Raf kinase inhibitor protein positively regulates cell-substratum adhesion while negatively regulating cell-cell adhesion.
Raf kinase inhibitor protein (RKIP) regulates a number of cellular processes, including cell migration. Exploring the role of RKIP in cell adhesion, we found that overexpression of RKIP in Madin-Darby canine kidney (MDCK) epithelial cells increases adhesion to the substratum, while decreasing adhesion of the cells to one another. The level of the adherens junction protein E-cadherin declines profoundly, and there is loss of normal localization of the tight junction protein ZO-1, while expression of the cell-substratum adhesion protein beta1 integrin dramatically increases. The cells also display increased adhesion and spreading on multiple substrata, including collagen, gelatin, fibronectin and laminin. In three-dimensional culture, RKIP overexpression leads to marked cell elongation and extension of long membrane protrusions into the surrounding matrix, and the cells do not form hollow cysts. RKIP-overexpressing cells generate considerably more contractile traction force than do control cells. In contrast, RNA interference-based silencing of RKIP expression results in decreased cell-substratum adhesion in both MDCK and MCF7 human breast adenocarcinoma cells. Treatment of MDCK and MCF7 cells with locostatin, a direct inhibitor of RKIP and cell migration, also reduces cell-substratum adhesion. Silencing of RKIP expression in MCF7 cells leads to a reduction in the rate of wound closure in a scratch-wound assay, although not as pronounced as that previously reported for RKIP-knockdown MDCK cells. These results suggest that RKIP has important roles in the regulation of cell adhesion, positively controlling cell-substratum adhesion while negatively controlling cell-cell adhesion, and underscore the complex functions of RKIP in cell physiology.
Department of Chemistry, University of Connecticut, Storrs, Connecticut 06269, USA.
Kevin T
KT
Mc Henry
Roberto
R
Montesano
Shoutian
S
Zhu
Anwar B
AB
Beshir
Hui-Hui
HH
Tang
Kam C
KC
Yeung
Gabriel
G
Fenteany
D022005:Adherens Junctions / Q000378:metabolism*; D000818:Animals; D001943:Breast Neoplasms / Q000378:metabolism / Q000473:pathology; D015820:Cadherins / Q000378:metabolism; D002465:Cell Movement / Q000187:drug effects; D021982:Cell-Matrix Junctions / Q000187:drug effects / Q000378:metabolism*; D004285:Dogs; D015536:Down-Regulation; D004847:Epithelial Cells / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D019012:Integrin beta1 / Q000378:metabolism; D007680:Kidney Neoplasms / Q000378:metabolism / Q000473:pathology; D008565:Membrane Proteins / Q000378:metabolism; D023303:Oxazolidinones / Q000494:pharmacology; D049488:Phosphatidylethanolamine Binding Protein / Q000378:metabolism* / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000378:metabolism* / Q000494:pharmacology; D034622:RNA Interference; D014407:Tumor Cells, Cultured; D015854:Up-Regulation; D014945:Wound Healing / Q000187:drug effects; D062826:Zonula Occludens-1 Protein; D048490:raf Kinases / Q000037:antagonists & inhibitors*
D015820:Cadherins; D016326:Extracellular Matrix Proteins; D019012:Integrin beta1; D008565:Membrane Proteins; D023303:Oxazolidinones; D049488:Phosphatidylethanolamine Binding Protein; D010750:Phosphoproteins; D047428:Protein Kinase Inhibitors; C569042:TJP1 protein, human; D062826:Zonula Occludens-1 Protein; C529490:locostatin; D048490:raf Kinases