Journal: Nature Communications

Article Title: A regulatory circuit of miR-125b/miR-20b and Wnt signalling controls glioblastoma phenotypes through FZD6-modulated pathways

doi: 10.1038/ncomms12885

Figure Lengend Snippet: ( a ) IB analysis on PN 23 and 17 spheres with or without FZD6 overexpression using indicated antibodies. ( b ) qRT–PCR analysis of PN-associated genes OLIG2 and SOX2 , and MES-associated genes ALDH1A3 and CD44 in PN 23 spheres with or without FZD6 overexpression. ( c ) FZD6 expression promoted PN 23 spheres proliferation and self-renewal. ( d ) Ectopic expression of FZD6 promoted tumour growth of PN 23 spheres in GBM xenografts in the brain ( n =5). Scale bar, 1.0 mm. ( e ) TOPFlash/FOPFlash assays on PN 23 with overexpressing FZD6 or control, with or without CaMKII inhibitor KN93 (5 μM), TAK1 inhibitor 5Z-7-oxo (5 μM), or a shRNA-NLK. ( f , g ) qRT–PCR analyses of the relative expression of miR-20b and miR-125b ( f ), and Wnt signalling target gene LEF1 ( g ) in PN 23 overexpressing FZD6, or a vector control, with or without CaMKII inhibitor KN93 (5 μM), TAK1 inhibitor 5Z-7-oxo (5 μM), or a shRNA-NLK. ( h ) sphere formation assays for PN 23 and 17 spheres with or without overexpression of FZD1, FZD3, and FZD5, respectively. ( i ) Summary of data shown in . An overview of transmembrane receptors and Wnt ligands that affect FZD6 overexpression-induced phenotypes. Data were normalized to the control. *Relative inhibitory effects on indicated protein expression or cellular properties. Error bars (s.d.) represent the data of triplicate samples for each cell line or tumours from five individual mice in each group. * P <0.05, ** P <0.01, paired two-way Student's t -test. Data are representative from three independent experiments with similar results.

Article Snippet: M50 super 8 × TOPFlash (ID: 12456) and its mutation M51 super 8 × FOPFlash (ID: 12457), lentiviral-TOP-dGFP (ID: 14715), lentiviral-FOP-dGFP (ID: 14885), 7TFC (ID: 24307), TOP-GFP.mC (ID: 35491), FOP-GFP.mC (ID: 35492), Myc-tagged human full-length TCF4 pcDNA (ID: 32738), dnTCF4 (ID: 24310), pcDONR-NLK (ID: 23693) and TAK1 (ID: 23642), FZD5 (ID: 16799), puro-STAT3 shRNA (ID:26596), and shRNA for β-catenin (1248, ID:19761; 2279, ID:19762) constructs were purchased from Addgene .

Techniques: Over Expression, Quantitative RT-PCR, Expressing, Control, shRNA, Plasmid Preparation

Journal: Angiogenesis

Article Title: Frizzled-5: a high affinity receptor for secreted frizzled-related protein-2 activation of nuclear factor of activated T-cells c3 signaling to promote angiogenesis.

doi: 10.1007/s10456-017-9574-5

Figure Lengend Snippet: Fig. 1 The theoretical topology of interaction between SFRP2 and FZD5 is similar to other Wnt/FZD family members. a Full length homology model of SFRP2 and FZD5. CRD, the cysteine-rich domain of FZD5, is colored green. TM, the transmembrane domain of FZD5, is colored orange along with the intracellular domains. b, c Detail of twelve inter- molecular interactions between SFRP2 and the CRD of FZD5. (Color figure online)

Article Snippet: Gain and loss of function studies with FZD5 Stable transfection of 2H11 endothelial cells with FZD5 shRNA To silence FZD5 in endothelial cells, we used HuSH shRNA plasmids containing FZD5 (OriGene, Rockville MD, USA).

Techniques:

Journal: Angiogenesis

Article Title: Frizzled-5: a high affinity receptor for secreted frizzled-related protein-2 activation of nuclear factor of activated T-cells c3 signaling to promote angiogenesis.

doi: 10.1007/s10456-017-9574-5

Figure Lengend Snippet: Fig. 4 SFRP2 and FZD5 co-immunoprecipitate in 2H11 and MS1 cells. a Western blot analysis on samples from sham (control) and shFZD5-transfected 2H11 cells treated for 0, 5 and 15 min with 10 nM SFRP2, showing the levels of SFRP2 and FZD5. Actin was used as a loading control. Dosimetry units (DU) normalized to actin. b Sham and shFZD5 samples from 2H11 cells treated for 0, 5 and 15 with 10 nM SFRP2 were immunoprecipitated with an anti-SFRP2 antibody (lanes 1–6) or a control IgG (lanes 7–12) and the levels of SFRP2 and FZD5 were then measured by western blot. c Western blot

Article Snippet: Gain and loss of function studies with FZD5 Stable transfection of 2H11 endothelial cells with FZD5 shRNA To silence FZD5 in endothelial cells, we used HuSH shRNA plasmids containing FZD5 (OriGene, Rockville MD, USA).

Techniques: Western Blot, Control, Transfection, Immunoprecipitation