Fig. 9

RhoH co-localizes and interacts with Rac1. a RhoH partially co-localizes with Rac1. Cells were seeded on Matrigel-coated coverslips and transfected with GFP-Rac1 and/or HA-RhoH as indicated. After 24 h cells were fixed and stained for F-actin with Alexa Fluor 633 phalloidin, for anti-HA antibodies for HA-RhoH and for DNA with DAPI. Images were acquired by confocal microscopy. Scale bar 10 μm. b, c RhoH promotes cell elongation, which is not affected by PAK2. 48 h after siRNA transfection with siPAK2-2/3 or control siRNA, cells were transfected with GFP or GFP-RhoH. Cells were fixed and stained for F-actin with Alexa Fluor 546 phalloidin and DNA with DAPI. Cell elongation factor and shape factor were measured using ImageJ from F-actin-stained images. Scatter dot plots show mean +/− SEM; cells were analysed from three independent experiments, n.s. non-significant, *p ≤ 0.05, Student’s t test. d Rac1 and RhoH co-immunoprecipitate. COS7 cells were transfected with constructs encoding GFP, HA-RhoH and/or GFP-Rac1 as indicated. After 24 h, lysates were immunoprecipitated with anti-GFP and analysed by western blotting with the indicated antibodies