Phospholamban Thiols Play a Central Role in Activation of the Cardiac Muscle Sarcoplasmic Reticulum Calcium Pump by Nitroxyl

Froehlich, Jeffrey P.; Mahaney, James E.; Keceli, Gizem; Pavlos, Christopher M.; Goldstein, Russell; Redwood, Abiona J.; Sumbilla, Carlota; Lee, Dong I.; Tocchetti, Carlo G.; Kass, David A.; Paolocci, Nazareno; Toscano, John P.

doi:10.1021/bi801925p

Cited by 89 publications

(100 citation statements)

References 17 publications

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“…2) show some important features: (i) PLN R9C pentamers have slightly less mobility than the PLN wt , which suggests a change in the protein structural topology (i.e., hydrodynamic radius), and (ii) the presence of heat-resistant dimers. The latter was also observed in the oxidation studies of PLN wt carried out by Froehlich et al (34) (Fig. 2).…”

Section: Resultssupporting

confidence: 71%

“…In vitro assays reported in the previous sections and work from other groups (26,34) (Fig. 4E), with approximately 5% increase in the emission of YFP-PLN R9C .…”

Section: Plnsupporting

confidence: 58%

“…An important finding is the presence of dimers in oxidized PLN R9C . The latter has been previously observed by Froehlich et al upon PLN oxidation by nitroxyl radicals, which promote the formation of disulfide bonds in the transmembrane region, generating noninhibitory oligomers that prevent SERCA regulation (34). Under oxidative conditions, PLN R9C oligomerization is enhanced.…”

Section: Discussionmentioning

confidence: 58%

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Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A

Masterson

Hou

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

103

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The regulatory interaction of phospholamban (PLN) with Ca 2þ -ATPase controls the uptake of calcium into the sarcoplasmic reticulum, modulating heart muscle contractility. A missense mutation in PLN cytoplasmic domain (R9C) triggers dilated cardiomyopathy in humans, leading to premature death. Using a combination of biochemical and biophysical techniques both in vitro and in live cells, we show that the R9C mutation increases the stability of the PLN pentameric assembly via disulfide bridge formation, preventing its binding to Ca 2þ -ATPase as well as phosphorylation by protein kinase A. These effects are enhanced under oxidizing conditions, suggesting that oxidative stress may exacerbate the cardiotoxic effects of the PLN R9C mutant. These results reveal a regulatory role of the PLN pentamer in calcium homeostasis, going beyond the previously hypothesized role of passive storage for active monomers.SERCA | ventricular dilatation | calcium regulation | heart failure | membrane proteins H eart failure (HF) is the leading cause of morbidity and mortality worldwide (1, 2). The most prominent disorder leading to HF is dilated cardiomyopathy (DCM), a disease characterized by left ventricular dilatation and impaired systolic function (1, 2). DCM has both acquired and genetic etiologies (1, 2). Recent genome sequencing has revealed a high incidence of DCM-associated mutations in cytoskeletal, nuclear, as well as sarcomeric proteins (3). A number of mutations have been indentified in calcium handling proteins, which play a central role in the mechanics of heart muscle contractility (3-6).Cardiac muscle contraction (systole) begins when an action potential causes membrane depolarization, activating the sarcolemmal L-type calcium (Ca 2þ ) channels. Ca 2þ flows through the L-type Ca 2þ -channels into the cytosol. This increase in Ca 2þ concentration induces a large-scale release of Ca 2þ into the cytosol from intracellular stores by the sarcoplasmic reticulum (SR) Ca 2þ -release channels (or ryanodine receptors). Ca 2þ then moves toward the contractile apparatus, where it binds the troponin complex and initiates contraction. Muscle relaxation (diastole) occurs when Ca 2þ is sequestered into the SR by the SR Ca 2þ -ATPase (SERCA) (7) a membrane-embedded Ca 2þ pump (8). SERCA is regulated by phospholamban (PLN), which reduces its apparent Ca 2þ affinity (9, 10). PLN's inhibition is reversed by cAMP-dependent protein kinase A (PKA), which phosphorylates PLN at Ser16, enhancing cardiac contractility and reestablishing Ca 2þ flux (11).PLN is a single-pass membrane protein, which comprises three structural domains (12-14), further subdivided into four dynamic domains [cytoplasm: domain Ia (residues 1-16), loop (residues 17-22), domain Ib (residues 23-30); transmembrane: domain II (residues 31-52)] (15) (Fig. S1). In membranes, PLN forms homopentamers arranged in a pinwheel topology that are in equilibrium with monomers (16, 17) that bind SERCA with 1∶1 stoichiometry (6, 18-21). Also, it has been proposed that the PLN monomer-pen...

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Section: Resultssupporting

confidence: 71%

“…In vitro assays reported in the previous sections and work from other groups (26,34) (Fig. 4E), with approximately 5% increase in the emission of YFP-PLN R9C .…”

Section: Plnsupporting

confidence: 58%

Section: Discussionmentioning

confidence: 58%

See 1 more Smart Citation

Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A

Masterson

Hou

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

103

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“…However, recent evidence suggests an alternative, phosphorylation-independent mechanism of SERCA2a-regulated Ca 2þ reuptake. In fact, Froehlich et al (2008) showed that activation of SERCA2a could be achieved by modifying critical thiol residues in the PLN, which through protein conformation change relieves the inhibition of the pump. In particular, the covalent adduction of a nitroso group to a cysteine thiol side chain has recently emerged as a major mechanism by which NO mediates a large number of intracellular processes (Hess et al 2005) and an increase in proteins S-nitrosylation by NO was recently observed, also in eel cardiac tissues (Cerra et al 2009).…”

Section: Discussionmentioning

confidence: 99%

Phospholamban S-nitrosylation modulates Starling response in fish heart

et al. 2009

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The Frank -Starling mechanism is a fundamental property of the vertebrate heart, which allows the myocardium to respond to increased filling pressure with a more vigorous contraction of its lengthened fibres. In mammals, myocardial stretch increases cardiac nitric oxide (NO) release from both vascular endothelium and cardiomyocytes. This facilitates myocardial relaxation and ventricular diastolic distensibility, thus influencing the Frank -Starling mechanism.In the in vitro working heart of the eel Anguilla anguilla, we previously showed that an endogenous NO release affects the Frank -Starling response making the heart more sensitive to preload. Using the same bioassay, we now demonstrate that this effect is confirmed in the presence of the exogenous NO donor S-nitroso-N-acetyl penicillamine, is independent from endocardial endothelium and guanylate cyclase/ cGMP/protein kinase G and cAMP/protein kinase A pathways, involves a PI(3)kinase-mediated activation of endothelial NO synthase and a modulation of the SR-CA 2þ ATPase (SERCA2a) pumps. Furthermore, we show that NO influences cardiac response to preload through S-nitrosylation of phospholamban and consequent activation of SERCA2a. This suggests that in the fish heart NO modulates the Frank -Starling response through a beat-to-beat regulation of calcium reuptake and thus of myocardial relaxation.We propose that this mechanism represents an important evolutionary step for the stretch-induced intrinsic regulation of the vertebrate heart, providing, at the same time, a stimulus for mammalian-oriented studies.

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“…In fact, it has been reported that myocardial production of NO by eNOS and/or neuronal NOS may stimulate the activity of SERCA (7,28,51) by removing the inhibitory effect of phospholamban via nitrosylation of its cysteine residues (7). The role of NO on SERCA activity might explain the severe contracture attributable to L-NNA administration, when also the basal release of NO is impaired if not prevented.…”

Section: Is No Involved In Ap-postc Protection?mentioning

confidence: 99%

Apelin-13 limits infarct size and improves cardiac postischemic mechanical recovery only if given after ischemia

Rastaldo

Cappello

Folino

et al. 2011

American Journal of Physiology-Heart and Circulatory Physiology

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Rastaldo R, Cappello S, Folino A, Berta GN, Sprio AE, Losano G, Samaja M, Pagliaro P. Apelin-13 limits infarct size and improves cardiac postischemic mechanical recovery only if given after ischemia. Am J Physiol Heart Circ Physiol 300: H2308 -H2315, 2011. First published March 4, 2011 doi:10.1152/ajpheart.01177.2010.-We studied whether apelin-13 is cardioprotective against ischemia/reperfusion injury if given as either a pre-or postconditioning mimetic and whether the improved postischemic mechanical recovery induced by apelin-13 depends only on the reduced infarct size or also on a recovery of function of the viable myocardium. We also studied whether nitric oxide (NO) is involved in apelin-induced protection and whether the reported ischemia-induced overexpression of the apelin receptor (APJ) plays a role in cardioprotection. Langendorffperfused rat hearts underwent 30 min of global ischemia and 120 min of reperfusion. Left ventricular pressure was recorded. Infarct size and lactate dehydrogenase release were determined to evaluate the severity of myocardial injury. Apelin-13 was infused at 0.5 M concentration for 20 min either before ischemia or in early reperfusion, without and with NO synthase inhibition by N G -nitro-L-arginine (L-NNA). In additional experiments, before ischemia also 1 M apelin-13 was tested. APJ protein level was measured before and after ischemia. Whereas before ischemia apelin-13 (0.5 and 1.0 M) was ineffective, after ischemia it reduced infarct size from 54 Ϯ 2% to 26 Ϯ 4% of risk area (P Ͻ 0.001) and limited the postischemic myocardial contracture (P Ͻ 0.001). L-NNA alone increased postischemic myocardial contracture. This increase was attenuated by apelin-13, which, however, was unable to reduce infarct size. Ischemia increased APJ protein level after 15-min perfusion, i.e., after most of reperfusion injury has occurred. Apelin-13 protects the heart only if given after ischemia. In this protection NO plays an important role. Apelin-13 efficiency as postconditioning mimetic cannot be explained by the increased APJ level. APJ receptor; ischemia/reperfusion; myocardial protection; preconditioning APELIN IS THE ENDOGENOUS LIGAND for the G protein-coupled APJ receptor. Human, mouse, rat, and cow apelin genes encode a 77-amino acid preprotein with the active sequence in the COOH-terminal region (19,43). Various fragments of apelin have been isolated and classified according to the number of amino acids. Among fragments, apelin-13 and -36 are the most frequently studied, and apelin-13 is considered the most active of them (40). Apelin receptors APJ have been found to be similar to angiotensin II receptor type 1 (25,30,43), but angiotensin II does not bind to APJ receptors (14, 30).Apelin mRNA is expressed in several organs and tissues. In the cardiovascular system, apelin and APJ receptors occur in vascular smooth muscle, endothelial cells, and cardiomyocytes (21,22). Apelin exerts a positive inotropic effect (2, 42) and a nitric oxide (NO)-driven vasodilator activity (14, 15). It protect...

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Phospholamban Thiols Play a Central Role in Activation of the Cardiac Muscle Sarcoplasmic Reticulum Calcium Pump by Nitroxyl

Cited by 89 publications

References 17 publications

Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A

Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A

Phospholamban S-nitrosylation modulates Starling response in fish heart

Apelin-13 limits infarct size and improves cardiac postischemic mechanical recovery only if given after ischemia

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Phospholamban Thiols Play a Central Role in Activation of the Cardiac Muscle Sarcoplasmic Reticulum Calcium Pump by Nitroxyl

Cited by 89 publications

References 17 publications

Lethal Arg9Cys phospholamban mutation hinders Ca 2+ -ATPase regulation and phosphorylation by protein kinase A

Lethal Arg9Cys phospholamban mutation hinders Ca 2+ -ATPase regulation and phosphorylation by protein kinase A

Phospholamban S-nitrosylation modulates Starling response in fish heart

Apelin-13 limits infarct size and improves cardiac postischemic mechanical recovery only if given after ischemia

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Product

Resources

About

Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A

Lethal Arg9Cys phospholamban mutation hinders Ca ²⁺ -ATPase regulation and phosphorylation by protein kinase A