Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Inactivation of the tight junction gene CLDN11 by aberrant hypermethylation modulates tubulins polymerization and promotes cell migration in nasopharyngeal carcinoma

doi: 10.1186/s13046-018-0754-y

Figure Lengend Snippet: TUBA1B and TUBB3 are the interacting proteins of CLDN11; CLDN11 blocks cell migration by interfering with tubulin polymerization. a TW02 cells transfected with FLAG-tagged CLDN11 construct (C) or vector control plasmid (V) were harvested after 48 h. TW02 cell extracts and M2 beads were used for a co-immunoprecipitation assay. CLDN11-interacting proteins identified through LC–MS/MS and top-ranking proteins are listed in the table. b Immunoblot analyses were performed to confirm the interaction between FLAG-tagged CLDN11 and endogenous TUBA1B (left panel) or TUBB3 (right panel). c Subcellular distribution of exogenous FLAG-tagged CLDN11 and endogenous TUBA1B or TUBB3 in TW02 cells was assayed through immunofluorescence staining 24 h after transfection. The fluorescent signal (pixel) of each individual cell was quantitated by using the Zen 2.0 software ( n = 50). The proportion of colocalized fluorescent signals (%) is indicated (FLAG-tagged CLDN11, green; Tubulins, red; DAPI, blue). d Schematic illustrations of WT and four deletion FLAG-tagged CLDN11 molecules—transmembrane (TM), extracellular loop (ECL), intracellular loop (ICL), and C-terminus (C). Various FLAG-tagged CLDN11 deletion clones or a vector control were used to dissect the interacting domains on CLDN11 that are crucial for the interaction of endogenous TUBA1B and TUBB3. Input (3%) and immunoprecipitates (30% IP) were assayed through immunoblot analysis by using appropriate antibodies (anti-Flag, anti-TUBA1B and anti-TUBB3). The asterisks denote the major bands of ectopic CLDN11 in the immunoblot assays. e CLDN11 deletion clones were used to perform cell migration assay and to determine which domains on CLDN11 are necessary for blocking cell migration in TW02 cells. f Cell migration and ( g ) cell viability assays were performed in the presence or absence of nocodazole in TW02 or HK1 cells

Article Snippet: Finally, the nuclei were stained with DAPI fluorescent dye (AAT Bioquest, Sunnyvale, CA).

Techniques: Migration, Transfection, Construct, Plasmid Preparation, Control, Co-Immunoprecipitation Assay, Liquid Chromatography with Mass Spectroscopy, Western Blot, Immunofluorescence, Staining, Software, Clone Assay, Cell Migration Assay, Blocking Assay

Journal: Oncotarget

Article Title: Antibody h-R3-dendrimer mediated siRNA has excellent endosomal escape and tumor targeted delivery ability, and represents efficient siPLK1 silencing and inhibition of cell proliferation, migration and invasion

doi: 10.18632/oncotarget.7368

Figure Lengend Snippet: ( A ) Ex vivo fluorescence imaging of five major organs and tumor 24 h post-injection after injection with dendriplex and h-R3/EGF/HSA-dendriplex (T, tumor; H, heart; Lv, liver; S, spleen; Lu, lung; K, kidney). ( B ) Region-of-interest analysis of fluorescent signals from the tumors and five major organs 24 h post-injection. (T, tumor; H, heart; Lv, liver; S, spleen; Lu, lung; K, kidney). Error bars indicate s.d. ( n = 3). ** P < 0.01. * p < 0.05 compared with dendriplexes as the control. ( C ) Ex vivo distribution of siRNA in the tumor treated with dendriplex and h-R3/EGF/HSA-dendriplex 24 h after i.v. injection to nude mice-bearing HepG2 tumor. Cryosections of tumor tissues (6 um thick) were examined by CLSM. DAPI and fluorescein isothiocyanate-labeled phalloidin were used to stain nuclei and F actin (to display the rough cell outline), respectively. Red homogeneous spots present the Cy5-siRNA. Scale bars = 50 μm. ( D ) Quantitative analysis of Cy5-siRNA in tumor was executed by Image-Pro Plus. Error bars indicate s.d. ( n = 3). ** P < 0.01.

Article Snippet: The fluorescent dye DAPI was purchase from Fanbo Biochemical Co (Beijing, China).

Techniques: Ex Vivo, Fluorescence, Imaging, Injection, Control, Labeling, Staining