MicroRNA-34a: a novel tumor suppressor in p53-mutant glioma cell line U251.
OBJECTIVE
Previous studies showed that microRNA-34 (miR-34a) family was found to be a direct target of p53, functioning downstream of the p53 pathway as tumor suppressors. MiR-34a was identified to represent the status of p53 and participate in initiation and progress of cancers. We undertook this study to investigate the role of miR-34a in glioma cells.


METHODS
Expression levels of miR-34a in glioma cell lines and normal brains were detected using qRT-PCR. Human U251 glioma cells were transfected with miR-34a mimics, and the effects of miR-34a restoration were assessed by MTT assays, cell cycle analysis, caspase-3 activation, and in vitro migration and invasion assays. A computational search revealed a conserved target site of miR-34a within the 3'-untranslated region of SIRT1. Luciferase reporter assay was performed to examine the effects of miR-34a on expression of potential target gene SIRT1, and mRNA and protein expression of SIRT1 after miR-34a transfection were detected by qRT-PCR and Western blot analysis.


RESULTS
MiR-34a expression was markedly reduced in p53-mutant cells U251 compared with A172 and SHG-44 cells expressing wild-type p53 and normal brains. Overexpression of miR-34a in U251 cells resulted in inhibition of cell growth and arrest in G0-G1 phase and induced apoptosis. Also, restoration of miR-34a significantly reduced in vitro migration and invasion capabilities. Reporter assays indicated that SIRT1 was a direct target of miR-34a. In U251 cells, overexpression of miR-34a decreased SIRT1 protein levels but not mRNA expressions, which demonstrated miR-34a-induced SIRT1 inhibition occurred at the posttranscriptional level.


CONCLUSIONS
Our results demonstrate that miR-34a acts as a tumor suppressor in p53-mutant glioma cells U251, partially through regulating SIRT1.
Luan
Shihai
S
Department of Neurosurgery, Huashan Hospital of Fudan University, Shanghai, China.
Sun
Linlin
L
Huang
Fengping
F
D001483:Base Sequence; D001921:Brain / Q000166:cytology / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D005910:Glioma / Q000235:genetics / Q000378:metabolism; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D056564:Sirtuin 1 / Q000235:genetics / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
C522425:MIRN34 microRNA, human; D035683:MicroRNAs; D016159:Tumor Suppressor Protein p53; C447939:SIRT1 protein, human; D056564:Sirtuin 1