Fig. 3

Hax1 recruits IQGAP1 to microtubules and the leading edge of migrating cells. A, MCF7 cells were transfected with DsRed-Hax1 and labeled with antibodies against tubulin (green), and IQGAP1 (magenta). Scale bar, 10 μm. Note that white arrows indicate the membrane ruffle-like localization of Hax1 is superimposed onto that of the IQGAP1 signal and the three channels are merged. B, Immunofluorescence staining for tubulin (green) and IQGAP1 (red) in MCF7 cells transfected with the indicated siRNAs and followed with or without treatment of nocodazole (10 μM) for 30 min. Cells were imaged by super-resolution N-SIM microscopy. The boxed areas are magnified as insets. Scale bar, 10 μm. C, Co-localization of IQGAP1 and MT (tubulin) was determined by Pearson correlation coefficient (n = 20 cells for each). Note that Hax1 localization with MT is reduced significantly in both nocodazole treated cells and Hax1-depleted cells (p < 0.0001). D, The interaction with microtubules (MT) was examined by co-sedimentation assay. The presence of proteins in the original lysate (Input), pellet of ultracentrifugation, as well as the supernatant (Sup) was determined by immunoblot with anti-IQGAP1 antibody