The WD protein Rack1 mediates protein kinase C and integrin-dependent cell migration.
The scaffolding protein, Rack1, is a seven-WD-domain-containing protein that has been implicated in binding to integrin beta subunit cytoplasmic domains and to members of two kinase families (src and protein kinase C, PKC) that mediate integrin bidirectional signaling. To explore the role of Rack1 in integrin function we have transfected this protein in Chinese hamster ovary (CHO) cells. We have observed no effect of Rack1 overexpression on inside-out signaling as the ligand binding properties of CHO cells also expressing constitutively active or inactive integrins were not affected. In contrast, we observed that cells stably or transiently overexpressing Rack1 had decreased migration compared to mock transfected cells. Stable Rack1 transfectants also demonstrated an increased number of actin stress fibers and focal contacts. These effects on motility and cytoskeletal organization did not appear to result from Rack1 inhibition of src function as downstream substrates of this kinase were phosphorylated normally. In addition, expression of an active src construct did not reverse the migratory deficit induced by Rack1 overexpression. On the other hand when we overexpressed a Rack1 variant with alanine substitutions in the putative PKC binding site in its third WD domain, we observed no deficit in migration. Thus the ability of Rack1 to bind, localize and stabilize PKC isoforms is likely to be involved in aspects of integrin outside-in signaling.
Buensuceso
C S
CS
Department of Vascular Biology, Scripps Research Institute-VB2, La Jolla, CA 92037, USA.
Woodside
D
D
Huff
J L
JL
Plopper
G E
GE
O'Toole
T E
TE
D000818:Animals; D016466:CHO Cells; D002465:Cell Movement / Q000502:physiology*; D006224:Cricetinae; D003599:Cytoskeleton / Q000378:metabolism; D016023:Integrins / Q000502:physiology*; D010455:Peptides / Q000378:metabolism*; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D000075370:Receptors for Activated C Kinase; D015398:Signal Transduction / Q000502:physiology; D019061:src-Family Kinases / Q000378:metabolism
D016023:Integrins; D010455:Peptides; D000075370:Receptors for Activated C Kinase; C070301:peptide I; D019061:src-Family Kinases; D011493:Protein Kinase C