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Regulation of ryanodine receptor by nitric oxide

Cheong, Eunji (2004) Regulation of ryanodine receptor by nitric oxide. Doctoral Dissertation, University of Pittsburgh.

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    Abstract

    The control of redox state of free thiols on ryanodine receptor (RyR) has been implicated as an important mechanism to regulate RyR channel activity and tune its responses to the physiological modulators. Both the skeletal and cardiac RyRs have been shown to be activated by S-nitrosylation of free thiols on them by a chemical process analogous to the oxidation of ¡°critical¡± or ¡°hyperreactive¡± thiols on RyR proteins. Inositol 1,4,5-triphophate receptors (IP3Rs) that control Ca2+ release from internal stores in non-excitable cells were found to be activated by oxidation, which emphasizes the redox reaction as a common mechanism to regulate intracellular Ca2+ channels. Therefore, the study on nitric oxide-mediated regulation of these ion channels will be important to understand the regulation of Ca2+ homeostasis in all cells including excitable and non-excitable cells since it regulates internal Ca2+ stores via RyR and /or IP3 receptors.The aims were to investigate the chemical reaction underlying the thiol-oxidation and activation of ryanodine receptors (RyRs) by various types of NO donors namely authentic NO¡¤, S-nitrosothiols and other NO¡¤ species such as nitroxyl anions (i.e. HNO). The different actions of these various NO¡¤ species were used to better evaluate the physiological significance of RyR activation by biologically relevant forms of NO, to investigate the role of oxygen on these chemical reactions and to identify the critical cysteine residues involved in redox mediated regulation of RyRs. The main findings are that RyRs are direct targets of S-nitrosothiols which trans-nitrosate hyper-reactive thiols and activate RyRs at biologically relevant concentration. In contrast, NO¡¤ gas cannot modify RyRs at biological circumstances found in cells. HNO is considerably more potent activator of RyR1 than NO¡¤, activates RyRs at nM concentrations. The study with truncated RyR1 indicated that all of transmembrane domains are located close to the C-terminus of the protein and the ¡®critical¡¯ regulatory thiols are part of conserved cysteines residing in it.Further studies will be required to elucidate the interplay of oxidants and reductants found in the cytosolic milieu of all cells and how these activators and inhibitors act to regulate the opening and closure of Ca2+ release channels.


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    Item Type: University of Pittsburgh ETD
    ETD Committee:
    ETD Committee TypeCommittee MemberEmail
    Committee ChairSalama, Guygsalama@pitt.edu
    Committee MemberFarkas, DanielDaniel.Farkas@cshs.org
    Committee MemberBorovetz, Harveyborovetzhs@upmc.edu
    Committee MemberWalker, Williamwalkerw@pitt.edu
    Title: Regulation of ryanodine receptor by nitric oxide
    Status: Unpublished
    Abstract: The control of redox state of free thiols on ryanodine receptor (RyR) has been implicated as an important mechanism to regulate RyR channel activity and tune its responses to the physiological modulators. Both the skeletal and cardiac RyRs have been shown to be activated by S-nitrosylation of free thiols on them by a chemical process analogous to the oxidation of ¡°critical¡± or ¡°hyperreactive¡± thiols on RyR proteins. Inositol 1,4,5-triphophate receptors (IP3Rs) that control Ca2+ release from internal stores in non-excitable cells were found to be activated by oxidation, which emphasizes the redox reaction as a common mechanism to regulate intracellular Ca2+ channels. Therefore, the study on nitric oxide-mediated regulation of these ion channels will be important to understand the regulation of Ca2+ homeostasis in all cells including excitable and non-excitable cells since it regulates internal Ca2+ stores via RyR and /or IP3 receptors.The aims were to investigate the chemical reaction underlying the thiol-oxidation and activation of ryanodine receptors (RyRs) by various types of NO donors namely authentic NO¡¤, S-nitrosothiols and other NO¡¤ species such as nitroxyl anions (i.e. HNO). The different actions of these various NO¡¤ species were used to better evaluate the physiological significance of RyR activation by biologically relevant forms of NO, to investigate the role of oxygen on these chemical reactions and to identify the critical cysteine residues involved in redox mediated regulation of RyRs. The main findings are that RyRs are direct targets of S-nitrosothiols which trans-nitrosate hyper-reactive thiols and activate RyRs at biologically relevant concentration. In contrast, NO¡¤ gas cannot modify RyRs at biological circumstances found in cells. HNO is considerably more potent activator of RyR1 than NO¡¤, activates RyRs at nM concentrations. The study with truncated RyR1 indicated that all of transmembrane domains are located close to the C-terminus of the protein and the ¡®critical¡¯ regulatory thiols are part of conserved cysteines residing in it.Further studies will be required to elucidate the interplay of oxidants and reductants found in the cytosolic milieu of all cells and how these activators and inhibitors act to regulate the opening and closure of Ca2+ release channels.
    Date: 02 February 2004
    Date Type: Completion
    Defense Date: 13 October 2003
    Approval Date: 02 February 2004
    Submission Date: 12 November 2003
    Access Restriction: No restriction; Release the ETD for access worldwide immediately.
    Patent pending: No
    Institution: University of Pittsburgh
    Thesis Type: Doctoral Dissertation
    Refereed: Yes
    Degree: PhD - Doctor of Philosophy
    URN: etd-11122003-081026
    Uncontrolled Keywords: Ca2+ release channel; nitric oxide; redox regulation; ryanodine receptor
    Schools and Programs: Swanson School of Engineering > Bioengineering
    Date Deposited: 10 Nov 2011 15:04
    Last Modified: 11 May 2012 11:34
    Other ID: http://etd.library.pitt.edu/ETD/available/etd-11122003-081026/, etd-11122003-081026

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