Abstract:Pannexins constitute a family of proteins exhibiting predominantly hemichannel activity. Pannexin channels have been suggested to participate in a wide spectrum of biological functions such as propagation of calcium waves, release of IL-1, and responses to ischemic conditions. At present, the molecular mechanisms regulating pannexin hemichannel activity are essentially unknown. Because cysteines have been shown to constitute key elements in regulating hemichannel properties of the connexin-type we performed s… Show more
“…For example, we found that a synthetic peptide containing just this sequence was not sufficient to block current from truncated hPANX1 channels, whereas larger C-terminal proteins were effective. In addition, a more proximal C-terminal residue (Cys-347) that regulates basal channel activity has been identified (16). It is also interesting that hPANX1 inhibition associated with mutations in this restricted region or obtained by cell-free reconstitution of C-terminal proteins with truncated channels was most pronounced at hyperpolarized potentials.…”
Section: Discussionmentioning
confidence: 90%
“…mSpo20 (14) and PLC␦-PH (15)). Likewise, hPANX1⌬371 currents were unaffected by a Cterminal construct that includes a point mutation (hPANX1(Ct)C347S) known to yield constitutive currents in full-length hPANX1 channels (16). Finally, a C-terminal construct truncated at Glu-391 (hPANX1(Ct)⌬391) provided a block of current that was comparable with that seen with the full-length C terminus, consistent with our observation that residues downstream of Glu-391 are not required for C-terminal inhibition of hPANX1 (cf.…”
Background: Pannexin 1 is activated by caspase cleavage of its C-terminal tail during apoptosis. Results: Cleavage removes a critical adjacent region to activate membrane-associated PANX1; activation requires dissociation of the C terminus from the pore. Conclusion: An intrinsic inhibitory interaction between the C terminus and the pore constrains PANX1 activity. Significance: PANX1 activation is caused by disruption of C-terminal-mediated inhibition.
“…For example, we found that a synthetic peptide containing just this sequence was not sufficient to block current from truncated hPANX1 channels, whereas larger C-terminal proteins were effective. In addition, a more proximal C-terminal residue (Cys-347) that regulates basal channel activity has been identified (16). It is also interesting that hPANX1 inhibition associated with mutations in this restricted region or obtained by cell-free reconstitution of C-terminal proteins with truncated channels was most pronounced at hyperpolarized potentials.…”
Section: Discussionmentioning
confidence: 90%
“…mSpo20 (14) and PLC␦-PH (15)). Likewise, hPANX1⌬371 currents were unaffected by a Cterminal construct that includes a point mutation (hPANX1(Ct)C347S) known to yield constitutive currents in full-length hPANX1 channels (16). Finally, a C-terminal construct truncated at Glu-391 (hPANX1(Ct)⌬391) provided a block of current that was comparable with that seen with the full-length C terminus, consistent with our observation that residues downstream of Glu-391 are not required for C-terminal inhibition of hPANX1 (cf.…”
Background: Pannexin 1 is activated by caspase cleavage of its C-terminal tail during apoptosis. Results: Cleavage removes a critical adjacent region to activate membrane-associated PANX1; activation requires dissociation of the C terminus from the pore. Conclusion: An intrinsic inhibitory interaction between the C terminus and the pore constrains PANX1 activity. Significance: PANX1 activation is caused by disruption of C-terminal-mediated inhibition.
“…31,32 Moreover, there is a loss of channel function when any of the four extracellular cysteines of Panx1 are mutated. 33 The possibility of intrinsic control of pannexin channel activity by the cysteines hints at the potential regulation of the channel by such post-translational modifications as glutathionation or S-nitrosylation.…”
Section: Post-translational Modification Of Pannexinsmentioning
The mammalian pannexin family of channel-forming proteins consisting of Panx1, Panx2, and Panx3 has received considerable attention in the last 10 years given their newly discovered physiological roles in development and disease. Pannexins exhibit diverse subcellular profiles indicating that they may serve distinct roles in cells and tissues of different origin. This complexity in cellular residencies may be rooted in the fact that pannexin genes consist of multiple exons that have led to the identification of several splice variants. Additionally, post-translational modifications, especially N-glycosylation, appear to be important in regulating trafficking and intermixing of pannexin family members increasing the diversity of assembled channels. These long-lived membrane proteins are typically trafficked through the classical secretory pathway before reaching the plasma membrane although Panx2 tends to often be retained in intracellular compartments. Trafficking, stability, and function of pannexins likely enlist the services of an interactome that continues to expand. The research field has been amazed by the fact that Panx1 null mice are generally healthy with distinct phenotypes only being revealed when mutant mice encounter additional stress or have comorbidities. The emerging field of pannexin biology has also begun to explore the relationships and potential cross-talk between pannexin channels and connexin hemichannels. It is imperative to dissect the different constituents of the channels and the molecules that pass through these distinct channel types. Finally, as witnessed in connexin biology throughout the 90s, the field awaits to see if germline mutations in the genes that encode pannexins also cause disease.
“…Interestingly, the two sites that we identified as critical for this inhibitory modification --C40 and C346 --were previously reported to enhance activity of Panx1 channels in mutagenesis studies [259,260]. In that other work, serine substitution at either C40 [260] or C346 [259] produced "leaky" or constitutively active channels. Together with our results, these observations suggest that C40 and C346 may be localized to regions that are important for dynamic up-and downregulation of Panx1 channel activity.…”
Section: Discussionmentioning
confidence: 81%
“…Also, a previous study identified a loss in the Gly2 species in functional, plasma membrane-localized Panx1C346S [259].…”
The nucleotide adenosine 5'-triphosphate (ATP) has classically been considered the cell's primary energy currency; however, a novel role for ATP as an extracellular autocrine/paracrine signaling molecule has evolved over the past century. Purinergic signaling is now known to regulate a plethora of physiological and pathophysiological processes in almost every organ system. In the vasculature, ATP and its metabolites elicit dual control over blood vessel tone and tissue perfusion, and purinergic signaling events have been implicated in vascular pathologies including atherosclerosis and inflammation.While the importance of extracellular ATP in the vascular system is well recognized, the mechanism(s) mediating the regulated release of the purine from vascular cells are less well understood and are a key target of current investigation. One such ATP-liberation mechanism has been ascribed to the recently identified pannexin (Panx) channels, namely Panx1. Initially, we characterized the expression and localization profiles of Panx isoforms across the systemic vasculature, identifying predominant representation by the Panx1 isoform in both smooth muscle (SMC) and endothelial cells (EC) comprising the blood vessel wall, with more heterogeneous expression profiles observed in specialized vascular systems including the heart, lung and kidney. In particular, the Panx1 isoform is highly expressed in ECs regardless of blood vessel type or size, while Panx1 expression is limited to SMCs of small arteries and arterioles. Panx1 forms hexameric channels in the plasma membrane of ECs, functioning to release ATP in response to a number of stimuli. However, prolonged Panx1 channel activity is extremely detrimental to cell viability. Here we report a novel negative regulatory mechanism that may govern the activity of Panx1 channels in ECs, by which the bioactive gas nitric oxide (NO) covalently modifies two cysteine (Cys) residues in the channel by a process termed S- reinforce the dual effect of purines on peripheral resistance and blood pressure.
Purinergic Control of Vascular InflammationWhile the foundation for purinergic control of vascular functions was initially characterized extensively in the context of vascular reactivity and overall blood flow and pressure regulation, it has become increasingly clear that extracellular ATP also plays important regulatory roles in the vascular inflammatory response. Vascular inflammation can be characterized as acute (physiological) or chronic (pathological) in nature.The acute vascular inflammatory response is an innate process of inflammatory cell homing to infected or damaged tissues resulting from integrated cross-talk between circulating leukocytes and the vascular endothelium, primarily at the level of postcapillary venules [61]. This process has been highly studied and is now known to occur in a step-wise manner beginning with local recruitment, rolling (fast and slow), adhesion and final extravasation into the surrounding tissue (reviewed in [62...
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