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Argonaute 2 immunoprecipitation revealed large tumor suppressor kinase 1 as a novel proapoptotic target of miR‐21 in T cells
Author(s) -
Teteloshvili Nato,
SmigielskaCzepiel Katarzyna,
Yuan Ye,
Seitz Annika,
Jong Debora,
Rutgers Bea,
Jellema Pytrick,
Lei Roelof Jan,
SlezakProchazka Izabella,
Brouwer Elisabeth,
Boots Annemieke M.H.,
Kroesen BartJan,
Berg Anke,
Kluiver Joost
Publication year - 2017
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14011
Subject(s) - jurkat cells , gene knockdown , immunoprecipitation , microrna , biology , microbiology and biotechnology , cell growth , microarray analysis techniques , apoptosis , small interfering rna , chromatin immunoprecipitation , cell culture , gene expression , rna , cancer research , t cell , gene , immunology , promoter , genetics , immune system
Micro RNA (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 α CD 3/α CD 28 activation of Jurkat cells. Inhibition of miR‐21 reduced cell growth which could be explained by an increase in apoptosis. Micro RNA target gene identification by AGO 2 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 AGO 2‐ IP fraction of miR‐21 overexpressing cells. Of these, 71 were at least twofold more enriched in the AGO 2‐ IP fraction of miR‐21 overexpressing cells as compared to AGO 2‐ IP fraction of control cells. The target gene for which the AGO 2‐ IP enrichment was most prominently increased upon miR‐21 overexpression was the proapoptotic protein LATS 1. Luciferase reporter assays and western blot analysis confirmed targeting of LATS 1 by miR‐21. qRT ‐ PCR analysis in primary T cells showed an inverse expression pattern between LATS 1 transcript levels and miR‐21 upon T‐cell stimulation. Finally, LATS 1 knockdown partially rescued the miR‐21 inhibition‐induced impaired cell growth. Collectively, these data identify LATS 1 as a miR‐21 target important for the antiapoptotic function of miR‐21 in T cells and likely also in many types of cancer.