Systematic analysis of viral and cellular microRNA targets in cells latently infected with human gamma-herpesviruses by RISC immunoprecipitation assay.
The mRNA targets of microRNAs (miRNAs) can be identified by immunoprecipitation of Argonaute (Ago) protein-containing RNA-induced silencing complexes (RISCs) followed by microarray analysis (RIP-Chip). Here we used Ago2-based RIP-Chip to identify transcripts targeted by Kaposi's sarcoma-associated herpesvirus (KSHV) miRNAs (n = 114), Epstein-Barr virus (EBV) miRNAs (n = 44), and cellular miRNAs (n = 2337) in six latently infected or stably transduced human B cell lines. Of the six KSHV miRNA targets chosen for validation, four showed regulation via their 3'UTR, while two showed regulation via binding sites within coding sequences. Two genes governing cellular transport processes (TOMM22 and IPO7) were confirmed to be targeted by EBV miRNAs. A significant number of viral miRNA targets were upregulated in infected cells, suggesting that viral miRNAs preferentially target cellular genes induced upon infection. Transcript half-life both of cellular and viral miRNA targets negatively correlated with recruitment to RISC complexes, indicating that RIP-Chip offers a quantitative estimate of miRNA function.
Dölken
Lars
L
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Malterer
Georg
G
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Erhard
Florian
F
Institute for Informatics, Ludwig-Maximilians-University Munich, Amalienstrasse 17, 80333 Munich, Germany.
Kothe
Sheila
S
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Friedel
Caroline C
CC
Institute for Informatics, Ludwig-Maximilians-University Munich, Amalienstrasse 17, 80333 Munich, Germany.
Suffert
Guillaume
G
Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 rue René Descartes, Université de Strasbourg, 67084 Strasbourg, France.
Marcinowski
Lisa
L
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Motsch
Natalie
N
Institute of Virology, Haus 47, Universitätsklinikum des Saarlandes, 66421 Homburg/Saar, Germany.
Barth
Stephanie
S
Institute of Virology, Haus 47, Universitätsklinikum des Saarlandes, 66421 Homburg/Saar, Germany.
Beitzinger
Michaela
M
Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
Lieber
Diana
D
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Bailer
Susanne M
SM
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Hoffmann
Reinhard
R
Institute of Medical Microbiology, Technical University Munich, Trogerstrasse 30, 81675 Munich, Germany.
Ruzsics
Zsolt
Z
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Kremmer
Elisabeth
E
Helmholtz Zentrum München, Institut für Molekulare Immunologie, Marchioninistraße 25, 81377 Munich, Germany.
Pfeffer
Sébastien
S
Institut de Biologie Moléculaire et Cellulaire du CNRS, 15 rue René Descartes, Université de Strasbourg, 67084 Strasbourg, France.
Zimmer
Ralf
R
Institute for Informatics, Ludwig-Maximilians-University Munich, Amalienstrasse 17, 80333 Munich, Germany.
Koszinowski
Ulrich H
UH
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany.
Grässer
Friedrich
F
Institute of Virology, Haus 47, Universitätsklinikum des Saarlandes, 66421 Homburg/Saar, Germany.
Meister
Gunter
G
Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany.
Haas
Jürgen
J
Max von Pettenkofer-Institute, Ludwig-Maximilians-University Munich, Pettenkofer Strasse 9a, 80336 Munich, Germany; Division of Pathway Medicine, University of Edinburgh, 49 Little France Crescent, Edinburgh EH16 4SB, UK. Electronic address: haas@lmb.uni-muenchen.de.
D002460:Cell Line; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D005786:Gene Expression Regulation; D015967:Gene Expression Regulation, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics*; D019288:Herpesvirus 8, Human / Q000235:genetics*; D054884:Host-Pathogen Interactions; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D046228:Microarray Analysis; D012367:RNA, Viral / Q000378:metabolism; D014773:Virology / Q000379:methods*
D035683:MicroRNAs; D012367:RNA, Viral