Journal: Molecular Biology of the Cell
Article Title: Actin-binding Protein-1 Interacts with WASp-interacting Protein to Regulate Growth Factor-induced Dorsal Ruffle Formation
doi: 10.1091/mbc.E09-02-0106
Figure Lengend Snippet: mAbp localizes with WIP at dorsal ruffles. (A) GFP-mAbp1 and mCherry-WIP were transiently transfected into NIH-3T3 cells. Cells were plated on FN-coated, glass-bottomed dishes and serum-starved. Cells were stimulated with PDGF and subsequently analyzed by dual, time-lapse fluorescence microscopy. Montages are representative of at least three independent experiments. Bar, 10 μm. See accompanying Video 5. (B) GFP-WIP was transiently transfected into NIH-3T3 cells. Cells were plated on FN-coated coverslips, serum-starved, and stimulated with PDGF. Cells were fixed and stained with rhodamine phalloidin and anti-cortactin antibody (top panel) or anti-mAbp1 and anti-cortactin antibodies (bottom panel). Bar, 10 μm. (C) NIH-3T3 cells were transiently transfected with GFP, GFP-C-terminal mAbp1, or GFP-C-terminal mAbp1 and mCherry WIP. Cells were plated on FN-coated coverslips, serum-starved, and stimulated with PDGF. Cells were fixed and stained with anti-cortactin antibody. A representative image of a GFP-C-terminal mAbp1 coexpressing mCherry-WIP is shown (left). Bar, 10 μm. Dorsal ruffles were quantified by counting the number of transfected cells containing at least one dorsal ruffle after stimulation with PDGF. Greater than 50 cells were counted per condition, and each condition represents the average value from three independent experiments; error bars, SEM; *p < 0.01 compared with GFP control by one-way ANOVA.
Article Snippet: Live imaging of cells expressing GFP-mAbp1 and mCherry-WIP was performed using a 60× objective on a Nikon Eclipse TE300 inverted microscope (Melville, NY).
Techniques: Transfection, Fluorescence, Microscopy, Staining
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