title: Argonaute 2 immunoprecipitation revealed large tumor suppressor kinase 1 as a novel proapoptotic target of miR-21 in T cells.
abstract: MicroRNA (miR)-21 is an important suppressor of T-cell apoptosis that is also overexpressed in many types of cancers. The exact mechanisms underlying the antiapoptotic effects of miR-21 are not well understood. In this study, we used the Jurkat T-cell line as a model to identify apoptosis-associated miR-21 target genes. We showed that expression of miR-21 rapidly increases upon αCD3/αCD28 activation of Jurkat cells. Inhibition of miR-21 reduced cell growth which could be explained by an increase in apoptosis. MicroRNA target gene identification by AGO2 RNA-immunoprecipitation followed by gene expression microarray (RIP-Chip) resulted in the identification of 72 predicted miR-21 target genes that were at least twofold enriched in the AGO2-IP fraction of miR-21 overexpressing cells. Of these, 71 were at least twofold more enriched in the AGO2-IP fraction of miR-21 overexpressing cells as compared to AGO2-IP fraction of control cells. The target gene for which the AGO2-IP enrichment was most prominently increased upon miR-21 overexpression was the proapoptotic protein LATS1. Luciferase reporter assays and western blot analysis confirmed targeting of LATS1 by miR-21. qRT-PCR analysis in primary T cells showed an inverse expression pattern between LATS1 transcript levels and miR-21 upon T-cell stimulation. Finally, LATS1 knockdown partially rescued the miR-21 inhibition-induced impaired cell growth. Collectively, these data identify LATS1 as a miR-21 target important for the antiapoptotic function of miR-21 in T cells and likely also in many types of cancer.
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.|Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Nato
authors.initials: N
authors.lastname: Teteloshvili
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.|Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Smigielska-Czepiel
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.|Institute of Clinical Pharmacology of the Second Affiliated Hospital, Harbin Medical University, Heilongjiang Province, China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Yuan
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Annika
authors.initials: A
authors.lastname: Seitz
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Debora
authors.initials: D
authors.lastname: de Jong
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Bea
authors.initials: B
authors.lastname: Rutgers
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Pytrick
authors.initials: P
authors.lastname: Jellema
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Roelof Jan
authors.initials: RJ
authors.lastname: van der Lei
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Izabella
authors.initials: I
authors.lastname: Slezak-Prochazka
authors.affiliation: Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.|Department of Rheumatology and Clinical Immunology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Brouwer
authors.affiliation: Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.|Department of Rheumatology and Clinical Immunology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Annemieke M H
authors.initials: AM
authors.lastname: Boots
authors.affiliation: Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.|Department of Laboratory Medicine, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Bart-Jan
authors.initials: BJ
authors.lastname: Kroesen
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.|Groningen Research initiative on healthy Ageing and Immune Longevity (GRAIL), University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Anke
authors.initials: A
authors.lastname: van den Berg
authors.affiliation: Department of Pathology and Medical Biology, University of Groningen, University Medical Center Groningen, The Netherlands.
authors.forename: Joost
authors.initials: J
authors.lastname: Kluiver
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017209:Apoptosis / Q000235:genetics* / Q000276:immunology; D060565:Argonaute Proteins / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D018106:CD28 Antigens / Q000819:agonists / Q000235:genetics / Q000276:immunology; D017252:CD3 Complex / Q000235:genetics / Q000276:immunology; D019556:COS Cells; D002461:Cell Line, Transformed; D002522:Chlorocebus aethiops; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D019169:Jurkat Cells; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects; D035683:MicroRNAs / Q000235:genetics* / Q000276:immunology; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000276:immunology; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
chemical_list: C466422:AGO2 protein, human; D000911:Antibodies, Monoclonal; D060565:Argonaute Proteins; D018106:CD28 Antigens; D017252:CD3 Complex; C524940:MIRN21 microRNA, human; D035683:MicroRNAs; D034741:RNA, Small Interfering; D008156:Luciferases; C092745:LATS1 protein, human; D017346:Protein Serine-Threonine Kinases
keywords: AGO2-RIP; LATS1; T cells; apoptosis; miR-21