Calcium Input Potentiates the Transforming Growth Factor (TGF)-β1-dependent Signaling to Promote the Export of Inorganic Pyrophosphate by Articular Chondrocyte

Abstract : Transforming growth factor (TGF)-1 stimulates extracellular PP i (ePP i) generation and promotes chondrocalcinosis, which also occurs secondary to hyperparathyroidism-induced hypercalcemia. We previously demonstrated that ANK was up-regulated by TGF-1 activation of ERK1/2 and Ca 2-dependent protein kinase C (PKC). Thus, we investigated mechanisms by which calcium could affect ePP i metabolism, especially its main regulating proteins ANK and PC-1 (plasma cell membrane gly-coprotein-1). We stimulated articular chondrocytes with TGF-1 under extracellular (eCa 2) or cytosolic Ca 2 (cCa 2) modulations. We studied ANK, PC-1 expression (quantitative RT-PCR, Western blotting), ePP i levels (radiometric assay), and cCa 2 input (fluorescent probe). Voltage-operated Ca 2-channels (VOC) and signaling pathways involved were investigated with selective inhibitors. Finally, Ank promoter activity was evaluated (gene reporter). TGF-1 elevated cCa 2 and ePP i levels (by up-regulating Ank and PC-1 mRNA/proteins) in an eCa 2 dose-dependent manner. TGF-1 effects were suppressed by cCa 2 chelation or Land T-VOC blockade while being mostly reproduced by ionomycin. In the same experimental conditions, the activation of Ras, the phosphorylation of ERK1/2 and PKC, and the stimulation of Ank promoter activity were affected similarly. Activation of SP1 (specific protein 1) and ELK-1 (Ets-like protein-1) transcription factors supported the regulatory role of Ca 2. SP1 or ELK-1 overexpression or blockade experiments demonstrated a major contribution of ELK-1, which acted synergistically with SP1 to activate Ank promoter in response to TGF-1. TGF-1 promotes input of eCa 2 through opening of Land T-VOCs, to potentiate ERK1/2 and PKC signaling cascades, resulting in an enhanced activation of Ank promoter and ePP i production in chondrocyte.
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Frederic Cailotto, Pascal Reboul, Sylvie Sébillaud, Patrick Netter, Jean-Yves Jouzeau, et al.. Calcium Input Potentiates the Transforming Growth Factor (TGF)-β1-dependent Signaling to Promote the Export of Inorganic Pyrophosphate by Articular Chondrocyte. Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2011, 286 (22), pp.19215-19228. ⟨10.1074/jbc.M110.175448⟩. ⟨hal-01704454⟩

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