FCHSD1 and FCHSD2 Are Expressed in Hair Cell Stereocilia and Cuticular Plate and Regulate Actin Polymerization In Vitro
Figure 4
FCHSD2, but not FCHSD1, binds to WASP and N-WASP and regulates WASP-Arp2/3-mediated F-actin polymerization in vitro.
(A) Western blots showing that FCHSD2, but not FCHSD1, was co-immunoprecipitated with WASP and N-WASP. Expression vectors were transfected into HEK293 cells to express epitope-tagged proteins, and cell lysates were subjected to immunoprecipitation. IP indicates antibody used for immunoprecipitation and WB indicates antibody used for detection. (B) WASP-Arp2/3-mediated F-actin polymerization was enhanced by GST-FCHSD2ΔC, but not by GST-FCHSD1ΔC (or GST). Monomeric actin (3 µM, 10% pyrene-labeled) was incubated with 1 µM GST-FCHSD1ΔC (or GST-FCHSD2ΔC, or GST as control) in the presence of 40 nM His-WASP145 and 20 nM Arp2/3 protein complex, and F-actin polymerization was tracked by monitoring the increase of pyrene fluorescence using a spectrofluorometer. a.u., absorbance units.