Figure S1.
Characterisation of tensin specific antibodies and tensin localization. (A) Schematic of the four human tensin family members. Tensin1-3 have an N-terminal region homologous to phosphatase and tensin homolog (PTEN). Tensin2 has an N-terminal lipid-binding C1 domain. All four family members have C-terminal Src-homology 2 (SH2) and phosphotyrosine-binding (PTB) domains. The middle regions are largely unstructured with little amino acid sequence homology between family members. (B) Schematic showing various reported interaction partners of the tensin family, including: cytoplasmic domains of beta integrin subunits (β1, β3, β5, β7) with the PTB domain that reportedly supports integrin activation; the SH2 domain with p130CAS; actin to the N-terminus; the Rho GAPs DLC1-3 to both the N- and C-termini; growth factor receptors (EGFR, cMET) and additional FA proteins (e.g., FAK and p130Cas, Hic5 and ILK) to the C-terminus. (C) NIH3T3 mouse fibroblasts were transfected with the indicated GFP-tagged human tensin constructs and immunostained with antibodies against (human) TNS1, TNS2, or TNS3. Note that the respective antibodies detect only the expressed tensin member they are directed against (i.e., tensin1 antibody detects only over-expressed GFP-tensin1, tensin2 antibody only the expressed GFP-tensin2 and the tensin3 antibody only the GFP-tensin3; indicated by red boxes). (D) Expression of tensin1, 2, and 3 in U2OS and TIF cells evaluated by Western blot and immunofluorescence. A whole cell lysate from U2OS cells was run on the same gel separated by ladder lanes. After transfer, the membrane was cut and incubated with individual tensin antibodies and GAPDH separately (since tensin1, 2, and 3 have a similar molecular weight). (E) Left panel: TIF cell stained for vinculin and tensin3, note that tensin3 localises more to the centrally located adhesions and vinculin to the peripheral adhesions; right panel: TIF cell stained for tensin3 and fibronectin, note the similar localization. (F) Left panels: TIF cells were treated in suspension with blebbistatin (50 µM) or an equivalent volume of DMSO, for 60 min. Cells were fixed after spreading on fibronectin-coated glass for 60 min. Note the absence of tensin3- or vinculin-positive structures in blebbistatin-treated cells. Right panels: TIF cells cultured overnight on fibronectin-coated glass were treated with blebbistatin (50 µM) or an equivalent volume of DMSO, for 45 min prior to fixation. Scale bars in C–F indicate 10 µm. Source data are available for this figure: SourceData FS1.

Characterisation of tensin specific antibodies and tensin locali z ation. (A) Schematic of the four human tensin family members. Tensin1-3 have an N-terminal region homologous to phosphatase and tensin homolog (PTEN). Tensin2 has an N-terminal lipid-binding C1 domain. All four family members have C-terminal Src-homology 2 (SH2) and phosphotyrosine-binding (PTB) domains. The middle regions are largely unstructured with little amino acid sequence homology between family members. (B) Schematic showing various reported interaction partners of the tensin family, including: cytoplasmic domains of beta integrin subunits (β1, β3, β5, β7) with the PTB domain that reportedly supports integrin activation; the SH2 domain with p130CAS; actin to the N-terminus; the Rho GAPs DLC1-3 to both the N- and C-termini; growth factor receptors (EGFR, cMET) and additional FA proteins (e.g., FAK and p130Cas, Hic5 and ILK) to the C-terminus. (C) NIH3T3 mouse fibroblasts were transfected with the indicated GFP-tagged human tensin constructs and immunostained with antibodies against (human) TNS1, TNS2, or TNS3. Note that the respective antibodies detect only the expressed tensin member they are directed against (i.e., tensin1 antibody detects only over-expressed GFP-tensin1, tensin2 antibody only the expressed GFP-tensin2 and the tensin3 antibody only the GFP-tensin3; indicated by red boxes). (D) Expression of tensin1, 2, and 3 in U2OS and TIF cells evaluated by Western blot and immunofluorescence. A whole cell lysate from U2OS cells was run on the same gel separated by ladder lanes. After transfer, the membrane was cut and incubated with individual tensin antibodies and GAPDH separately (since tensin1, 2, and 3 have a similar molecular weight). (E) Left panel: TIF cell stained for vinculin and tensin3, note that tensin3 localises more to the centrally located adhesions and vinculin to the peripheral adhesions; right panel: TIF cell stained for tensin3 and fibronectin, note the similar localization. (F) Left panels: TIF cells were treated in suspension with blebbistatin (50 µM) or an equivalent volume of DMSO, for 60 min. Cells were fixed after spreading on fibronectin-coated glass for 60 min. Note the absence of tensin3- or vinculin-positive structures in blebbistatin-treated cells. Right panels: TIF cells cultured overnight on fibronectin-coated glass were treated with blebbistatin (50 µM) or an equivalent volume of DMSO, for 45 min prior to fixation. Scale bars in C–F indicate 10 µm. Source data are available for this figure: SourceData FS1.

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