2-LTR circles remained impaired. Likewise, proteasome inhibition alleviated human TRIM5␣ restriction of N-tropic murine leukemia virus reverse transcription. Finally, HIV-1 reverse transcription products escaping rhesus TRIM5␣ restriction by proteasome inhibition were fully competent for integration in vitro, demonstrating that TRIM5␣ likely prevents the viral cDNA from accessing chromosomal target DNA. Collectively, these data indicate that the diverse TRIM5 proteins inhibit retroviral infection in multiple ways and that inhibition of reverse transcription products is not necessary for TRIM5-mediated restriction of retroviral infection.

Cellular Restriction of Retrovirus Particle-Mediated mRNA Transfer

“…6). These data are consistent with recent observations that inhibition of the proteasome during restriction by TRIM5␣ rescues RT and support the notion that the RT block is due to the destruction of the particle and the RNA by the proteasome (1,39). While the proteasome might not degrade the RNA directly, we imagine that degradation of the virion protein would render the genome sensitive to degradation by cellular nucleases.…”

Ovine TRIM5α Can Restrict Visna/Maedi Virus

“…4). The involvement of the proteasome in OvT5 restriction is in line with findings on other lentiviruses (1,40).…”

“…Since the proteasome has been shown to be involved in TRIM5␣ restriction in other species (1,25,40), we inhibited the proteasome and examined OvT5-mediated restriction of VMV. MDTF cells expressing TRIM5␣ Ov2 or empty vector were treated with prewarmed (37°C) proteasome inhibitor MG132 (SigmaAldrich) at a final concentration of 25 M for 1 h before infection with Ev1 at an MOI of 0.2.…”

Generation of human TRIM5α mutants with high HIV-1 restriction activity

“…16,21,22 The proteasome is also involved and causes a decrease in retroviral complementary DNA accumulation in acutely infected cells. 23 TRIM5a proteins can seemingly selfubiquitinate 24,25 and are rapidly degraded by the proteasome on exposure to a restriction-sensitive virus. 26 Finally, TRIM5a interferes with the transport of postentry retroviral complexes toward the nucleus, and this antiviral activity is independent from the one involving the proteasome.…”