Next-Generation Sequencing Analysis Reveals Differential Expression Profiles of MiRNA-mRNA Target Pairs in KSHV-Infected Cells.
Kaposi's sarcoma associated herpesvirus (KSHV) causes several tumors, including primary effusion lymphoma (PEL) and Kaposi's sarcoma (KS). Cellular and viral microRNAs (miRNAs) have been shown to play important roles in regulating gene expression. A better knowledge of the miRNA-mediated pathways affected by KSHV infection is therefore important for understanding viral infection and tumor pathogenesis. In this study, we used deep sequencing to analyze miRNA and cellular mRNA expression in a cell line with latent KSHV infection (SLKK) as compared to the uninfected SLK line. This approach revealed 153 differentially expressed human miRNAs, eight of which were independently confirmed by qRT-PCR. KSHV infection led to the dysregulation of ~15% of the human miRNA pool and most of these cellular miRNAs were down-regulated, including nearly all members of the 14q32 miRNA cluster, a genomic locus linked to cancer and that is deleted in a number of PEL cell lines. Furthermore, we identified 48 miRNAs that were associated with a total of 1,117 predicted or experimentally validated target mRNAs; of these mRNAs, a majority (73%) were inversely correlated to expression changes of their respective miRNAs, suggesting miRNA-mediated silencing mechanisms were involved in a number of these alterations. Several dysregulated miRNA-mRNA pairs may facilitate KSHV infection or tumor formation, such as up-regulated miR-708-5p, associated with a decrease in pro-apoptotic caspase-2 and leukemia inhibitory factor LIF, or down-regulated miR-409-5p, associated with an increase in the p53-inhibitor MDM2. Transfection of miRNA mimics provided further evidence that changes in miRNAs are driving some observed mRNA changes. Using filtered datasets, we also identified several canonical pathways that were significantly enriched in differentially expressed miRNA-mRNA pairs, such as the epithelial-to-mesenchymal transition and the interleukin-8 signaling pathways. Overall, our data provide a more detailed understanding of KSHV latency and guide further studies of the biological significance of these changes.
HIV and AIDS Malignancy Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America; The Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom.
Coralie
C
Viollet
HIV and AIDS Malignancy Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
David A
DA
Davis
Institute of Molecular Oncology, Alexander Fleming Biomedical Sciences Research Center, Vari, Greece.
Martin
M
Reczko
HIV and AIDS Malignancy Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Joseph M
JM
Ziegelbauer
Nuffield Division of Clinical Laboratory Sciences, University of Oxford, Oxford, United Kingdom.
Francesco
F
Pezzella
The Wellcome Trust Centre for Human Genetics, University of Oxford, Oxford, United Kingdom; Institute of Molecular Oncology, Alexander Fleming Biomedical Sciences Research Center, Vari, Greece; McGill University and Génome Québec Innovation Centre, Montréal, Canada.
Jiannis
J
Ragoussis
HIV and AIDS Malignancy Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America.
Robert
R
Yarchoan
D001483:Base Sequence; D045744:Cell Line, Tumor; D058750:Epithelial-Mesenchymal Transition / Q000235:genetics; D019288:Herpesvirus 8, Human / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics; D054685:Lymphoma, Primary Effusion / Q000235:genetics* / Q000821:virology; D035683:MicroRNAs / Q000235:genetics*; D051736:Proto-Oncogene Proteins c-mdm2 / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D012514:Sarcoma, Kaposi / Q000235:genetics* / Q000821:virology; D017422:Sequence Analysis, DNA; D016159:Tumor Suppressor Protein p53 / Q000037:antagonists & inhibitors
C508606:CXCL8 protein, human; D016209:Interleukin-8; C572367:MIRN409 microRNA, human; C568528:MIRN708 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2