Effect of the T(D/E)₅XE region of Fhod3 on actin assembly.

<p>(A) Effect of the Fhod3 T(D/E)₅XE region encoded by exon 25 on <i>in vivo</i> actin assembly. HeLa cells were transfected with plasmids encoding Fhod3-ΔN (wt) (amino acids 931–1,586) with or without the T(D/E)₅XE region in the FH2 domain; or plasmids encoding Fhod3-ΔN (I1127A) with or without the T(D/E)₅XE region. Cells were fixed and visualized by GFP-fluorescence (green) or phalloidin staining (red). Scale bar, 10 µm. (B) SDS-PAGE analysis of purified proteins used in an actin polymerization assay. Purified proteins were subjected to 10% SDS-PAGE and stained with <i>Coomassie Brilliant Blue</i>. Positions for marker proteins are indicated in kDa. (C) Effect of the T(D/E)₅XE region of Fhod3 on <i>in vitro</i> actin assembly. G-actin (10% pyrene-labeled) at 2 µM was incubated with 50 nM Fhod3-T(D/E)₅XE(+)-ΔN (wt), Fhod3-T(D/E)₅XE(+)-ΔN (I1127A), Fhod3-T(D/E)₅XE(−)-ΔN (wt), or mDia1-FH1FH2 (amino acids 549–1,175) in the presence of 2 µM profilin I.