Abstract
Diverse signaling pathways have been proposed to regulate store-operated calcium entry (SOCE) in a wide variety of cell types. However, it still needs to be determined if all of these known pathways operate in a single cell type. In this study, we examined involvement of various signaling molecules in SOCE using human fibroblast cells (HSWP). Bradykinin (BK)-stimulated Ca2+ entry, previously shown to be via SOCE, is enhanced by the addition of vanadate, an inhibitor of tyrosine phosphatases. Furthermore, SOCE is regulated by cytochrome P-450, as demonstrated by the fact that the products of cytochrome P-450 activity (14,15 EET) stimulated SOCE while econazole, an inhibitor of cytochrome P450, suppressed BK-stimulated Ca2+ entry. In contrast, Ca2+ entry was unaffected by the guanylate cyclase inhibitor LY83583, or the membrane permeant cyclic GMP analog 8-bromo-cyclic GMP (8-Br-cGMP). Neither nitric oxide donors nor phorbol esters affected BK-stimulated Ca2+ entry. SOCE in HSWP cells is primarily regulated by tyrosine phosphorylation and the cytochrome P-450 pathway, but not by cyclic GMP, nitric oxide, or protein kinase C. Thus, multiple pathways do operate in a single cell type leading to the activation of Ca2+ entry and some of these signaling pathways are more prominently involved in regulating calcium entry in different cell types.
Highlights
One of the early events following stimulation of many G protein coupled receptors is an increase in the intracellular Ca2+ concentration ([intracellular free calcium concentration (Ca2+]i)) (Bird et al, 2004), which initiates a cascade of signal transduction events often leading to DNA synthesis and cell proliferation (Parekh and Putney, 2005)
Since the level of tyrosine phosphorylation is determined by the activity of tyrosine kinases and tyrosine phosphatases, we investigated whether modulation of cellular tyrosine phosphatase activity affects the Ca2+ entry induced by BK, as well as the level of tyrosine phosphorylation
We have examined the role of intracellular signaling molecules in regulating store-operated calcium entry (SOCE) and protein tyrosine phosphorylation in human fibroblast cells
Summary
One of the early events following stimulation of many G protein coupled receptors is an increase in the intracellular Ca2+ concentration ([Ca2+]i) (Bird et al, 2004), which initiates a cascade of signal transduction events often leading to DNA synthesis and cell proliferation (Parekh and Putney, 2005). The peak is due to the release of intracellular Ca2+ from Ins(1,4,5)P3 -sensitive internal stores and the sustained plateau is due to Ca2+ entry from the extracellular space. A direct interaction between store-operated channels and IP3 receptors, the insertion of sub-membrane vesicles containing store-operated channels into the plasma membrane, or a secretion-like coupling may be involved in regulation of SOCE
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