Regulation of tumor necrosis factor-like weak inducer of apoptosis receptor protein (TWEAKR) expression by Kaposi's sarcoma-associated herpesvirus microRNA prevents TWEAK-induced apoptosis and inflammatory cytokine expression.
Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is the causative agent of KS, the second most common AIDS-associated malignancy. KSHV expresses at least 18 different mature microRNAs (miRNAs) during latency. To identify cellular targets of KSHV miRNAs, we have analyzed a previously reported series of microarrays examining changes in cellular gene expression in the presence of KSHV miRNAs. Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) receptor (TWEAKR) was among the most consistently and robustly downregulated genes in the presence of KSHV miR-K12-10a (miR-K10a). Results from luciferase assays with reporter plasmids containing the 3' untranslated region (UTR) of TWEAKR suggest a targeting of TWEAKR by miR-K10a. The mutation of two predicted miR-K10a recognition sites within the 3' UTR of TWEAKR completely disrupts inhibition by miR-K10a. The expression of TWEAKR was downregulated in cells transfected with miR-K10a as well as during de novo KSHV infection. In a KS tumor-derived endothelial cell line, the downregulation of TWEAKR by miR-K10a resulted in reduced levels of TWEAK-induced caspase activation. In addition, cells transfected with miR-K10a showed less induction of apoptosis by annexin V staining and terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling (TUNEL) assays. Finally, the downregulation of TWEAKR by miR-K10a in primary human endothelial cells resulted in a decrease in levels of expression of the proinflammatory cytokines interleukin-8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) in response to TWEAK. These results identify and validate an important cellular target of KSHV miRNAs. Furthermore, we demonstrate that a viral miRNA protects cells from apoptosis and suppresses a proinflammatory response, which may have significant implications in the complex context of KS lesions.
Abend
Johanna R
JR
HIV and AIDS Malignancy Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Uldrick
Thomas
T
Ziegelbauer
Joseph M
JM
D017304:Annexin A5; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D018932:Chemokine CCL2 / Q000378:metabolism; D000074049:Cytokine TWEAK; D017931:DNA Primers / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology*; D019288:Herpesvirus 8, Human / Q000235:genetics*; D006801:Humans; D020287:In Situ Nick-End Labeling; D016209:Interleukin-8 / Q000378:metabolism; D008156:Luciferases; D035683:MicroRNAs / Q000235:genetics / Q000502:physiology*; D016296:Mutagenesis; D018124:Receptors, Tumor Necrosis Factor / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000074050:TWEAK Receptor; D048069:Tumor Necrosis Factors / Q000502:physiology*
D017304:Annexin A5; C111835:CCL2 protein, human; D018932:Chemokine CCL2; D000074049:Cytokine TWEAK; D017931:DNA Primers; D016209:Interleukin-8; D035683:MicroRNAs; D018124:Receptors, Tumor Necrosis Factor; C109743:TNFSF12 protein, human; D000074050:TWEAK Receptor; D048069:Tumor Necrosis Factors; D008156:Luciferases; D020169:Caspases