We investigated the signal transduction pathway(s) of leukotriene D4 (LTD4) in the human promonocytic U937 cells, a cell line known to constitutively express CysLT1 receptors. Herein, we demonstrate that LTD4 specifically acts on a CysLT1 receptor to dose-dependently increase (3-5 fold over basal) RasGTP through a Gi/o protein. In fact, while cytosolic Ca2+ ([Ca2+]i) increase was only partially sensitive to pertussis toxin (PTx), Ras activation was almost completely inhibited by the same toxin. Furthermore, the phospholipase C (PLC) inhibitor U73122 completely inhibited both [Ca2+]i and RasGTP increase, suggesting that in these cells PLC is the point of convergence for both PTx insensitive and sensitive pathways leading to [Ca2+]i release and Ras activation. Indeed, chelating intracellular Ca2+ strongly (>70%) prevented LTD4-induced Ras activation, indicating that this ion plays an essential role for CysLT1-induced downstream signaling in differentiated U937 (dU937) cells. In addition, while Src did not appear to be substantially involved in CysLT1-induced signaling, genistein was able to partially inhibit LTD4-induced [Ca2+]I transient (~34%) and almost completely prevented Ras activation (>90%), suggesting a potential role for other Ca2+-dependent tyrosine kinases in LTD4-induced signaling. Finally, agonist-induced CysLT1 stimulation was followed by a specific extracellular regulated kinase (ERK) 1/2 phosphorylation, an event with a pharmacological profile similar to that of Ras activation, partially (~40%) sensitive to Clostridium sordellii lethal toxin and totally blocked by PTx. In conclusion, LTD4-induced CysLT1 receptor activation in dU937 cells leads to Ras activation and ERK phosphorylation mostly through a PTx-sensitive Gi/o protein, PLC, and Ca2+-dependent tyrosine kinase(s).
|Titolo:||CysLT1 signal transduction in differentiated U937 cells involves the activation of the small GTP-binding protein Ras|
|Autori interni:||ROVATI, GIANENRICO (Ultimo)|
CAPRA, VALERIE CAROLINE (Primo)
ACCOMAZZO, MARIA ROSA
|Parole Chiave:||[Ca 2+]i; cys-LTs; Cysteine-containing leukotrienes; Cytosolic Ca2+ concentration; Differentiated U937; dU937; G protein-coupled receptors; GPCR; Leukotriene D4; LTD4; Pertussis toxin; PLC; PTx|
|Settore Scientifico Disciplinare:||Settore BIO/14 - Farmacologia|
|Data di pubblicazione:||2004|
|Digital Object Identifier (DOI):||10.1016/j.bcp.2003.12.027|
|Appare nelle tipologie:||01 - Articolo su periodico|
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