FIGURE 5:

Mesendoderm keratin filament dynamic exchange is decreased by 14-3-3 inhibition. (A) Immunoblot analyses of protein extracts (1% Triton X-100) of stage 10.5 Xenopus embryos expressing mCh-R18 or mCh-R18M. (B) Coimmunoprecipitation (1% Tergitol type NP-40) performed using stage 10.5 lysates expressing human FLAG-14-3-3 β with either mCherry-R18 or mCherry-R18M. (C, D) Still images from photobleach and recovery time lapse movies (Supplemental Movie S1). Mesendoderm explants expressing either 14-3-3 inhibitor peptide mCherry-R18 or control peptide mCherry-R18M (red) with eGFP-K19 (green) were exposed to GFP photobleaching and fluorescence recovery at the site was measured. (C’–D’’) Enlarged view of the region of filament bleaching (white boxes) during recovery measurements. The time annotations in seconds refer to start of capture (t = 0 s) and end of capture (t = 330 s). (E) Representative analysis plotting fluorescence recovery against time of image capture. (F) Comparison of mean eGFP-K19 recovery rate in explants expressing either mCherry-R18 or mCherry-R18M. Analysis was performed using a one-tailed t test, *p < 0.05. Error bars are ± SEM. Scale bars are 10 μm.