MiR-4282 suppresses proliferation and mobility of human colorectal carcinoma cells by targeting semaphorin 3E.
BACKGROUND
MicroRNAs play an important role in cancer development. Deregulation of microRNAs can lead to tumorigenesis. Class 3 semaphorin, semaphorin 3E (Sema3E), has been shown to be implicated in tumor growth and metastasis. The role of miR-4282 in regulating colorectal carcinoma and its correlation to Sema3E remain uncertain.


METHODS
Real-time quantitative reverse transcription polymerase chain reaction was used to detect the levels of miR-4282 and Sema3E in colorectal carcinoma cells and colorectal tumor tissues. Sema3E protein level in cell lines and human tissues was analyzed by western blot Transient transfections of miR-4282 inhibitor or mimics were conducted to silence or overexpress miR-4282. Sema3E siRNA was transfected to knockdown Sema3E in tumor cell lines. MTT assay was employed to measure colorectal tumor cell growth. Migration and invasion of the cells were examined by trans-well assays. Luciferase reporter assays were performed to confirm miR-4282 targeted at Sema3E.


RESULTS
In the present study, reduced miR-4282 expression was observed in the colorectal carcinoma cell lines and human carcinoma tissues in comparison with normal human colon cells (P<0.05) or matched non-tumor tissues (P<0.05), whereas, Sema3E was up-regulated in colorectal carcinoma cells lines (P<0.05) and human colorectal tumor tissues (P<0.05). MiR-4282 was then reduced by the inhibitor and overexpressed by its mimics transfection. It was found that miR-4282 inhibition promoted cell growth, migration and invasion (P<0.05) of HT29 and HCT116 colorectal carcinoma cells while miR-4282 overexpression suppressed cell growth and mobility (P<0.05). Sema3E was predicted as a target of miR-4282 in miRDB database. We found that miR-4282 overexpression significantly reduced luciferase activity of pRL-Sema3E-3'-UTR (P<0.05), but failed to alter the activity of pRL-sema3E-3'-UTR-mutation. Also, miR4282 overexpression suppressed Sema3E expression in the colorectal carcinoma cell lines. To further confirm the role of Sema3E suppression in the function of the colorectal carcinoma cells by miR-4282, HT29 and HCT116 cells were transfected with Sema3E siRNA. We found that cell growth, migration and invasion of HT29 and HCT116 cells were dramatically inhibited by Sema3E knockdown (P<0.05).


CONCLUSIONS
Our findings suggested that miR-4282 is a tumor suppressor in colorectal carcinoma cells and exerted its inhibitory effect on the tumor cells through targeting Sema3E by inhibiting Sema3E translation or enhancing Sema3E mRNA degradation. Thus, manipulation of miR-4282 and interfere with Sema3E might represent a potential target for the treatment of colorectal cancer.
Department of General Surgery, The Affiliated Drum Tower Hospital of Nanjing University Medical School, Nanjing, China - wenxianguan@yahoo.com.
Xing
X
Kang
Meng
M
Wang
Hao
H
Wang
Xiaofei
X
Shen
Wenxian
W
Guan
D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D015179:Colorectal Neoplasms / Q000235:genetics / Q000473:pathology*; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D006801:Humans; D035683:MicroRNAs / Q000502:physiology*; D009361:Neoplasm Invasiveness; D039961:Semaphorins / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000502:physiology
C000616433:MIRN-4282 microRNA, human; D035683:MicroRNAs; C494123:SEMA3E protein, human; D039961:Semaphorins