Tumour suppressor microRNA-584 directly targets oncogene Rock-1 and decreases invasion ability in human clear cell renal cell carcinoma.
BACKGROUND
The purpose of this study was to identify new tumour suppressor microRNAs (miRs) in clear cell renal cell carcinoma (ccRCC), carry out functional analysis of their suppressive role and identify their specific target genes.


METHODS
To explore suppressor miRs in RCC, miR microarray and real-time PCR were performed using HK-2 and A-498 cells. Cell viability, invasion and wound healing assays were carried out for functional analysis after miR transfection. To determine target genes of miR, we used messenger RNA (mRNA) microarray and target scan algorithms to identify target oncogenes. A 3'UTR luciferase assay was also performed. Protein expression of target genes in ccRCC tissues was confirmed by immunohistochemistry and was compared with miR-584 expression in ccRCC tissues.


RESULTS
Expression of miR-584 in RCC (A-498 and 769-P) cells was downregulated compared with HK-2 cells. Transfection of miR-584 dramatically decreased cell motility. The ROCK-1 mRNA was inhibited by miR-584 and predicted to be target gene. The miR-584 decreased 3'UTR luciferase activity of ROCK-1 and ROCK-1 protein expression. Low expression of miR-584 in ccRCC tissues was correlated with high expression of ROCK-1 protein. The knockdown of ROCK-1 by siRNA inhibited cell motility.


CONCLUSIONS
miR-584 is a new tumour suppressor miR in ccRCC and inhibits cell motility through downregulation of ROCK-1.
Department of Urology, San Francisco Veterans Affairs Medical Center, University of California, San Francisco, 4150 Clement Street, San Francisco, CA 94121, USA.
K
K
Ueno
H
H
Hirata
V
V
Shahryari
Y
Y
Chen
M S
MS
Zaman
K
K
Singh
Z L
ZL
Tabatabai
Y
Y
Hinoda
R
R
Dahiya
D020413:3' Untranslated Regions; D001483:Base Sequence; D002292:Carcinoma, Renal Cell / Q000235:genetics / Q000473:pathology*; D045744:Cell Line, Tumor; D017931:DNA Primers; D055785:Gene Knockdown Techniques; D006801:Humans; D007680:Kidney Neoplasms / Q000235:genetics / Q000473:pathology*; D035683:MicroRNAs / Q000502:physiology*; D009361:Neoplasm Invasiveness / Q000235:genetics*; D054460:rho-Associated Kinases / Q000235:genetics*
D020413:3' Untranslated Regions; D017931:DNA Primers; D035683:MicroRNAs; D054460:rho-Associated Kinases