Impact of the Thr-184 site on localization to filopodia structures in COS7 cells.
A, GFP-tagged full-length Myo3A localizes along the length of filopodia in COS7 cells, unlike GFP-MYO3A ΔK (lacking the kinase domain) that shows concentration at filopodial tips. Neither the T184A nor the T184E mutations in GFP-MYO3A result in increased tip localization. Representative images are epifluorescence of paraformaldehyde-fixed and phalloidin-stained COS7 cells. GFP constructs are shown in cyan, and Alexa 568-phalloidin is shown in yellow. All images were captured under identical imaging conditions and scaled identically for brightness. Scale bar, 10 μm. B, T184A and T184E mutations do not alter the tip localization of MYO3A as indicated by the ratio of tip to cell body fluorescence. C, similarly, expression of GFP-MYO3A-T184A and GFP-MYO3A-T184E did not increase the density of filopodia along the cell periphery. B, n >46 filopodial tips from 10 or more cells per condition; C, n >24 cells per condition. * indicates significant difference (p < 0.003, Tukey test) from all other constructs.