miR-200b suppresses invasiveness and modulates the cytoskeletal and adhesive machinery in esophageal squamous cell carcinoma cells via targeting Kindlin-2

Abstract

To further our understanding of the pathobiology of esophageal squamous cell carcinoma (ESCC), we previously performed microRNA profiling that revealed downregulation of miR-200b in ESCC. Using quantitative real-time PCR applied to 88 patient samples, we confirmed that ESCC tumors expressed significantly lower levels of miR-200b compared with the respective adjacent benign tissues (P = 0.003). Importantly, downregulation of miR-200b significantly correlated with shortened survival (P = 0.025), lymph node metastasis (P = 0.002) and advanced clinical stage (P = 0.020) in ESCC patients. Quantitative mass spectrometry identified 57 putative miR-200b targets, including Kindlin-2, previously implicated in the regulation of tumor invasiveness and actin cytoskeleton in other cell types. Enforced expression of miR-200b mimic in ESCC cells led to a decrease of Kindlin-2 expression, whereas transfection of miR-200b inhibitor induced Kindlin-2 expression. Furthermore, transfection of miR-200b mimic or knockdown of Kindlin-2 in ESCC cells decreased cell protrusion and focal adhesion (FA) formation, reduced cell spreading and invasiveness/migration. Enforced expression of Kindlin-2 largely abrogated the inhibitory effects of miR-200b on ESCC cell invasiveness. Mechanistic studies revealed that Rho-family guanosine triphosphatases and FA kinase mediated the biological effects of the miR-200b-Kindlin-2 axis in ESCC cells. To conclude, loss of miR-200b, a frequent biochemical defect in ESCC, correlates with aggressive clinical features. The tumor suppressor effects of miR-200b may be due to its suppression of Kindlin-2, a novel target of miR-200b that modulates actin cytoskeleton, FA formation and the migratory/invasiveness properties of ESCC.

miR-200b Suppresses Cell Growth, Migration and Invasion by Targeting Notch1 in Nasopharyngeal Carcinoma

“…Recently, Tang et al have reported that ectopic expression of miR-200b and miR-200c impaired cell growth and invasion by directly targeting DNMT3A, DNMT3B and SP1 [40]. Zhang et al found that miR-200b suppressed invasiveness and modulated the cytoskeletal and adhesive machinery in esophageal squamous cell carcinoma cells via targeting Kindlin-2 [41]. Our study adds Notch1 to the growing group of miR-200b target genes and provides evidence that miR-200b suppresses NPC cell growth, migration and invasion by repressing Notch1 expression.…”

Of Kindlins and Cancer

“…Kindlin-2 expression has been found to be dysregulated in several cancer types: prostate 74-77 breast 64, 78-80 , lung 65, 81 , colorectal cancer 82 , pancreas 83, 84 ovarian 85, 86 , esophageal squamous cell carcinoma 87-89 , liver 90 , brain 91 , gastric cancer 92, 93 , bladder 94 , and acute myeloid leukemia 95 . Given the association of kindlin-2 with several cancers of different origins, one can predict an important role that kindlin-2 may play in cancer pathogenesis.…”

Quantitative proteomics reveals the downregulation of GRB2 as a prominent node of F806-targeted cell proliferation network

“…Each gel lane was cut into 10 bands containing approximately the same protein amount, and bands from the three identical gel lanes were pooled [12]. Then the pooled bands were subjected to in-gel digestion, as described previously [13,14]. Briefly, the gel bands were sliced into small pieces (~1 mm 3 ) and destained with 25 mM ammonium bicarbonate in ethanol/ water (50:50, vol/vol).…”

“…This experiment was performed in a single run [13,14]. The tryptic digests were injected into a Nano-LC system (Eksigent Technologies, Dublin, CA), and analyzed by an LTQ-Orbitrap Velos mass spectrometer (Thermo Fisher Scientific, Waltham, MA).…”