The inhibitory glutathione transferase M2-2 binding site is located in divergent region 3 of the cardiac ryanodine receptor

Liú, Dan; Hewawasam, Ruwani Punyakanthi; Karunasekara, Yamuna; Dulhunty, Angela F.; Board, Philip G.

doi:10.1016/j.bcp.2012.02.020

Cited by 10 publications

(8 citation statements)

References 34 publications

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“…We attempted to identify the brain RyR isoform using the peptide GSTM2C H5-8, a C-terminal fragment of the musclespecific glutathione transferase (GSTM2). GSTM2C H5-8 specifically inhibits RyR2, but not RyR1 (Hewawasam et al, 2010), by binding to divergent region three of RyR2 (which has little sequence homology with RyR1 or RyR3) (Liu et al, 2012a). The brain RyRs failed to respond to 8 µM GSTM2C added after 2 μM H101Q CLIC2, suggesting that the channels were not RyR2.…”

Section: Sub-state Opening Induced By Wt Clic2 and H101q Clic2mentioning

confidence: 98%

Association of FK506 binding proteins with RyR channels – effect of CLIC2 binding on sub-conductance opening and FKBP binding

Richardson

Steele

Gallant

et al. 2017

Journal of Cell Science

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Ryanodine receptor (RyR) Ca channels are central to striated muscle function and influence signalling in neurons and other cell types. Beneficially low RyR activity and maximum conductance opening may be stabilised when RyRs bind to FK506 binding proteins (FKBPs) and destabilised by FKBP dissociation, with submaximal opening during RyR hyperactivity associated with myopathies and neurological disorders. However, the correlation with submaximal opening is debated and quantitative evidence is lacking. Here, we have measured altered FKBP binding to RyRs and submaximal activity with addition of wild-type (WT) CLIC2, an inhibitory RyR ligand, or its H101Q mutant that hyperactivates RyRs, which probably causes cardiac and intellectual abnormalities. The proportion of sub-conductance opening increases with WT and H101Q CLIC2 and is correlated with reduced FKBP-RyR association. The sub-conductance opening reduces RyR currents in the presence of WT CLIC2. In contrast, sub-conductance openings contribute to excess RyR 'leak' with H101Q CLIC2. There are significant FKBP and RyR isoform-specific actions of CLIC2, rapamycin and FK506 on FKBP-RyR association. The results show that FKBPs do influence RyR gating and would contribute to excess Ca release in this CLIC2 RyR channelopathy.

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Section: Sub-state Opening Induced By Wt Clic2 and H101q Clic2mentioning

confidence: 98%

Association of FK506 binding proteins with RyR channels – effect of CLIC2 binding on sub-conductance opening and FKBP binding

Richardson

Steele

Gallant

et al. 2017

Journal of Cell Science

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“…Such inward currents can be generated by surface membrane HCN2 and HCN4 channels [ 27 ] and/or follow activation of the NCX by increases in cytoplasmic [Ca 2+ ] following release from IP3 and/or ryanodine sensitive stores [ 28 , 31 ]. The effect of GSTM2C on the number of spontaneously active cells and on the frequency of spontaneous contractility most likely depends on an effect of the drug on Ca 2+ leak through RyR2, since GSTM2C is a specific inhibitor of RyR2 channels [ 11 , 12 , 16 ]. Indeed a BLAST alignment indicates that the specific GSTM2C binding sequence in the RyR2 D3 region [ 16 ] is not present in rat or human HCN2.…”

Section: Discussionmentioning

confidence: 99%

“…The effect of GSTM2C on the number of spontaneously active cells and on the frequency of spontaneous contractility most likely depends on an effect of the drug on Ca 2+ leak through RyR2, since GSTM2C is a specific inhibitor of RyR2 channels [ 11 , 12 , 16 ]. Indeed a BLAST alignment indicates that the specific GSTM2C binding sequence in the RyR2 D3 region [ 16 ] is not present in rat or human HCN2. There is a region of alignment in HCN4, but only one identical residue was found to lie within the GSTM2C-binding region.…”

Section: Discussionmentioning

confidence: 99%

“…We did not specifically examine the effects of the constructs on other factors such as SERCA or NCX activity. However the fact that the specific inhibition of the cardiac RyR2 depends on GSTM2C binding to a cardiac RyR2-specific sequence in the divergent region 3 of RyR2 [ 16 ] suggests that the compound would be unlikely to interact with other proteins that do not contain this specific sequence.…”

Section: Discussionmentioning

confidence: 99%

“…Specific mutations in helix 6 ( Fig 1 ) showed that two residues, F157A and Y160A prevent the C-terminal half of the protein from exerting its normal inhibitory action on RyR2 channels [ 11 ]. Furthermore, we find that the selective inhibition of RyR2 by GSTM2C is due to its interaction with amino acids within the divergent region 3 (D3 region) of cardiac ryanodine receptor [ 16 ]. We have also shown that GSTM2C can enter cells and carry cargo [ 17 ] and is taken up by adult mouse cardiomyocytes, altering Ca 2+ signaling and contraction in a manner consistent with a direct action on RyR2 [ 18 ].…”

Section: Introductionmentioning

confidence: 99%

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The GSTM2 C-Terminal Domain Depresses Contractility and Ca2+ Transients in Neonatal Rat Ventricular Cardiomyocytes

et al. 2016

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The cardiac ryanodine receptor (RyR2) is an intracellular ion channel that regulates Ca2+ release from the sarcoplasmic reticulum (SR) during excitation–contraction coupling in the heart. The glutathione transferases (GSTs) are a family of phase II detoxification enzymes with additional functions including the selective inhibition of RyR2, with therapeutic implications. The C-terminal half of GSTM2 (GSTM2C) is essential for RyR2 inhibition, and mutations F157A and Y160A within GSTM2C prevent the inhibitory action. Our objective in this investigation was to determine whether GSTM2C can enter cultured rat neonatal ventricular cardiomyocytes and influence contractility. We show that oregon green-tagged GSTM2C (at 1 μM) is internalized into the myocytes and it reduces spontaneous contraction frequency and myocyte shortening. Field stimulation of myocytes evoked contraction in the same percentage of myocytes treated either with media alone or media plus 15 μM GSTM2C. Myocyte shortening during contraction was significantly reduced by exposure to 15 μM GSTM2C, but not 5 and 10 μM GSTM2C and was unaffected by exposure to 15 μM of the mutants Y160A or F157A. The amplitude of the Ca2+ transient in the 15 μM GSTM2C - treated myocytes was significantly decreased, the rise time was significantly longer and the decay time was significantly shorter than in control myocytes. The Ca2+ transient was not altered by exposure to Y160A or F157A. The results are consistent with GSTM2C entering the myocytes and inhibiting RyR2, in a manner that indicates a possible therapeutic potential for treatment of arrhythmia in the neonatal heart.

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Physiology and Pharmacology of Ryanodine Receptor Calcium Release Channels

Dulhunty

Board

Beard

2017

Advances in Pharmacology

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The inhibitory glutathione transferase M2-2 binding site is located in divergent region 3 of the cardiac ryanodine receptor

Cited by 10 publications

References 34 publications

Association of FK506 binding proteins with RyR channels – effect of CLIC2 binding on sub-conductance opening and FKBP binding

Association of FK506 binding proteins with RyR channels – effect of CLIC2 binding on sub-conductance opening and FKBP binding

The GSTM2 C-Terminal Domain Depresses Contractility and Ca2+ Transients in Neonatal Rat Ventricular Cardiomyocytes

Physiology and Pharmacology of Ryanodine Receptor Calcium Release Channels

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