The RhoA regulators Myo9b and GEF-H1 are targets of cyclic nucleotide-dependent kinases in platelets

DC FieldValueLanguage
dc.contributor.authorComer, Shane-
dc.contributor.authorNagy, Zoltan-
dc.contributor.authorBolado, Alfonso-
dc.contributor.authorSmolenski, Albert P.-
dc.contributor.authoret al.-
dc.date.accessioned2021-11-10T10:38:54Z-
dc.date.available2021-11-10T10:38:54Z-
dc.date.copyright2020 International Society on Thrombosis and Haemostasisen_US
dc.date.issued2020-11-
dc.identifier.citationJournal of Thrombosis and Haemostasisen_US
dc.identifier.issn1538-7933-
dc.identifier.urihttp://hdl.handle.net/10197/12616-
dc.description.abstractBackground: Circulating platelets are maintained in an inactive state by the endothelial lining of the vasculature. Endothelium-derived prostacyclin and nitric oxide stimulate cAMP- and cGMP-dependent kinases, PKA and PKG, to inhibit platelets. PKA and PKG effects include the inhibition of the GTPase RhoA, which has been suggested to involve the direct phosphorylation of RhoA on serine 188. Objectives: We wanted to confirm RhoA S188 phosphorylation by cyclic nucleotide-dependent kinases and to identify possible alternative mechanisms of RhoA regulation in platelets. Methods: Phosphoproteomics data of human platelets were used to identify candidate PKA and PKG substrates. Phosphorylation of individual proteins was studied by Western blotting and Phos-tag gel electrophoresis in human platelets and transfected HEK293T cells. Pull-down assays were performed to analyze protein interaction and function. Results: Our data indicate that RhoA is not phosphorylated by PKA in platelets. Instead, we provide evidence that cyclic nucleotide effects are mediated through the phosphorylation of the RhoA-specific GTPase-activating protein Myo9b and the guanine nucleotide exchange factor GEF-H1. We identify Myo9b S1354 and guanine nucleotide exchange factor-H1 (GEF-H1) S886 as PKA and PKG phosphorylation sites. Myo9b S1354 phosphorylation enhances its GTPase activating protein function leading to reduced RhoA-GTP levels. GEF-H1 S886 phosphorylation stimulates binding of 14-3-3β and has been shown to inhibit GEF function by facilitating binding of GEF-H1 to microtubules. Microtubule disruption increases RhoA-GTP levels confirming the importance of GEF-H1 in platelets. Conclusion: Phosphorylation of RhoA regulatory proteins Myo9b and GEF-H1, but not RhoA itself, is involved in cyclic nucleotide-mediated control of RhoA in human platelets.en_US
dc.description.sponsorshipUniversity College Dublinen_US
dc.format.mediumPrint-Electronic-
dc.language.isoenen_US
dc.publisherWileyen_US
dc.rightsThis is the peer reviewed version of the following article: Comer, S, Nagy, Z, Bolado, A, et al. The RhoA regulators Myo9b and GEF‐H1 are targets of cyclic nucleotide‐dependent kinases in platelets. J Thromb Haemost. 2020; 00: 1– 11, which has been published in final form at https://doi.org/10.1111/jth.15028. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.en_US
dc.subjectPeripheral vascular diseaseen_US
dc.subject14-3-3 proteinsen_US
dc.subjectCyclic AMP-dependent protein kinasesen_US
dc.subjectCyclic GMP-dependent protein kinasesen_US
dc.subjectGTPase-activating proteinsen_US
dc.subjectGuanine nucleotide exchange factorsen_US
dc.subjectPhosphorylationen_US
dc.subjectGTPase-activating proteinen_US
dc.subjectShape changeen_US
dc.subjectMyosin IXBen_US
dc.subjectSignaling pathwayen_US
dc.subjectPhosphorylationen_US
dc.subjectPhosphataseen_US
dc.subjectRevealsen_US
dc.subjectBindingen_US
dc.subjectContractionen_US
dc.subjectThrombinen_US
dc.titleThe RhoA regulators Myo9b and GEF-H1 are targets of cyclic nucleotide-dependent kinases in plateletsen_US
dc.title.alternativeRhoA regulation by Myo9b and GEF-H1 in plateletsen_US
dc.typeJournal Articleen_US
dc.internal.authorcontactotheralbert.smolenski@ucd.ieen_US
dc.statusPeer revieweden_US
dc.identifier.volume18en_US
dc.identifier.issue11en_US
dc.identifier.startpage3002en_US
dc.identifier.endpage3012en_US
dc.identifier.doi10.1111/jth.15028-
dc.neeo.contributorComer|Shane|aut|-
dc.neeo.contributorNagy|Zoltan|aut|-
dc.neeo.contributorBolado|Alfonso|aut|-
dc.neeo.contributorSmolenski|Albert P.|aut|-
dc.neeo.contributoret al.||aut|-
dc.date.embargo2021-07-21en_US
dc.description.othersponsorshipRussian Foundation For Basic Researchen_US
dc.description.othersponsorshipDeutsche Forschungsgemeinschaften_US
dc.date.updated2020-09-17T15:28:26Z-
dc.identifier.grantid17‐00‐00141-
dc.identifier.grantid17‐00‐00139-
dc.identifier.grantidJU 2735/2‐1-
dc.identifier.grantidZA 639/4‐1-
dc.rights.licensehttps://creativecommons.org/licenses/by-nc-nd/3.0/ie/en_US
item.fulltextWith Fulltext-
item.grantfulltextopen-
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