Department of Biochemistry and Molecular Biophysics, Columbia University, New York, New York, United States of America.
PLoS Pathog. 2011 Apr;7(4):e1002019. doi: 10.1371/journal.ppat.1002019. Epub 2011 Apr 7.
Human TRIM5α potently restricts particular strains of murine leukemia viruses (the so-called N-tropic strains) but not others (the B- or NB-tropic strains) during early stages of infection. We show that overexpression of SUMO-1 in human 293T cells, but not in mouse MDTF cells, profoundly blocks N-MLV infection. This block is dependent on the tropism of the incoming virus, as neither B-, NB-, nor the mutant R110E of N-MLV CA (a B-tropic switch) are affected by SUMO-1 overexpression. The block occurred prior to reverse transcription and could be abrogated by large amounts of restricted virus. Knockdown of TRIM5α in 293T SUMO-1-overexpressing cells resulted in ablation of the SUMO-1 antiviral effects, and this loss of restriction could be restored by expression of a human TRIM5α shRNA-resistant plasmid. Amino acid sequence analysis of human TRIM5α revealed a consensus SUMO conjugation site at the N-terminus and three putative SUMO interacting motifs (SIMs) in the B30.2 domain. Mutations of the TRIM5α consensus SUMO conjugation site did not affect the antiviral activity of TRIM5α in any of the cell types tested. Mutation of the SIM consensus sequences, however, abolished TRIM5α antiviral activity against N-MLV. Mutation of lysines at a potential site of SUMOylation in the CA region of the Gag gene reduced the SUMO-1 block and the TRIM5α restriction of N-MLV. Our data suggest a novel aspect of TRIM5α-mediated restriction, in which the presence of intact SIMs in TRIM5α, and also the SUMO conjugation of CA, are required for restriction. We propose that at least a portion of the antiviral activity of TRIM5α is mediated through the binding of its SIMs to SUMO-conjugated CA.
人类 TRIM5α 在感染早期能强烈限制特定株的鼠白血病病毒（所谓的 N 型株），但不能限制其他株（B 型或 NB 型株）。我们发现，在人 293T 细胞中过表达 SUMO-1，但不在鼠 MDTF 细胞中过表达，会显著阻断 N-MLV 感染。这种阻断依赖于进入病毒的嗜性，因为无论是 B-、NB-，还是 N-MLV CA 的突变型 R110E（B 型嗜性转换）都不受 SUMO-1 过表达的影响。阻断发生在逆转录之前，可以被大量受限的病毒消除。在 293T SUMO-1 过表达细胞中敲低 TRIM5α，会导致 SUMO-1 的抗病毒作用丧失，而这种限制的丧失可以通过表达人 TRIM5α shRNA 抗性质粒来恢复。对人 TRIM5α 的氨基酸序列分析显示，在 N 端有一个公认的 SUMO 连接位点，在 B30.2 结构域中有三个假定的 SUMO 相互作用基序（SIM）。TRIM5α 公认的 SUMO 连接位点的突变，在任何测试的细胞类型中都不会影响 TRIM5α 的抗病毒活性。然而，SIM 共识序列的突变，会使 TRIM5α 对 N-MLV 的抗病毒活性丧失。在 Gag 基因的 CA 区域中潜在 SUMO 化位点的赖氨酸突变，会降低 SUMO-1 阻断和 TRIM5α 对 N-MLV 的限制。我们的数据表明，TRIM5α 介导的限制有一个新的方面，即 TRIM5α 中完整的 SIMs 的存在，以及 CA 的 SUMO 连接，对于限制是必需的。我们提出，TRIM5α 的至少一部分抗病毒活性是通过其 SIMs 与 SUMO 连接的 CA 的结合来介导的。
