miR-15a-3p and miR-16-1-3p Negatively Regulate Twist1 to Repress Gastric Cancer Cell Invasion and Metastasis

Abstract

MicroRNAs are a novel class of gene regulators that function as oncogenes or tumor suppressors. In our current study, we investigated the role of miR-15a-3p and miR-16-1-3p in the regulation of Twist1 expression and EMT process. Our bioinformatics analysis suggested that on the 3' UTR of Twist1, there are two conserved miRNA recognition sites for miR-15a-3p and miR-16-1-3p respectively. Interestingly, overexpression of miR-15a-3p and miR-16-1-3p significantly suppressed the activity of luciferase reporter containing Twist1-3' UTR, reduced mRNA and protein level of EMT related genes such as TWIST1, N-cadherin, α-SMA and Fibronectin, and repressed MMP9 and MMP2 activity, as well as cell migration and invasion. Conversely, inhibition of miR-15a-3p and miR-16-1-3p significantly increased TWIST1, N-cadherin, α-SMA and Fibronectin protein expression. In addition, Twist1 co-transfection significantly ameliorated the loss of cell migration and invasion. Moreover, overexpression of miR-15a-3p and miR-16-1-3p dramatically suppressed the ability of BGC823 cells to form colonies in vitro and develop tumors in vivo in nude mice. Finally, qPCR and Western blot analysis showed that miR-15a-3p and miR-16-1-3p were significantly reduced in clinical gastric cancer tissue, whereas Twist1 mRNA and protein were significantly up-regulated, suggesting that this aberrant down-regulation of miR-15a-3p and miR-16-1-3p might be associated with the abnormal regulation of Twist1 and the EMT process in gastric cancer development. Our results help to elucidate a novel and important mechanism for the regulation of Twist1 in the development of cancer.

MicroRNAs and the PTEN/PI3K/Akt pathway in gastric cancer (Review)

“…CDX1 was targeted by oncogenic miR-296-5p, which was detected to be overexpressed in GC tissues and abolished the suppressive effects induced by CDX1 (10). Different from the elevated expression level of oncogenic miRNAs in GC, the expressions of tumor-suppressor miRNAs, including miR-137, miR-34a, miR-15a and miR-16-1, which exert negative regulations on cell proliferation, epithelial-mesenchymal transition (EMT), migration, invasion, colony formation in vitro, in addition to tumorigenicity and metastasis in vivo, have been documented to be evidently decreased in the tumor tissues and cell lines compared with normal controls (18)(19)(20)(21). The targets responsible for their effects including Akt2, platelet-derived growth factor receptor (PDGFR) and twist family bHLH transcription factor 1 (18)(19)(20)(21).…”

Long non-coding RNA UCA1 exerts growth modulation by miR-15a in human thyroid cancer TPC-1 cells

“…In addition, miR-145 is regarded as a target of lncRNA-TUG1 and associates with tumour progression in thyroid cancer cells [11]. What is more, miR-15a-5p is a tumour suppressor in various kinds of cancers [12][13][14]. However, the effect of miR-15a in thyroid cancer is granted elusive.…”

MiR-16-1*and miR-16-2*possess strong tumor suppressive and anti-metastatic properties in osteosarcoma

“…This is the first report of any function for miR-16-1* and/or miR-16-2*. Although anti-metastatic properties of miR-16-1* in gastric cancer cells were reported earlier by Wang T and his co-workers 29 , it was not sufficiently detailed and explored as in our study. In fact, Wang T et al overexpressed miR-16-1 precursor for all functional experiments and did not check for miR-16 overexpression 29 .…”

“…Although anti-metastatic properties of miR-16-1* in gastric cancer cells were reported earlier by Wang T and his co-workers 29 , it was not sufficiently detailed and explored as in our study. In fact, Wang T et al overexpressed miR-16-1 precursor for all functional experiments and did not check for miR-16 overexpression 29 . Since miR-16-1* precursor overexpression is expected to lead to overexpression of both, the "lead" miR-16 and the "passenger" miR-16-1*, strands (Fig 2A), then, it is not clear which strand caused functional effects in gastric 2 0 cancer cells 29 .…”

“…In fact, Wang T et al overexpressed miR-16-1 precursor for all functional experiments and did not check for miR-16 overexpression 29 . Since miR-16-1* precursor overexpression is expected to lead to overexpression of both, the "lead" miR-16 and the "passenger" miR-16-1*, strands (Fig 2A), then, it is not clear which strand caused functional effects in gastric 2 0 cancer cells 29 . Noteworthy, our data also indicate that miR-16-1* and miR-16-2* have different although strongly overlapping functions.…”