Latrunculin A Treatment Causes Both CRIB Dispersal from Cell Tips and Sty1 MAP Kinase Activation
(A) Still images from movies of Lifeact-mCherry (LA-mCh) and CRIB-3xmCitrine (CRIB-3mCit) in wild-type cells after addition of DMSO or 50 μM latrunculin A (LatA). Asterisks indicate dispersal of CRIB from cell tips. Arrowheads indicate examples of ectopic CRIB patches after dispersal. Nuclear CRIB signal is unrelated to Cdc42 (see the Supplemental Experimental Procedures).
(B) Anti-phospho-Sty1 western blot of wild-type cell extracts after addition of DMSO, 50 μM LatA, or 0.6 M KCl for the indicated times. Ponceau S stain of the same region of the blot is shown below.
(C) Quantification of phospho-Sty1 (a.u.) from experiments of the type shown in (B). Mean values are from three independent experiments. Error bars indicate SEM.
(D) Still images from movies of Sty1-mECitrine in wild-type cells after addition of DMSO or LatA. LatA addition leads to net import of Sty1-mECitrine into the nucleus.
(E) Quantification of Sty1-mECitrine nuclear fluorescence after addition of DMSO or LatA. Values shown are mean relative intensity (a.u.) of nuclear fluorescence above cytoplasmic fluorescence. Error bars indicate SD.
All times shown are relative to addition of DMSO, LatA, or KCl. Scale bars, 5 μm. See also Figure S1 and Movie S1.