Abstract
We hypothesize that miR-3151, localized to a GWAS-identified chronic lymphocytic leukemia (CLL) risk locus (8q22.3), is a tumor suppressor miRNA silenced by promoter DNA methylation in CLL. The promoter of miR-3151 was methylated in 5/7 (71%) CLL cell lines, 30/98 (31%) diagnostic primary samples, but not normal controls. Methylation of miR-3151 correlated inversely with expression. Treatment with 5-Aza-2'-deoxycytidine led to promoter demethylation and miR-3151 re-expression. Luciferase assay confirmed MAP-kinase activating death domain (MADD) and phosphoinositide-3-kinase, regulatory subunit 2 (PIK3R2) as direct targets of miR-3151. Moreover, restoration of miR-3151 resulted in inhibition of cellular proliferation and enhanced apoptosis, repression of MADD and PIK3R2, downregulation of MEK/ERK and PI3K/AKT signaling, and repression of MCL1. Lastly, miR-3151 methylation was significantly associated with methylation of miR-203 and miR-34b/c in primary CLL samples. Therefore, this study showed that miR-3151 is a tumor suppressive miRNA frequently hypermethylated and hence silenced in CLL. miR-3151 silencing by DNA methylation protected CLL cells from apoptosis through over-expression of its direct targets MADD and PIK3R2, hence constitutive activation of MEK/ERK and PI3K/AKT signaling respectively, and consequently over-expression of MCL1.
Keywords:
DNA methylation; chronic lymphocytic leukemia; miR-3151; microRNA; tumor suppressor.
MeSH terms
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3' Untranslated Regions
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Adult
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Aged
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Aged, 80 and over
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Apoptosis
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Asian People / genetics
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Azacitidine / analogs & derivatives
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Azacitidine / pharmacology
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Binding Sites
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Cell Line, Tumor
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Cell Proliferation
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China
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DNA Methylation* / drug effects
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DNA Modification Methylases / antagonists & inhibitors
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DNA Modification Methylases / metabolism
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Death Domain Receptor Signaling Adaptor Proteins / genetics
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Death Domain Receptor Signaling Adaptor Proteins / metabolism*
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Decitabine
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Enzyme Activation
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Enzyme Inhibitors / pharmacology
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Epigenesis, Genetic* / drug effects
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Extracellular Signal-Regulated MAP Kinases / metabolism*
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Female
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Gene Expression Regulation, Neoplastic
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Gene Silencing* / drug effects
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Guanine Nucleotide Exchange Factors / genetics
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Guanine Nucleotide Exchange Factors / metabolism*
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Humans
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Leukemia, Lymphocytic, Chronic, B-Cell / enzymology*
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Leukemia, Lymphocytic, Chronic, B-Cell / genetics*
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Leukemia, Lymphocytic, Chronic, B-Cell / pathology
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Male
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MicroRNAs / genetics*
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MicroRNAs / metabolism
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Middle Aged
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Myeloid Cell Leukemia Sequence 1 Protein / metabolism
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Phosphatidylinositol 3-Kinases / genetics
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Phosphatidylinositol 3-Kinases / metabolism*
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Phosphorylation
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Proto-Oncogene Proteins c-akt / metabolism*
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RNA Interference
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Signal Transduction
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Transfection
Substances
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3' Untranslated Regions
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Death Domain Receptor Signaling Adaptor Proteins
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Enzyme Inhibitors
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Guanine Nucleotide Exchange Factors
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MADD protein, human
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MCL1 protein, human
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MIRN203 microRNA, human
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MIRN3151 microRNA, human
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MIRN34 microRNA, human
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MicroRNAs
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Myeloid Cell Leukemia Sequence 1 Protein
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Decitabine
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DNA Modification Methylases
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phosphoinositol-3 kinase regulatory subunit 2, human
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Proto-Oncogene Proteins c-akt
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Extracellular Signal-Regulated MAP Kinases
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Azacitidine