The identification of novel targets of miR-16 and characterization of their biological functions in cancer cells.
BACKGROUND
In eukaryotes, miR-16 is an important microRNA (miRNA) that is involved in numerous biological processes. However, it is not fully understood how miR-16 executes its physiological functions. In the present study, we aimed to identify novel miR-16 targets and study their biological functions.


METHODS
Candidate target genes of miR-16 were screened by microarray analysis of mRNA levels in several cancer cell lines with enhanced miR-16. Three bioinformatics algorithms, including TargetScan, PicTar, and miRanda, were used in combination to calculate the miR-16 targets. The expression levels of miR-16 and target mRNA were examined by relative quantification RT-PCR, and the expression levels of target protein were detected by Western blot. Luciferase reporter plasmids were constructed to confirm direct targeting. The effect of miR-16 and target gene on cell viability was evaluated using MTT assays. The effects of miR-16 and target gene on apoptosis and cell cycle distribution were evaluated by flow cytometry analysis.


RESULTS
By overexpressing miR-16 in several cancer cell lines and measuring global mRNA levels using microarray analysis, we identified 27 genes that may be regulated by miR-16. After the bioinformatics filtering process, 18 genes were selected as candidate miR-16 targets. Furthermore, we experimentally validated three of these candidates, MAP7 (microtubule-associated protein 7), PRDM4 (PR domain containing 4) and CDS2 (CDP-diacylglycerol synthase 2), as direct targets of miR-16. Finally, we demonstrated that miR-16 targeting MAP7 played a critical role in regulating proliferation but not apoptosis and cell cycle progression in cancer cells.


CONCLUSIONS
In summary, the present study identifies several novel miR-16 targets and illustrates a novel function of miR-16 targeting MAP7 in modulating proliferation in cancer cells.
Tianjin Medical University Cancer Institute and Hospital, Huanhuxi Road, Tiyuanbei, Tianjin 300060, China.
Xin
X
Yan
Hongwei
H
Liang
Ting
T
Deng
Kegan
K
Zhu
Suyang
S
Zhang
Nan
N
Wang
Xueyuan
X
Jiang
Xueliang
X
Wang
Rui
R
Liu
Ke
K
Zen
Chen-Yu
CY
Zhang
Yi
Y
Ba
Xi
X
Chen
D017209:Apoptosis / Q000235:genetics; D020029:Base Pairing; D001483:Base Sequence; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic / Q000235:genetics; D019295:Computational Biology; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D006801:Humans; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism; D009369:Neoplasms / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D004268:DNA-Binding Proteins; C505362:MIRN16 microRNA, human; D035683:MicroRNAs; D008869:Microtubule-Associated Proteins; C403744:PRDM4 protein, human; D012333:RNA, Messenger; D014157:Transcription Factors