Phosphorylation of the CAS N-terminus reduces the nuclear retention of CAS, which colocalizes with F-actin. (A) Immunostaining of Kapβ1 in cells expressing mCherry-CAS in the presence and absence of oxidative stress (H2O2) and MEK1 inhibitor. (B) Oxidative stress reduces the N/C ratio of mCherry-CAS. This is rescued by adding MEK1 inhibitor, which has a null effect in the absence of H2O2. Box plots denote the median, first, and third quartiles. The whiskers represent the minimum and maximum values. N = 3. The data were tested for normality using a built-in function of GraphPad Prism software. P values were obtained using an unpaired two-tailed t test. (C) Representative images of HEK293T cells expressing mCherry-CAS, mCherry-CAS_T18G, or mCherry-CAS_T18D. Filamentous actin (F-actin) is stained with mEos2-Lifeact-7 peptide (Addgene 54809; green). mCherry-CAS_T18D colocalizes with F-actin at invadopodia but does not show enrichment in the nucleus. Scale bar, 5 µm.
