Ras-guanine-nucleotide-releasing factors 1 and 2 interact with PLC gamma at focal adhesions to enable IL-1-induced Ca2+ signalling, ERK activation and MMP-3 expression

摘要

IL (interleukin)-1 signalling in anchorage-dependent cells involves focal-adhesion-restricted and Ca2+ -dependent Ras and ERK (extracellular-signal-regulated kinase) activation that leads to MMP (matrix metalloproteinase) release and extracellular matrix remodelling. Ras activity is regulated, in part, by the Ca2+ responsive Ras GRFs (guanine-nucleotide-releasing factors) 1 and 2, but the mechanisms that link and localize IL-1-induced Ca2+ signalling to focal adhesions are not defined. In the present study we characterized the role of Ras-GRF1/2 in ce, and Ras -> ERK signalling after IL-1 stimulation. By immunoprecipitation we found that Ras-GRF1/2 associates with PLC gamma 1 (phospholipase C gamma 1). This association enables PLC gamma 1 recruitment to focal adhesions and is required for Ras signalling, ERK activation and MMP-3 release downstream of IL-1 stimulation., Depletion of PLC gamma 1 by siRNA (small interfering RNA) abolished IL-1-induced Ras activation and MMP-3 expression. Buffering of cytosolic Ca2+ reduced Ras interactions with Ras-GRF1/2 and blocked MMP-3 release. The results of the present study show that, in addition to their functions as Ras-exchange factors, Ras-GRF1 and -GRF2 may act as adaptors that bind PLC gamma 1 and restrict Ca2+ signalling to the vicinity of focal adhesions, indicating a new role for these GRFs that is required for IL-1 induction of the Ras -> ERK pathway and MMP-3 expression.