Identification of Cellular Genes Targeted by KSHV-Encoded MicroRNAs

MicroRNAs (miRNAs) are 19 to 23 nucleotide–long RNAs that post-transcriptionally regulate gene expression. Human cells express several hundred miRNAs which regulate important biological pathways such as development, proliferation, and apoptosis. Recently, 12 miRNA genes have been identified within the genome of Kaposi sarcoma–associated herpesvirus; however, their functions are still unknown. To identify host cellular genes that may be targeted by these novel viral regulators, we performed gene expression profiling in cells stably expressing KSHV-encoded miRNAs. Data analysis revealed a set of 81 genes whose expression was significantly changed in the presence of miRNAs. While the majority of changes were below 2-fold, eight genes were down-regulated between 4- and 20-fold. We confirmed miRNA-dependent regulation for three of these genes and found that protein levels of thrombospondin 1 (THBS1) were decreased >10-fold. THBS1 has previously been reported to be down-regulated in Kaposi sarcoma lesions and has known activity as a strong tumor suppressor and anti-angiogenic factor, exerting its anti-angiogenic effect in part by activating the latent form of TGF-β. We show that reduced THBS1 expression in the presence of viral miRNAs translates into decreased TGF-β activity. These data suggest that KSHV-encoded miRNAs may contribute directly to pathogenesis by down-regulation of THBS1, a major regulator of cell adhesion, migration, and angiogenesis.

微小RNA（miRNAs）是一类长度为19至23个核苷酸的RNA分子，可在转录后水平调控基因表达。人类细胞可表达数百种miRNAs，这些miRNAs参与调控发育、增殖、凋亡等重要生物学通路。近期研究在卡波西肉瘤相关疱疹病毒（Kaposi sarcoma–associated herpesvirus, KSHV）的基因组中鉴定出12个miRNA基因，但其功能仍未明确。
为鉴定可能受这些新型病毒调控因子靶向的宿主细胞基因，我们对稳定表达KSHV编码miRNAs的细胞开展了基因表达谱分析。数据分析显示，在miRNAs存在的条件下，共有81个基因的表达发生显著改变。尽管大多数表达变化幅度低于2倍，但有8个基因的表达被下调了4至20倍。
我们对其中3个基因的miRNA依赖性调控进行了验证，发现血小板反应蛋白1（THBS1）的蛋白水平下调幅度超过10倍。既往研究表明，THBS1在卡波西肉瘤病灶中呈下调状态，其作为强效肿瘤抑制因子与抗血管生成因子发挥功能，可通过激活转化生长因子β（TGF-β）的潜伏形式部分介导其抗血管生成作用。
本研究显示，病毒miRNAs介导的THBS1表达降低会导致TGF-β活性下降。上述数据表明，KSHV编码的miRNAs可能通过下调THBS1——一种调控细胞黏附、细胞迁移与血管生成的关键调节因子——直接参与病毒致病过程。