{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T22:58:07Z","timestamp":1790636287550,"version":"4.1.0"},"reference-count":45,"publisher":"MDPI AG","issue":"19","license":[{"start":{"date-parts":[[2020,9,23]],"date-time":"2020-09-23T00:00:00Z","timestamp":1600819200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000291","name":"Kidney Research UK","doi-asserted-by":"publisher","award":["RP30\/July 2014"],"award-info":[{"award-number":["RP30\/July 2014"]}],"id":[{"id":"10.13039\/501100000291","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Hyperphosphataemia increases cardiovascular mortality in patients with kidney disease. Direct effects of high inorganic phosphate (Pi) concentrations have previously been demonstrated on endothelial cells (ECs), including generation of procoagulant endothelial microvesicles (MVs). However, no mechanism directly sensing elevated intracellular Pi has ever been described in mammalian cells. Here, we investigated the hypothesis that direct inhibition by Pi of the phosphoprotein phosphatase PP2A fulfils this sensing role in ECs, culminating in cytoskeleton disruption and MV generation. ECs were treated with control (1 mM [Pi]) vs. high (2.5 mM [Pi]), a condition that drives actin stress fibre depletion and MV generation demonstrated by confocal microscopy of F-actin and NanoSight Nanoparticle tracking, respectively. Immuno-blotting demonstrated that high Pi increased p-Src, p-PP2A-C and p-DAPK-1 and decreased p-TPM-3. Pi at 100 \u03bcM directly inhibited PP2A catalytic activity. Inhibition of PP2A enhanced inhibitory phosphorylation of DAPK-1, leading to hypophosphorylation of Tropomyosin-3 at S284 and MV generation. p-Src is known to perform inhibitory phosphorylation on DAPK-1 but also on PP2A-C. However, PP2A-C can itself dephosphorylate (and therefore inhibit) p-Src. The direct inhibition of PP2A-C by Pi is, therefore, amplified by the feedback loop between PP2A-C and p-Src, resulting in further PP2A-C inhibition. These data demonstrated that PP2A\/Src acts as a potent sensor and amplifier of Pi signals which can further signal through DAPK-1\/Tropomyosin-3 to generate cytoskeleton disruption and generation of potentially pathological MVs.<\/jats:p>","DOI":"10.3390\/ijms21196993","type":"journal-article","created":{"date-parts":[[2020,9,23]],"date-time":"2020-09-23T09:28:08Z","timestamp":1600853288000},"page":"6993","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Inorganic Phosphate (Pi) Signaling in Endothelial Cells: A Molecular Basis for Generation of Endothelial Microvesicles in Uraemic Cardiovascular Disease"],"prefix":"10.3390","volume":"21","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-6906-2563","authenticated-orcid":false,"given":"Nima","family":"Abbasian","sequence":"first","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alan","family":"Bevington","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1176-7592","authenticated-orcid":false,"given":"James O.","family":"Burton","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"},{"name":"Department of Nephrology, Leicester General Hospital, Leicester LE5 4PW, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karl E.","family":"Herbert","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alison H.","family":"Goodall","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nigel J.","family":"Brunskill","sequence":"additional","affiliation":[{"name":"Department of Cardiovascular Sciences, University of Leicester, and Leicester NIHR Cardiovascular Biomedical Research Unit, Leicester LE3 9QP, UK"},{"name":"Department of Nephrology, Leicester General Hospital, Leicester LE5 4PW, UK"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2020,9,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1042\/CS20100377","article-title":"Phosphate toxicity: New insights into an old problem","volume":"120","author":"Razzaque","year":"2011","journal-title":"Clin. 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