ABSTRACT
Filopodia, dynamic membrane protrusions driven by polymerization of an actin filament core, can adhere to the extracellular matrix and experience both external and cell-generated pulling forces. The role of such forces in filopodia adhesion is however insufficiently understood. Here, we study filopodia induced by overexpression of myosin X, typical for cancer cells. The lifetime of such filopodia positively correlates with the presence of myosin IIA filaments at the filopodia bases. Application of pulling forces to the filopodia tips through attached fibronectin-coated laser-trapped beads results in sustained growth of the filopodia. Pharmacological inhibition or knockdown of myosin IIA abolishes the filopodia adhesion to the beads. Formin inhibitor SMIFH2, which causes detachment of actin filaments from formin molecules, produces similar effect. Thus, centripetal force generated by myosin IIA filaments at the base of filopodium and transmitted to the tip through actin core in a formin-dependent fashion is required for filopodia adhesion.
Subject(s)
Formins/metabolism , Myosins/metabolism , Neoplasms/metabolism , Nonmuscle Myosin Type IIA/metabolism , Pseudopodia/physiology , Actin Cytoskeleton , Animals , COS Cells , Chlorocebus aethiops , Formins/antagonists & inhibitors , Formins/genetics , Formins/ultrastructure , Gene Expression Regulation, Neoplastic , HeLa Cells , Humans , Microfilament Proteins , Nonmuscle Myosin Type IIA/antagonists & inhibitors , Nonmuscle Myosin Type IIA/genetics , Nonmuscle Myosin Type IIA/ultrastructure , Pseudopodia/pathology , Thiones/pharmacology , Uracil/analogs & derivatives , Uracil/pharmacologyABSTRACT
Chromosome movement during mitosis is powered in part by energy released through the depolymerization of kinetochore microtubules (MTs). Strong but indirect evidence suggests the existence of a specialized coupling between kinetochores and MT plus ends that enables this transduction of chemical energy into mechanical work. Analysis of this phenomenon is important for learning how energy is stored within the MT lattice, how it is transduced, and how efficient the process can be, given coupling devices of different designs. Here we use a recently developed molecular-mechanical model of MTs to examine the mechanism of disassembly dependent force generation. Our approach is based on changes in tubulin dimer conformation that occur during MT disassembly. We find that all of the energy of polymerization-associated GTP hydrolysis can be stored as deformations of the longitudinal bonds between tubulin dimers, and its optimal use does not require the weakening of lateral bonds between dimers. Maximum utilization of this stored energy and, hence, the generation of the strongest possible force, is achieved by a protofilament power-stroke mechanism, so long as the coupling device does not restrict full dissociation of the lateral bonds between tubulin dimers.
