Spatial regulation of CLASP affinity for microtubules by Rac1 and GSK3beta in migrating epithelial cells.
Proteins that in cells specifically bind to growing microtubule plus ends (+TIPs) are thought to play important roles in polarization of the cytoskeleton. However, most +TIPs do not show a bias of their microtubule-binding behavior toward different subcellular regions. Here, we examine the dynamics of the +TIP CLASP in migrating PtK1 epithelial cells. We find that, although CLASPs track microtubule plus ends in the cell body, they dynamically decorate the entire microtubule lattice in the leading edge lamella and lamellipodium. Microtubule lattice binding is mediated by the COOH-terminal region of the CLASP microtubule-binding domain and is regulated downstream of Rac1. Phosphorylation of sites in the NH2-terminal part of the microtubule-binding domain by glycogen synthase kinase 3beta likely regulates the affinity of CLASPs for microtubule lattices. These results demonstrate the striking difference of the microtubule cytoskeleton in the lamella as compared with the cell body and provide the first direct observation of subcellular regulation of a microtubule-associated protein in migrating cells.
Wittmann
Torsten
T
Department of Cell Biology, The Scripps Research Institute, La Jolla, CA 92037, USA. twittman@scripps.edu
Waterman-Storer
Clare M
CM
D000818:Animals; D001665:Binding Sites / Q000502:physiology; D002460:Cell Line; D002465:Cell Movement / Q000502:physiology*; D016764:Cell Polarity / Q000502:physiology; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D009687:Nuclear Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D011554:Pseudopodia / Q000378:metabolism / Q000648:ultrastructure; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*
C497851:CLASP2 protein, human; C469747:Clasp protein, mouse; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D038362:Glycogen Synthase Kinase 3; D020830:rac1 GTP-Binding Protein