Calreticulin modulates cell adhesiveness via regulation of vinculin expression.
Calreticulin is an ubiquitous and highly conserved high capacity Ca(2+)-binding protein that plays a major role in Ca2+ storage within the lumen of the ER. Here, using L fibroblast cell lines expressing different levels of calreticulin, we show that calreticulin plays a role in the control of cell adhesiveness via regulation of expression of vinculin, a cytoskeletal protein essential for cell-substratum and cell-cell attachments. Both vinculin protein and mRNA levels are increased in cells overexpressing calreticulin and are downregulated in cells expressing reduced level of calreticulin. Abundance of actin, talin, alpha 5 and beta 1 integrins, pp125 focal adhesion kinase, and alpha-catenin is not affected by the differential calreticulin expression. Overexpression of calreticulin increases both cell-substratum and cell-cell adhesiveness of L fibroblasts that, most surprisingly, establish vinculin-rich cell-cell junctions. Upregulation of calreticulin also affects adhesion-dependent phenomena such as cell motility (which decreases) and cell spreading (which increases). Downregulation of calreticulin brings about inverse effects. Cell adhesiveness is Ca2+ regulated. The level of calreticulin expression, however, has no effect on either the resting cytoplasmic Ca2+ concentration or the magnitude of FGF-induced Ca2+ transients. Calreticulin, however, participates in Ca2+ homeostasis as its level of expression affects cell viability at low concentrations of extracellular Ca2+. Consequently, we infer that it is not the Ca2+ storage function of calreticulin that affects cell adhesiveness. Neither endogenous calreticulin nor overexpressed green fluorescent protein-calreticulin construct can be detected outside of the ER. Since all of the adhesion-related effects of differential calreticulin expression can be explained by its regulation of vinculin expression, we conclude that it is the ER-resident calreticulin that affects cellular adhesiveness.
Opas
M
M
Department of Anatomy and Cell Biology, University of Toronto, Ontario, Canada. m.opas@utoronto.ca
Szewczenko-Pawlikowski
M
M
Jass
G K
GK
Mesaeli
N
N
Michalak
M
M
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000235:genetics*; D037282:Calreticulin; D002448:Cell Adhesion / Q000502:physiology; D002460:Cell Line / Q000737:chemistry / Q000166:cytology / Q000502:physiology; D002465:Cell Movement / Q000502:physiology; D048429:Cell Size; D003593:Cytoplasm / Q000737:chemistry; D003599:Cytoskeleton / Q000502:physiology; D004721:Endoplasmic Reticulum / Q000737:chemistry; D005347:Fibroblasts / Q000737:chemistry / Q000166:cytology* / Q000502:physiology; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000502:physiology; D016023:Integrins / Q000032:analysis; D007365:Intercellular Junctions / Q000737:chemistry; D051379:Mice; D018832:Molecular Chaperones / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D012261:Ribonucleoproteins / Q000235:genetics*; D014162:Transfection; D016596:Vinculin / Q000235:genetics*
D002135:Calcium-Binding Proteins; D037282:Calreticulin; D016023:Integrins; D018832:Molecular Chaperones; D012333:RNA, Messenger; D012261:Ribonucleoproteins; D016596:Vinculin; D002118:Calcium