[Target prediction and verification of miR-27a in pancreatic cancer].
OBJECTIVE
To predict and verify the target gene of miR-27a in pancreatic cancer by combining the result of comparative proteome analysis.


METHODS
The bioinformatics softwares of TargetScan,PicTar and miRanda were used to predict the possible target genes of miR-27a. Based on the results of comparative proteomics analysis, possible candidates of the target genes were selected. Expression vector of target gene 3'UTR was constructed, and then target gene was verified by dual-luciferase reporter assay system. The PANC-1 and BxPC-3 pancreatic cancer cells were treated with miR-27a mimics or negative control for 48 h. Western blot analysis was used to verify alterations of protein expression of the genes.


RESULTS
PSMA1 was selected as the candidate target gene of miR-27a by bioinformatics prediction and comparative proteome analysis. Dual-luciferase reporter assay showed that miR-27a decreased luciferase activity in cells co-transfected with pmirGLO-PSMA1-WT, compared to the negative control, although significant difference of luciferase activity was not observed in cells co-transfected with pmirGLO-PSMA1-MUT between the two groups. The protein level of PSMA1 was down-regulated in pancreatic cancer cells transfected with miR-27a mimics in comparison with pancreatic cancer cells transfected with negative control.


CONCLUSIONS
PSMA1 is the direct target gene of miR-27a in pancreatic cancer.
Department of Pathology,Peking Union Medical College Hospital, Peking Union Medical College, Chinese Academy of Medical Science, Beijing 100730, China.
Ting-ting
TT
Zhang
Yang
Y
Sun
Cong-wei
CW
Jia
Shuang-ni
SN
Yu
Zhao-hui
ZH
Lu
Jie
J
Chen
D020413:3' Untranslated Regions / Q000235:genetics; D045744:Cell Line, Tumor; D015536:Down-Regulation; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D010190:Pancreatic Neoplasms / Q000378:metabolism* / Q000473:pathology; D010957:Plasmids; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
D020413:3' Untranslated Regions; C531388:MIRN27 microRNA, human; D035683:MicroRNAs; D011994:Recombinant Proteins; D008156:Luciferases; D046988:Proteasome Endopeptidase Complex