miR-214/199a/199a* cluster levels predict poor survival in hepatocellular carcinoma through interference with cell-cycle regulators.
OBJECTIVE
To identify the clinical and functional association of miR-214/199a/199a* cluster in human hepatocellular carcinoma (HCC) and to clarify the mechanism of miR-214.


METHODS
Kaplan-Meier and Cox proportional regression analyses were used to determine the association of miR-214/199a/199a* cluster levels with the survival of HCC patients. The role of miR-214 in regulating HCC cell proliferation was studied with miR-214 mimics/inhibitor-treated cells. Furthermore, the inhibition effect of miR-214 on E2F2, cyclin-dependent kinase (CDK) 3 and CDK6 expression was assessed in HCC cell lines with miR-214 mimics/inhibitors to increase/decrease miR-214 expression. Direct binding of miR-214 to the 3'-untranslated regions of E2F2, CDK3, and CDK6 was verified by dual-luciferase reporter assay.


RESULTS
In analyzing HCC clinical specimens and cell lines, we discovered a uniform decrease in miR-214/199a/199a* expression in comparison with noncancerous tissue or normal liver epithelial cell lines. Higher miR-214 levels were related with improved patient survival. Overexpression of miR-214 in HCC cells inhibited proliferation by inducing G1-S checkpoint arrest. Conversely, RNA interference-mediated silencing of miR-214 promoted cell-cycle progression and accelerated the proliferation of HCC cells. E2F2, CDK3 and CDK6 were each directly targeted for inhibition by miR-214, and restoring their expression reversed miR-214 inhibition of cell-cycle progression. The relationship between expression of miR-214 and its targets was confirmed in HCC tumor xenografts and clinical specimens.


CONCLUSIONS
Our results demonstrate that miR-214 has tumor-suppressive activity in HCC through inhibition of E2F2, CDK3 and CDK6.
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.|Key Laboratory of Tropical Disease Control, Ministry of Education, Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Peipei
P
Wang
Department of Radiology, Guangzhou Red Cross Hospital/The Fourth Affiliated Hospital of Jinan University Medical College, Guangzhou, Guangdong Province, China.
Song
S
Chen
Department of Burn Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
He
H
Fang
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Xiaojuan
X
Wu
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Dabiao
D
Chen
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Liang
L
Peng
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.|Key Laboratory of Tropical Disease Control, Ministry of Education, Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Zhiliang
Z
Gao
Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong Province, China.|Key Laboratory of Tropical Disease Control, Ministry of Education, Sun Yat-sen University, Guangzhou, Guangdong Province, China.
Chan
C
Xie
D000818:Animals; D015153:Blotting, Western; D006528:Carcinoma, Hepatocellular / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D059447:Cell Cycle Checkpoints / Q000235:genetics*; D002460:Cell Line; D045744:Cell Line, Tumor; D056849:Cyclin-Dependent Kinase 3 / Q000235:genetics / Q000378:metabolism; D051361:Cyclin-Dependent Kinase 6 / Q000235:genetics / Q000378:metabolism; D050703:E2F2 Transcription Factor / Q000235:genetics / Q000378:metabolism; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D053208:Kaplan-Meier Estimate; D008113:Liver Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008297:Male; D008807:Mice, Inbred BALB C; D008819:Mice, Nude; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D005810:Multigene Family; D011379:Prognosis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014183:Transplantation, Heterologous
D050703:E2F2 Transcription Factor; C547397:MIRN214 microRNA, human; D035683:MicroRNAs; C531408:mirn199 microRNA, human; C098776:CDK3 protein, human; C495911:CDK6 protein, human; D056849:Cyclin-Dependent Kinase 3; D051361:Cyclin-Dependent Kinase 6
cell cycle; human hepatocellular carcinoma; microRNA