Probing the integrin-actin linkage using high-resolution protein velocity mapping.
Cell migration is regulated in part by the connection between the substratum and the actin cytoskeleton. However, the very large number of proteins involved in this linkage and their complex network of interactions make it difficult to assess their role in cell migration. We apply a novel image analysis tool, spatio-temporal image correlation spectroscopy (STICS), to quantify the directed movements of adhesion-related proteins and actin in protrusions of migrating cells. The STICS technique reveals protein dynamics even when protein densities are very low or very high, and works in the presence of large, static molecular complexes. Detailed protein velocity maps for actin and the adhesion-related proteins alpha-actinin, alpha5-integrin, talin, paxillin, vinculin and focal adhesion kinase are presented. The data show that there are differences in the efficiency of the linkage between integrin and actin among different cell types and on the same cell type grown on different substrate densities. We identify potential mechanisms that regulate efficiency of the linkage, or clutch, and identify two likely points of disconnect, one at the integrin and the other at alpha-actinin or actin. The data suggests that the efficiency of the linkage increases as actin and adhesions become more organized showing the importance of factors that regulate the efficiency in adhesion signaling and dynamics.
Brown
Claire M
CM
Department of Cell Biology, University of Virginia, Charlottesville, VA, USA. claire.brown@mcgill.ca
Hebert
Benedict
B
Kolin
David L
DL
Zareno
Jessica
J
Whitmore
Leanna
L
Horwitz
Alan Rick
AR
Wiseman
Paul W
PW
D016475:3T3 Cells; D000199:Actins / Q000235:genetics / Q000378:metabolism*; D000465:Algorithms; D000818:Animals; D016466:CHO Cells; D002448:Cell Adhesion / Q000235:genetics / Q000502:physiology; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007090:Image Interpretation, Computer-Assisted / Q000379:methods; D007091:Image Processing, Computer-Assisted / Q000379:methods; D016023:Integrins / Q000235:genetics / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008954:Models, Biological; D025941:Protein Interaction Mapping / Q000295:instrumentation / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D000097573:Red Fluorescent Protein
D000199:Actins; D003598:Cytoskeletal Proteins; D049452:Green Fluorescent Proteins; D016023:Integrins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D000097573:Red Fluorescent Protein