title: MiR-9 downregulates CDX2 expression in gastric cancer cells.
abstract: Ectopic expression of CDX2, a caudal-related homeobox protein, is known to be associated with the development of intestinal metaplasia in the stomach and gastric carcinogenesis. Previously, we reported that DNA methylation was partly responsible for CDX2 silencing in gastric cancer (GC). However, the mechanism underlying the aberrant expression of CDX2 during malignant transformation remained unclear. MicroRNAs (miRNAs) are small non-coding RNAs that function as post-transcriptional regulators. To elucidate the role of miRNAs in CDX2 downregulation in GC cells, putative miRNAs, such as miR-9, were computationally predicted. After exogenous pre-miR-9 precursor transfection, the luciferase activity of a reporter vector containing a part of the 3'-UTR of CDX2 was downregulated in HEK-293T cells. The inverse correlation between the miR-9 and CDX2 protein levels was demonstrated in GC cell lines. By means of miR-9 overexpression and knockdown techniques, the expression levels of the CDX2 protein and downstream target genes (p21, MUC2 and TFF3) were responsively altered in MKN45 and NUGC-3 cells. Transfection of an anti-miR-9 molecule significantly inhibited cell growth by promoting G(1)  cell cycle arrest in MKN45 cells similarly to the effect of CDX2 overexpression. Moreover, examination of the miR-9 levels in primary GC tissues revealed that the amounts of miR-9 in the CDX2-negative group were significantly higher than those in the CDX2-positive group (p = 0.004). Therefore, miR-9 might repress CDX2 expression via the binding site in the 3'-UTR, resulting in the promotion of cell proliferation in GCs.
authors.affiliation: Department of Molecular Oncology, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, Tokyo, Japan.
authors.forename: Pichayanoot
authors.initials: P
authors.lastname: Rotkrua
authors.forename: Yoshimitsu
authors.initials: Y
authors.lastname: Akiyama
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Hashimoto
authors.forename: Takeshi
authors.initials: T
authors.lastname: Otsubo
authors.forename: Yasuhito
authors.initials: Y
authors.lastname: Yuasa
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics*; D015153:Blotting, Western; D000071616:CDX2 Transcription Factor; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D019175:DNA Methylation; D015536:Down-Regulation; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013274:Stomach Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology
chemical_list: D020413:3' Untranslated Regions; D000071616:CDX2 Transcription Factor; C491270:CDX2 protein, human; D018398:Homeodomain Proteins; C541874:MIRN92 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger; D008156:Luciferases