P2 receptor networks regulate signaling duration over a wide dynamic range of ATP concentrations

Abstract

SummaryThe primordial intercellular signaling molecule ATP acts through two families of cell-surface P2 receptors -the P2Y family of Gprotein-coupled receptors and the P2X family of ligand-gated cation channels. Multiple P2 receptors are expressed in a variety of cell types. However, the significance of these networks of receptors in any biological system remains unknown. Using osteoblasts as a model system, we found that a low concentration of ATP (10 mM, ATP low ) induced transient elevation of cytosolic Ca 2+ , whereas a high concentration of ATP (1 mM, ATP high ) elicited more sustained elevation. Moreover, graded increases in the Ca 2+ signal were achieved over a remarkable million-fold range of ATP concentrations (1 nM to 1 mM). Next, we demonstrated that ATP low caused transient nuclear localization of the Ca 2+ -regulated transcription factor NFATc1; whereas, ATP high elicited more sustained localization. When stimulated with ATP high , osteoblasts from P2X7 loss-of-function mice showed only transient Ca 2+ -NFATc1 signaling; in contrast, sustained signaling was observed in wild-type cells. Additional experiments revealed a role for P2Y receptors in mediating transient signaling induced by low ATP concentrations. Thus, distinct P2 receptors with varying affinities for ATP account for this wide range of sensitivity to extracellular nucleotides. Finally, ATP high , but not ATP low , was shown to elicit robust expression of the NFAT target gene Ptgs2 (encoding COX-2), consistent with a crucial role for the duration of Ca 2+ -NFAT signaling in regulating target gene expression. Taken together, ensembles of P2 receptors provide a mechanism by which cells sense ATP over a wide concentration range and transduce this input into distinct cellular signals.

P2X7 nucleotide receptor signaling potentiates the Wnt/β-catenin pathway in cells of the osteoblast lineage

“…Data are means ± SEM (n = 3 independent preparations) alone, or more so by a combination of Wnt3a and BzATP, was associated with enhanced transcriptional activity. These observations are consistent with our previous findings, in which we demonstrated that prolonged but not transient Ca 2+ /NFATc1 signaling in osteoblasts resulted in increased expression of NFAT target genes [23]. Taken together, these findings highlight the positive association between duration of nuclear occupancy by a transcription factor (such as NFAT or β-catenin) and expression of its target genes.…”

“…Additionally, BMP-2 has been shown to stimulate β-catenin transcriptional activity leading to increased expression of Wnt ligands and their receptors in differentiating cultures of calvarial osteoblasts [36]. Previously, we demonstrated that prolonged Ca 2+ /NFATc1 signaling downstream of P2X7 leads to expression of cyclooxygenase-2 (COX-2) [23]. Interestingly, NFATc1 has been shown to function downstream of strontium ranelate in vitro to elicit expression of canonical Wnt3a [37].…”

“…Previous work from our lab has demonstrated that stimulation of P2X7 in osteoblasts activates Ca 2+ signaling and the phosphatidylinositol 3-kinase (PI3K) pathway [17,23]. To investigate the pathways responsible for mediating P2X7-induced inhibitory phosphorylation of GSK3α/β, MC3T3-E1 cells were treated with BzATP (300 μM) in the presence or absence of the PI3K inhibitor LY 294002 (30 μM), the PKC inhibitor Bis I (0.5 μM), or its inactive analog Bis V (0.5 μM) (Fig.…”

Use of Brilliant Blue FCF during vein graft preparation inhibits intimal hyperplasia

“…Compared with other P2X receptor subtypes, significantly higher ATP concentrations are required for P2X 7 receptor activation. 35,36 Although such high ATP concentrations are typically not produced under physiologic conditions, moderate levels of extracellular ATP may play a role in mechanotransduction in a variety of tissues. 36 Moreover, significant release of ATP occurs from stressed, ischemic, hypoxic, or traumatized tissues.…”

Control of bone development by P2X and P2Y receptors expressed in mesenchymal and hematopoietic cells.

“…Intracellular calcium oscillations are known to occur during a number of processes including embryogenesis and the associated signals are implicated in cell differentiation (Tonelli et al, 2012). These autocrine and paracrine functions involve the calcium-dependent transcription factor nuclear factor of activated T cells (NFAT), which is known to be induced by extracellular nucleotides, at least in microglia, human MSCs and osteoblasts (Shiratori et al, 2010; Glaser et al, 2013; Grol et al, 2013). NFAT activation is critical for gene regulation because it is activated during differentiation of thymocytes and may have an integral role in hematopoietic cell growth and differentiation (Adachi et al, 2000).…”