Actin-dependent Lamellipodia Formation and Microtubule-dependent Tail Retraction Control-directed Cell Migration

Abstract

Migrating cells are polarized with a protrusive lamella at the cell front followed by the main cell body and a retractable tail at the rear of the cell. The lamella terminates in ruffling lamellipodia that face the direction of migration. Although the role of actin in the formation of lamellipodia is well established, it remains unclear to what degree microtubules contribute to this process. Herein, we have studied the contribution of microtubules to cell motility by time-lapse video microscopy on green flourescence protein-actin-and tubulin-green fluorescence protein-transfected melanoma cells. Treatment of cells with either the microtubule-disrupting agent nocodazole or with the stabilizing agent taxol showed decreased ruffling and lamellipodium formation. However, this was not due to an intrinsic inability to form ruffles and lamellipodia because both were restored by stimulation of cells with phorbol 12-myristate 13-acetate in a Rac-dependent manner, and by stem cell factor in melanoblasts expressing the receptor tyrosine kinase c-kit. Although ruffling and lamellipodia were formed without microtubules, the microtubular network was needed for advancement of the cell body and the subsequent retraction of the tail.In conclusion, we demonstrate that the formation of lamellipodia can occur via actin polymerization independently of microtubules, but that microtubules are required for cell migration, tail retraction, and modulation of cell adhesion.

Paxillin: Adapting to Change

“…The process of directed migration can be broken down into several discrete steps, namely, protrusion/polarization, attachment/adhesion assembly, traction/translocation, and adhesion disassembly/retraction (12,98,252,300,301). After cellular stimulation with a chemotactic growth factor, the cell extends a protrusion resulting from the polymerization of a dense network of actin toward the chemoattractant in the form of fingerlike filopodial projections and thin lamellipodial sheets.…”

Regulation of Sustained Actin Dynamics by the TCR and Costimulation as a Mechanism of Receptor Localization

“…␤-actin-GFP and ␤5-tubulin-GFP (18,19) were expressed from a Moloney murine leukemia virus-derived retroviral expression vector (20) containing an additional internal phosphoglycerol kinase promoter or not, respectively. T cells were primed on day 0 with lymph node APCs plus 3 M moth cytochrome C (MCC) peptide, retrovirally transduced as described in Costa et al (20) on day 1, FACS sorted on day 5, and used for microscopy between days 7 and 9.…”

“…As an indicator of T cell polarity, we next visualized the T cell MTOC in the interaction of live, primary 5C.C7 T cells with activating APCs in real time using a retrovirally expressed tubulin-GFP fusion protein (19). Initially, we used an optimal T cell stimulus ( Fig.…”

IQGAP1 Promotes Neurite Outgrowth in a Phosphorylation-dependent Manner

“…More recent evidence reveals that phosphorylation is essential for Ena/VASP function in cell movement (19), and the activity of Rho is regulated by tyrosine phosphorylation of RhoGEFs (20). Moreover, phorbol esters modulate cell motility via Rac, Cdc42, and ezrin (21)(22)(23), and Hck-induced tyrosine phosphorylation of Wiskott-Aldrich syndrome protein results in filopodia (24). In this work we demonstrate that endogenous IQGAP1 is highly phosphorylated in cells and that IQGAP1 is a target for protein kinase C (PKC).…”