Characterization of human fidgetin and its functions during mitosis and cell migration

Abstract

Fidgetin is a member of the 'Meiotic' subfamily or Subfamily 7 of AAA (ATPase Associated with diverse cellular Activities) protein super family. Analyses of Fidgetin mutant mice have revealed important developmental roles for the protein though its specific cellular and catalytic activities have largely remained unknown. Here, we show that human Fidgetin is a potent microtubule severing protein and depolymerase used to regulate microtubule dynamics and organization throughout the cell cycle. In vitro, purified, baculovirus expressed human Fidgetin promotes the breakage of taxol-stabilized microtubules along their length as well as shortening from their ends. In cells, human Fidgetin targets to centrosomes and, during mitosis, its depletion with siRNA significantly reduces the velocity at which tubulin subunits flux poleward through spindle microtubules and anaphase A chromatid-to-pole motion. Fidgetin also regulates centrosome size, content and the number of astral microtubules within metaphase spindles. Similarly, interphase cells lacking Fidgetin become enlarged with an increased density in centrosome-associated microtubules and display reduced rates of migration as determined by a wound-healing assay. Based on these data, we propose that human Fidgetin is utilized to release microtubules from their centrosomal-nucleation sites, thereby maintaining the equilibrium between the centrosomal and non-centrosomal microtubule population within the cells.