Microtubule-targeting-dependent reorganization of filopodia

Interaction between the microtubule system and actin cytoskeleton has emerged as a fundamental process required for spatial regulation of cell protrusion and retraction activities. In our current studies, analysis of digital fluorescence images revealed targeting of microtubules to filopodia in B16F1 melanoma cells and fibroblasts. We investigated the functional consequence of targeting on filopodia reorganization and examined mechanisms by which microtubules may be guided to, or interact with, filopodia. Live cell imaging studies show that targeting events in lamellipodia wings temporally correlated with filopodia turning toward the lamellipodium midline and with filopodia merging. Rapid uncoupling of targeting with nocodazole decreased filopodia merging events and increased filopodia density. Total internal reflection fluorescence microscopy identified microtubules near the ventral surface and upward movement of targeted filopodia. The role of adhesion sites and microtubule plus-end proteins in targeting was investigated. Correlation of adhesion sites with microtubule targeting to filopodia was not observed and depletion of microtubule plus-end proteins did not significantly alter targeting frequency. We propose that microtubules target filopodia, independent of focal adhesions and plus-end proteins, causing filopodia movement and microtubules regulate filopodia density in lamellipodia wings through filopodia merging events.