Architecture et Réactivité de l'ARN, Institut de Biologie Moléculaire et Cellulaire du CNRS, Université de Strasbourg, Strasbourg, France.
PLoS Pathog. 2011 Dec;7(12):e1002405. doi: 10.1371/journal.ppat.1002405. Epub 2011 Dec 8.
Kaposi's sarcoma herpesvirus (KSHV) encodes a cluster of twelve micro (mi)RNAs, which are abundantly expressed during both latent and lytic infection. Previous studies reported that KSHV is able to inhibit apoptosis during latent infection; we thus tested the involvement of viral miRNAs in this process. We found that both HEK293 epithelial cells and DG75 cells stably expressing KSHV miRNAs were protected from apoptosis. Potential cellular targets that were significantly down-regulated upon KSHV miRNAs expression were identified by microarray profiling. Among them, we validated by luciferase reporter assays, quantitative PCR and western blotting caspase 3 (Casp3), a critical factor for the control of apoptosis. Using site-directed mutagenesis, we found that three KSHV miRNAs, miR-K12-1, 3 and 4-3p, were responsible for the targeting of Casp3. Specific inhibition of these miRNAs in KSHV-infected cells resulted in increased expression levels of endogenous Casp3 and enhanced apoptosis. Altogether, our results suggest that KSHV miRNAs directly participate in the previously reported inhibition of apoptosis by the virus, and are thus likely to play a role in KSHV-induced oncogenesis.
卡波氏肉瘤疱疹病毒（KSHV）编码一组 12 个微小（mi）RNA，这些 miRNA 在潜伏和裂解感染期间大量表达。先前的研究报告称，KSHV 能够在潜伏感染期间抑制细胞凋亡；因此，我们测试了病毒 miRNA 在这个过程中的参与情况。我们发现，表达 KSHV miRNA 的 HEK293 上皮细胞和稳定表达 KSHV miRNA 的 DG75 细胞均能免受凋亡。通过微阵列分析鉴定了在 KSHV miRNA 表达后显著下调的潜在细胞靶标。其中，我们通过荧光素酶报告基因检测、定量 PCR 和 Western blot 验证了 caspase 3（Casp3），这是控制细胞凋亡的关键因素。通过定点突变，我们发现三个 KSHV miRNA，miR-K12-1、3 和 4-3p，负责 Casp3 的靶向。在 KSHV 感染的细胞中特异性抑制这些 miRNA 会导致内源性 Casp3 的表达水平增加并增强细胞凋亡。总之，我们的结果表明，KSHV miRNA 直接参与了先前报道的病毒抑制细胞凋亡的过程，因此可能在 KSHV 诱导的肿瘤发生中发挥作用。
