Rac regulates the interaction of fascin with protein kinase C in cell migration.
Fascin is an actin-bundling protein that is low or absent in normal epithelia; its upregulation correlates with poor prognosis in many human carcinomas. We have recently demonstrated in mouse xenograft models that fascin contributes to tumour development and metastasis through its dual actin-bundling and active PKC-binding activities. Rac was implicated as a regulator of fascin-dependent colon carcinoma cell migration in vitro. Here, we tested the hypothesis that Rac regulates the interaction of fascin with active PKC. The major conventional PKC in colon carcinoma cells is protein kinase Cgamma (PKCgamma). Endogenous PKCgamma, fascin and Rac1 colocalised at lamellipodial margins of migrating cells. Colocalisation of fascin and PKCgamma depended on Rac activity, and inhibition of Rac decreased PKCgamma activity in cell extracts but not in vitro. Fluorescence resonance energy transfer/fluorescence lifetime imaging microscopy uncovered that fascin and PKCgamma interact in protrusions and filopodia of migrating cells. Mechanistically, the interaction depended on phosphorylated fascin, active PKCgamma and active Rac, but not on active Cdc42. The activity of Rac on the fascin/PKC complex was mediated in part by Pak. Elucidation of this novel pathway for regulation of the fascin/PKCgamma complex in migrating carcinoma cells suggests novel targets for therapeutic intervention in metastasis.
Parsons
Maddy
M
Randall Division of Cell and Molecular Biophysics, King's College London, Guy's Campus, London SE1 1UL, UK. maddy.parsons@kcl.ac.uk
Adams
Josephine C
JC
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement; D022081:Cell Surface Extensions / Q000201:enzymology; D003110:Colonic Neoplasms / Q000201:enzymology; D004789:Enzyme Activation; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism*; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism*; D021381:Protein Transport; D014981:Xenopus; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D054462:p21-Activated Kinases / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism*
D002352:Carrier Proteins; D008840:Microfilament Proteins; C020324:fascin; C087857:protein kinase C gamma; D054462:p21-Activated Kinases; D011493:Protein Kinase C; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins