Viral Pathogens and Biosafety Unit, Division of Immunology, Transplantation, and Infectious Diseases, San Raffaele Scientific Institute, Via Olgettina 58, 20132, Milan, Italy.
J Virol. 2011 May;85(10):5183-96. doi: 10.1128/JVI.02302-10. Epub 2011 Feb 23.
Previous studies identified clones of the U937 promonocytic cell line that were either permissive or nonpermissive for human immunodeficiency virus type 1 (HIV-1) replication. These clones were investigated further in the search for host restriction factors that could explain their differential capacity to support HIV-1 replication. Among known HIV-1 restriction factors screened, tripartite motif-containing protein 22 (TRIM22) was the only factor constitutively expressed in nonpermissive and absent in permissive U937 cells. Stable TRIM22 knockdown (KD) rescued HIV-1 long-terminal-repeat (LTR)-driven transcription in KD-nonpermissive cells to the levels observed in permissive cells. Conversely, transduction-mediated expression of TRIM22 in permissive cells reduced LTR-driven luciferase expression by ∼7-fold, supporting a negative role of TRIM22 in HIV-1 transcription. This finding was further confirmed in the human T cell line A3.01 expressing TRIM22. Moreover, overexpression of TRIM22 in 293T cells significantly impaired basal and phorbol myristate acetate-ionomycin-induced HIV-1 LTR-driven gene expression, whereas inhibition of tumor necrosis factor alpha-induced viral transcription was a consequence of lower basal expression. In agreement, TRIM22 equally inhibited an LTR construct lacking the tandem NF-κB binding sites. In addition, TRIM22 did not affect Tat-mediated LTR transactivation. Finally, these effects were independent of TRIM22 E3 ubiquitin-ligase activity. In the context of replication-competent virus, significantly higher levels of HIV-1 production were observed in KD-nonpermissive versus control nonpermissive U937 cells after infection. In contrast, lower peak levels of HIV-1 replication characterized U937 and A3.01 cells expressing TRIM22 versus their control transduced counterpart. Thus, nuclear TRIM22 significantly impairs HIV-1 replication, likely by interfering with Tat- and NF-κB-independent LTR-driven transcription.
先前的研究鉴定出 U937 前单核细胞系的克隆，这些克隆对人类免疫缺陷病毒 1（HIV-1）的复制是允许的或不允许的。在寻找可能解释其支持 HIV-1 复制的不同能力的宿主限制因素的进一步研究中，对这些克隆进行了进一步的研究。在筛选的已知 HIV-1 限制因子中，三结构域蛋白 22（TRIM22）是唯一在非允许性和允许性 U937 细胞中均不表达的因子。稳定的 TRIM22 敲低（KD）使 KD-非允许性细胞中的 HIV-1 长末端重复（LTR）驱动转录恢复到允许性细胞中观察到的水平。相反，在允许性细胞中转导表达 TRIM22 使 LTR 驱动的荧光素酶表达降低了约 7 倍，支持 TRIM22 在 HIV-1 转录中的负作用。这一发现在表达 TRIM22 的人 T 细胞系 A3.01 中得到了进一步证实。此外，在 293T 细胞中过表达 TRIM22 显著抑制了基础和佛波醇肉豆蔻酸乙酸-离子霉素诱导的 HIV-1 LTR 驱动基因表达，而肿瘤坏死因子 alpha 诱导的病毒转录的抑制是由于基础表达降低所致。一致地，TRIM22 同样抑制缺乏串联 NF-κB 结合位点的 LTR 构建体。此外，TRIM22 不影响 Tat 介导的 LTR 反式激活。最后，这些效应与 TRIM22 的 E3 泛素连接酶活性无关。在复制性病毒的背景下，感染后 KD-非允许性与对照非允许性 U937 细胞相比，HIV-1 的产量显著升高。相比之下，表达 TRIM22 的 U937 和 A3.01 细胞的 HIV-1 复制峰值水平较低，与对照转导的对应物相比。因此，核 TRIM22 显著损害 HIV-1 复制，可能通过干扰 Tat 和 NF-κB 非依赖性 LTR 驱动转录。
