Abstract
Calreticulin is an endoplasmic reticulum Ca2+-storage protein, which influences gene expression and cell adhesion. In this study, we show that calreticulin induces fibronectin gene expression and matrix deposition, leading to differences in cell spreading and focal adhesion formation in cells differentially expressing calreticulin. We further show that these effects of calreticulin occur via a c-Src-regulated pathway and that c-Src activity is inversely related to calreticulin abundance. Since c-Src is an important regulator of focal contact turnover, we investigated the effect of c-Src inhibition on cells differentially expressing calreticulin. Inhibition of c-Src rescued the poorly adhesive phenotype of the calreticulin-underexpressing cells in that they became well spread, commenced formation of numerous focal contacts, and deposited a rich fibronectin matrix. Importantly, we show that c-Src activity is dependent on releasable Ca2+ from the endoplasmic reticulum, thus implicating Ca2+-sensitive pathways that are affected by calreticulin in cell-substratum adhesion. We propose that calreticulin affects fibronectin synthesis and matrix assembly via the regulation of fibronectin gene expression. In parallel, calcium-dependent effects of calreticulin on c-Src activity influence the formation and/or stability of focal contacts, which are instrumental in matrix assembly and remodeling.
Highlights
Capacity and its effects on both the SERCA pumps and inositol 1,4,5-trisphosphate receptors [2, 3]
Thapsi- Our particular focus was on fibronectin, a major component gargin was used to reduce the concentration of Ca2ϩ in the of integrin-based cell substratum adhesions
Cells were treated with 1 M thapsigargin for 30 min at we found that cells differentially expressing calreticulin dif
Summary
Capacity and its effects on both the SERCA pumps and inositol 1,4,5-trisphosphate receptors [2, 3]. Fibronectin is bound to and regulated by cells at specific sites of cell-substratum adhesions that serve to link the ECM to the actin cytoskeleton via transmembrane integrin heterodimers [12]. Forces due to integrin binding to ECM ligands cause the bundling of the actin cytoskeleton into tension-generating stress fibers This tension causes the clustering of integrins and associated cytoskeletal proteins, such as vinculin, paxillin, talin, c-Src, and focal adhesion kinase (FAK), into strong cell substratum adhesions termed focal contacts (or focal adhesions) [8, 12,13,14]. Tyrosine phosphorylation has been shown to regulate cell-substratum adhesion [17,18,19,20]; the aim of this study was to investigate the role of protein tyrosine kinases, c-Src, in the regulation of cell-substratum adhesion in cells differentially expressing calreticulin. A JUNE 1, 2007 VOLUME 282 NUMBER 22
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