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Figure 5 | Cell Communication and Signaling

Figure 5

From: NHERF2 is crucial in ERM phosphorylation in pulmonary endothelial cells

Figure 5

NHERF2 overexpression induces EC filopodia formation and spreading. (A) Lysates of BPAEC without transfection (ctr) or transfected with pCMV-HA wild type or mutant NHERF2 constructs were analyzed by Western blot using antibodies against phospho-ERM, ERM, and HA-tag. A representative Western blot is shown. Protein levels were quantified by densitometric analysis. Phospho-ERM protein levels were normalized against ERM protein levels. Bars represent mean ± SE. Significant changes, determined by Student’s t- test, are indicated by asterisks; **(P < 0.01), n = 6. (B) Left panel: Immunofluorescent staining of recombinant wild type or mutant NHERF2 overexpressing BPAEC was performed using anti-HA primary antibody (green). Actin microfilaments were stained with Texas Red conjugated phalloidin (red). Scale bars: 10 μm. Right panel: Wild type NHERF2 transfected BPAEC co-stained for phospho-ERM and HA-tag is shown. Arrows indicate co-localization of phosho-ERM and overexpressed NHERF2 on the merged image. (C): Non transfected (control), wild type (wt) and mutant (mu) NHERF2 transfected cells (left panel) or non-siRNA and NHERF2 specific siRNA treated cells (right panel) were plated onto 8W10E arrays. Spreading and attachment of cells were followed in time by ECIS measurement. Results are presented as means ± SD at least of four chambers for each sample.

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