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SURA2 targeting for cardioprotection?
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SURA2 targeting for cardioprotection? Aleksandar Jovanovic´ and Sofija Jovanovic´ SUR2A is an ATP-binding protein known to serve as a regulatory subunit of metabolic-sensing, cardioprotective sarcolemmal ATP-sensitive K+ (KATP) channels. It has been recently found that a moderate increase in expression of SUR2A protects the heart against different types of metabolic stresses, including ischaemia/reperfusion and hypoxia. Although the sarcolemmal KATP channel is a multiprotein complex composed of many proteins in vivo, it seems that an increase in SUR2A levels is sufficient to increase the number of sarcolemmal KATP channels. This effect of SUR2A could be due to SUR2A being the rate-limiting factor in generating fully composed sarcolemmal KATP channels. An increased number of sarcolemmal KATP channels seems to protect the heart by regulating action membrane potential, inhibiting Ca2+ influx and preventing Ca2+ overload, although an additional yet to be recognised mechanism independent of KATP channels activity cannot be excluded. Address Division of Medical Sciences, Ninewells Hospital & Medical School, University of Dundee, Dundee DD1 9SY, UK Corresponding author: Jovanovic´, Aleksandar ([email protected]) and
Current Opinion in Pharmacology 2009, 9:189–193 This review comes from a themed issue on Cardiovascular and renal Edited by Michael Curtis Available online 10th December 2008 1471-4892/$ – see front matter # 2008 Elsevier Ltd. All rights reserved. DOI 10.1016/j.coph.2008.11.003
Introduction In 1986, Murry et al. discovered that brief periods of blood vessel occlusion and reperfusion administered before a sustained ischaemic episode lead to a reduction in myocardial infarct size [1]. This and subsequent studies have indicated that brief ischaemia or hypoxia are likely to switch on signalling pathways that prevent or delay lethal outcomes of stressed cardiac cells and this phenomenon is known as cardioprotection [2]. ATP-sensitive K+ (KATP) channels are present in different tissues and are combinations of an inwardly rectifying K+ channel, Kir6.1 or/and Kir6.2 and a regulatory subunit, the sulfonylurea receptor SUR1, SUR2A or SUR2B [3]. Physical association of the Kir6.2 and SUR2A isoforms www.sciencedirect.com
generates cardiac KATP channels that are expressed in high density at the sarcolemma [4]. More recent studies have suggested that the sarcolemmal KATP channel protein complex may be composed of more proteins than just Kir6.2 and SUR2A [5–10]. Since intracellular ATP regulates the activity of KATP channels, it is consensus view that this ion conductance couples metabolic state of the cell with the membrane excitability [4]. In numerous studies, it has been demonstrated that KATP channel openers, drugs that promote opening of KATP channels, decrease infarct size, mimic ischaemic preconditioning and improve functional and energetic recovery of cardiac muscle following ischaemic and hypoxic insults [11]. More recently, it has been shown that targeting specific subunits of sarcolemmal KATP channels protein complex might have significant consequences on myocardial resistance to different types of metabolic stresses, including ischaemia/reperfusion and hypoxia [12,13,14,15]. One of the subunits that can be targeted to manipulate with the cardiac resistance to metabolic stress is SUR2A. Molecular biology, biochemistry and physiology of SUR2A have been reviewed recently in great details [16] and readers can use this review to learn more about structural/physiological/functional aspects of SUR2A. Here, we will provide a brief outline of a relationship between SUR2A and cardioprotection and highlight how targeting SUR2A alone might be a potential strategy to protect the heart against metabolic stress.
Evidence that changes in SUR2A expression regulates myocardial resistance to ischaemia/ reperfusion and hypoxia The first evidence demonstrating that SUR2A level might be particularly important in myocardial resistance to ischaemia was presented in a study addressing genderspecific difference in sarcolemmal KATP channels. It has been shown that females have more assembled functional sarcolemmal KATP channels than males and this was associated with cardiac cells from females being more resistant to ischaemia reperfusion [17]. The most intriguing finding was that no gender-specific difference was observed in mRNA levels of any other KATP channel subunits apart from SUR2A [17]. Further studies have shown that any condition that increases the number of sarcolemmal KATP channels (young age, female gender, oestrogen treatment and chronic hypoxia) is associated with a sole increase in SUR2A mRNA and with increased resistance to ischaemia/reperfusion [17–20]. This has suggested that upregulation of SUR2A is sufficient to Current Opinion in Pharmacology 2009, 9:189–193
190 Cardiovascular and renal
Figure 1
Increase in SUR2A expression confers myocardial resistance against ischaemia/reperfusion. Typical photographs of myocardial slices from wild type and transgenic mice with increased SUR2A expression after exposure to 30 min-long ischaemia followed by 30 min-long reperfusion. Infarcted areas are pale/gray (indicated also with the blue arrows) whereas viable myocardium is dark/red. The average size of myocardial infarction was 21.2 5.1% (n = 17) in wild type and 5.3 1.2% (n = 6) in transgenic mice. This difference was statistically significant (P = 0.02). Modified from [13] with permission.
increase the resistance to ischaemia/reperfusion. This kind of conclusion was particularly supported by a study showing that chronic mild hypoxia upregulates SUR2A without affecting the expression of any other gene [20]. In a latter study, transgenic animals overexpressing SUR2A without changes of expression of other KATP channel forming subunits have been shown to acquire resistance against ischaemia reperfusion [13]. The hearts from this phenotype were significantly more resistant to ischaemia/ reperfusion and the isolated cells were also more resistant to hypoxia (Figure 1). Taken all together, it seems that increased expression of SUR2A is cardioprotective. However, it should also be pointed out that there is a recent report suggesting that a phenotype lacking SUR2A in the heart also confers cardioprotection [14]. From a standpoint of what is known about KATP channels up to date it is difficult to explain this outcome of knocking out SUR2 gene. As an example, a lack of SUR2A produces a phenotype lacking sarcolemmal KATP channels [14] that is similar to a Kir6.2 knockout phenotype that also lack sarcolemmal KATP channels [12]. A lack of sarcolemmal KATP channels as a consequence of knocking Kir6.2 out was associated with unchanged response to ischaemia/ reperfusion under non-preconditioned conditions and abolishment of preconditioning [12]. Thus, it is difficult to explain the observed difference between phenotypes lacking Kir6.2 and SUR2A. One possibility is that this difference could be associated with a lack of SUR2B in SUR2 knockout phenotype. SUR2B is expressed in the heart, but its role is yet to be understood, as this subunit does not seem to be a part of sarcolemmal KATP channel Current Opinion in Pharmacology 2009, 9:189–193
protein complex in the myocardium [11]. The KATP channel subunits that do not form sarcolemmal KATP channels in the heart could affect the heart resistance to metabolic stress is demonstrated by a study showing that a lack of SUR1, another KATP channel subunit with unknown function in the heart, decreases myocardial susceptibility to ischaemia reperfusion [15]. On the other hand, mice overexpressing SUR2A did not have affected expression of any other KATP channel subunit suggesting that the observed increase in myocardial resistance to ischaemia/reperfusion is solely due to upregulation of SUR2A [13].
The mechanism underlying SUR2A-mediated cardioprotection How increase in SUR2A alone increases the heart resistance to ischemia is still a matter of vigourous investigation. It has been suggested that an increase in SUR2A increases the level of fully assembled KATP channel complexes and number of sarcolemmal KATP channels in the heart [17–20]. The study of transgenic phenotype overexpressing SUR2A has demonstrated that an increase in SUR2A expression is associated with the increased number of sarcolemmal KATP channels [13]. How an increase of a single subunit would increase a number of fully assembled channels is still not certain, but we have proposed a theory based on our finding that SUR2A is probably the least expressed KATP channel forming subunit (Figure 2; [13]). More specifically, we have found that mRNA levels of SUR2A in the heart are 30 times lower than those Kir6.2 levels [13]. How this transposes www.sciencedirect.com
SURA2 targeting for cardioprotection? Jovanovic and Jovanovic 191
Figure 2
SUR2A is less expressed than Kir6.2 in the heart. Representative progress curves done in duplicate for the real-time PCR amplification of SUR2A and Kir6.2 cDNA (dilution 1:25 ng of template; dilution 2:5 ng of template; dilution 3:1 ng of template). Average cycle threshold was 21.9 0.3 and 28.4 0.8 at dilution 1 for Kir6.2 and SUR2A, respectively, n = 5 for each, P < 0.0001. Modified from [13] with permission.
into difference at protein level is not yet known, but it is likely that some difference between protein levels also exists. If this is the case, it would mean that changes in Kir6.2 levels, unless it is an extreme decrease, would not affect the number of fully assembled KATP channels (as this subunit is anyway in surplus); by contrast, any change in SUR2A amounts would change the number of functional KATP channels in sarcolemma as this is the subunit that is in deficit when KATP channels are to be assembled. However, there is an experimental model of transgenic expression of SUR2A that did not result in increased number of sarcolemmal KATP channels; on the contrary, a decreased number of functional KATP channels was reported [21]. In this particular phenotype, the expression of SUR2A was controlled by aMHC promoter and the SUR2A mRNA levels were 50 times increased when compared with the wild type. On the contrary, our phenotype had only 6.5 times more SUR2A mRNA than the wild type (unpublished observation), but the level of SUR2A protein and number of functional sarcolemmal KATP channels was increased [13]. It is difficult to explain an apparent difference between these two phenotypes. It is possible that a moderate increase in SUR2A levels stimulates formation of fully assembled KATP channels, while a more dramatic increase of SUR2A decreases the number of functional channels. As it was mentioned, cardiac Kir6.2 mRNA levels are physiologically 30 times higher than those values of SUR2A. It is possible that stimulating expression of SUR2A to surpass levels of Kir6.2 would suppress the functional expression of KATP channels from optimized dimeric SUR2A/Kir6.2. A similar notion has been previously raised by [21] to explain why the phenotype expressing SUR2A 50 times more than the wild type would actually have less fully assemble KATP channels. Thus, it seems that a moderate increase in SUR2A expression increases the number of sarcolemmal KATP channels while a more drastic increase of SUR2A has an opposite effect. www.sciencedirect.com
It has been shown that acute increase in the number of sarcolemmal KATP channels mediates ischaemic preconditioning and is associated with preconditioning-induced activation of sarcolemmal KATP channels [22,23]. An activation of sarcolemmal KATP channels induced shortening of action membrane potential, which, in turn, decreased influx of Ca2+ preventing Ca2+ overload, which is the main cause of cell death during ischaemia. The response to hypoxia in cardiac cells with moderately increased SUR2A resembles a response in preconditioned wild type cardiac cells. In cells with moderate increase in SUR2A, hypoxia activates KATP channels much earlier than in cells from the wild type, which could explain how more SUR2A protects against metabolic stress in the heart. What activates KATP channels in hypoxia and ischaemia is still a matter of discussion. Apart from decrease of intracellular ATP, it has been suggested that a decrease of intracellular pH, disruption of cytoskeleton and changes in intracellular levels of other nucleotides stimulate the activation of sarcolemmal KATP channels [24]. We and others have shown in the past few years that KATP channels in the heart exists in vivo as a multiprotein protein complex. It has been suggested that the sarcolemmal KATP channel protein complex in its natural environment may be composed of more proteins than just Kir6.2 and SUR2A. More specifically, it has been shown that Kir6.1, a KATP channel pore-forming subunit predominant in some non-cardiac tissues, and some enzymes regulating intracellular ATP levels and glycolysis (adenylate kinase (AK), creatine kinase (CK), muscle form of lactate dehydrogenase (M-LDH), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), triose-phosphate isomerase (TPI) and pyruvate kinase (PK)) are integral parts of sarcolemmal KATP channel protein complex [5– 10]. Accessory proteins could be essential for sarcolemmal KATP channel regulation and function. It has been suggested that products of catalytic activity of adenylate kinase (ADP), GAPDH (1,3-bisphosphoglycerate) and Current Opinion in Pharmacology 2009, 9:189–193
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M-LDH (lactate) activate KATP channels even in the presence of millimolar ATP [8,9,25,26]. It is therefore possible that more fully assembled KATP channels would produce more of the channel opening compounds, which, in turn, would activate KATP channels. This would be crucial for shortening of action membrane potential, decrease influx of Ca2+ and Ca2+ overload. There are also some additional aspects in KATP channelsmediated cardioprotection; some reports suggest that KATP channels might play a role in cardioprotection independently of their activity. As an example, patients with heart failure of unknown aetiology were discovered to have mutations in ABCC9, the gene encoding the SUR2A protein. This suggests that SUR2A is important for heart well being even in hearts that were not exposed to ischaemia [27]. The protective effect of the sarcolemmal KATP channels is also seen when these channels are expressed and activated in cells that do not generate action membrane potential, including quiescent cardiomyocytes [8,28]. It has been also shown that inhibition of increase in the number of sarcolemmal KATP channels inhibits preconditioninginduced cardioprotection [22]. An enzyme that is responsible for 95% of ATP production in the heart [29], creatine kinase (CK) is a physical part of sarcolemmal KATP channel protein complex [7], meaning that ATP can be produced in subsarcolemmal regions where it is needed to be utilized by non-KATP channel-related processes (active transports, ion homeostasis and so on). Taking these into consideration, it is possible that increase in number of sarcolemmal KATP channels protein complexes results in more subsarcolemmal (even total) ATP production, which is certainly beneficial under conditions of ischaemia and hypoxia. Thus, taken all together, it is possible that some nonchannel properties of KATP channels that are yet to be fully understood mediate cardioprotection afforded by moderately increased levels of SUR2A.
Conclusion A moderate increase in SUR2A expression generates a cardiac phenotype that acquires resistance to different types of metabolic stresses, including ischaemia/reperfusion and hypoxia. SUR2A seems to regulate the number of KATP channels in sarcolemma by being the least expressed KATP channel forming protein. An increased number of sarcolemmal KATP channels protect the heart by regulating duration of action membrane potential and possibly by mechanism that is yet to be fully understood.
References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as: of special interest of outstanding interest 1.
Murry CE, Jennings RB, Reimer KA: Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation 1986, 74:1124-1136.
Current Opinion in Pharmacology 2009, 9:189–193
2.
Murphy E, Steenbergen C: Mechanisms underlying acute protection from cardiac ischemia-reperfusion injury. Physiol Rev 2008, 88:581-609.
3.
Seino S: ATP-sensitive potassium channels: a model of heteromultimeric potassium channel/receptor assemblies. Annu Rev Physiol 1999, 61:337-362.
4.
Zingman LV, Alekseeve AE, Hodgson-Zingman DM, Terzic A: ATP-sensitive potassium channels: metabolic sensing and cardioprotection. J Appl Physiol 2007, 103:1888-1893. A comprehensive review about regulation and role of sarcolemmal KATP channels in the heart. 5.
Cui Y, Giblin JP, Clapp LH, Tinker A: A mechanism for ATPsensitive potassium channel diversity: Functional coassembly of two pore-forming subunits. Proc Natl Acad Sci U S A 2001, 98:729-734.
6.
Carrasco AJ, Dzeja PP, Alekseev AE, Pucar D, Zingman LV, Abraham MR, Hodgson D, Bienengraeber M, Puceat M, Janssen E, Wieringa B et al.: Adenylate kinase phosphotransfer communicates cellular energetic signals to ATP-sensitive potassium channels. Proc Natl Acad Sci U S A 2001, 98:7623-7628.
7.
Crawford RM, Ranki HJ, Botting CH, Budas GR, Jovanovic´ A: Creatine kinase is physically associated with the cardiac ATPsensitive K+ channel in vivo. FASEB J 2002, 16:102-104.
8.
Crawford RM, Budas GR, Jovanovic´ S, Ranki HJ, Wilson TJ, Davies AM, Jovanovic´ A: M-LDH serves as a sarcolemmal K(ATP) channel subunit essential for cell protection against ischemia. EMBO J 2002, 21:3936-3948.
9.
Jovanovic´ S, Du Q, Crawford RM, Budas GR, Stagljar I, Jovanovic´ A: Glyceraldehyde 3-phosphate dehydrogenase serves as an accessory protein of the cardiac sarcolemmal K(ATP) channel. EMBO Rep 2005, 6:848-852.
10. Dhar-Chowdhury P, Harrell MD, Han SY, Jankowska D, Parachuru L, Morrissey A, Srivastava S, Liu W, Malester B, Yoshida H, Coetzee WA: The glycolytic enzymes, glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, and pyruvate kinase are components of the K(ATP) channel macromolecular complex and regulate its function. J Biol Chem 2005, 281:38464-38470. 11. Kane GC, Liu XK, Yamada S, Olson TM, Terzic A: Cardiac KATP channels in health and disease. J Mol Cell Cardiol 2005, 38:937-943. 12. Suzuki M, Sasaki N, Miki T, Sakamoto N, Ohmoto-Sekine Y, Tamagawa M, Seino S, Marba´n E, Nakaya H: Role of sarcolemmal K(ATP) channels in cardioprotection against ischemia/reperfusion injury in mice. J Clin Invest 2002, 109:509-516. 13. Du Q, Jovanovic´ S, Clelland A, Sukhodub A, Budas GR, Phelan K, Murray-Tait V, Malone L, Jovanovic´ A: Overexpression of SUR2A generates a cardiac phenotype resistant to ischaemia. FASEB J 2006, 20:1131-1141. The study providing most of the results described in this review. 14. Stoller D, Kakkar R, Smelley M, Chalupsky K, Earley JU, Shi NQ, Makielski JC, McNally EM: Mice lacking sulfonylurea receptor 2 (SUR2) ATP-sensitive potassium channels are resistant to acute cardiovascular stress. J Mol Cell Cardiol 2007, 43:445-454. A study suggesting that cardioprotection is conferred by a lack of SUR2 in the heart. 15. Elrod JW, Harrell M, Flagg TP, Gundewar S, Magnuson MA, Nichols CG, Coetzee WA, Lefer DJ: Role of sulfonylurea receptor type 1 subunits of ATP-sensitive potassium channels in myocardial ischemia/reperfusion injury. Circulation 2008, 117:1405-1413. 16. Burke MA, Mutharasan RK, Ardehali H: The sulfonylurea receptor, an atypical ATP-binding cassette protein, and its regulation of the KATP channel. Circ Res 2008, 102:164-176. A comprehensive review about SUR2A in the heart. 17. Ranki HJ, Budas GR, Crawford RM, Jovanovic´ A: Genderspecific difference in cardiac ATP-sensitive K+ channels. J Am Coll Cardiol 2001, 38:906-915. www.sciencedirect.com
SURA2 targeting for cardioprotection? Jovanovic and Jovanovic 193
18. Ranki HJ, Crawford RM, Budas GR, Jovanovic´ A: Ageing is associated with decrease in number of sarcolemmal ATPsensitive K+ channels in a gender-dependent manner. Mech Ageing Dev 2002, 123:695-705.
24. Alekseev AE, Hodgson DM, Karger AB, Park S, Zingman LV, Terzic A: ATP-sensitive K+ channel channel/enzyme multimer: metabolic gating in the heart. J Mol Cell Cardiol 2005, 38:895-905.
19. Ranki HJ, Budas GR, Crawford RM, Davies AM, Jovanovic´ A: 17bestradiol regulates expression of KATP channels in heartderived H9c2 cells. J Am Coll Cardiol 2002, 40:367-374.
25. Elvir-Mairena JR, Jovanovic´ A, Gomez LA, Alekseev AE, Terzic A: Reversal of the ATP-liganded state of ATP-sensitive K+ channels by adenylate kinase activity. J Biol Chem 1996, 271:31903-31908.
20. Crawford RM, Jovanovic´ S, Budas GR, Davies AM, Lad H, Wenger RH, Robertson KA, Roy DJ, Ranki HJ, Jovanovic´ A: Chronic mild hypoxia protects heart-derived H9c2 cells against acute hypoxia/reoxygenation by regulating expression of the SUR2A subunit of the ATP-sensitive K+ channels. J Biol Chem 2003, 278:31444-31455. 21. Flagg TP, Remedi MS, Masia R, Gomes J, McLerie M, Lopatin AN, Nichols CG: Transgenic overexpression of SUR1 in the heart suppresses sarcolemmal K(ATP). J Mol Cell Cardiol 2005, 39:647-656. A study suggesting that a dramatic increase of SUR2A expression decreases the number of functional KATP channels. 22. Budas GR, Jovanovic´ S, Crawford RM, Jovanovic´ A: Hypoxiainduced preconditioning in adult stimulated cardiomyocytes is mediated by the opening and trafficking of sarcolemmal KATP channels. FASEB J 2004, 18:1046-1048. 23. Sukhodub A, Jovanovic´ S, Du Q, Budas GR, Clelland A, Shen M, Sakamoto K, Tian R, Jovanovic´ A: AMP-activated protein kinase mediates preconditioning in cardiomyocytes by regulating activity and trafficking of sarcolemmal ATP-sensitive K+ channels. J Cell Physiol 2007, 210:224-236.
www.sciencedirect.com
26. Jovanovic´ S, Jovanovic´ A: High glucose regulates the activity of cardiac sarcolemmal ATP-sensitive K+ channels via 1,3-bisphosphoglycerate: a novel link between cardiac membrane excitability and glucose metabolism. Diabetes 2005, 54:383-393. 27. Bienengraeber M, Olson TM, Selivanov VA, Kathmann EC, O’Cochlain F, Gao F, Karger AB, Ballew JD, Hodgson DM, Zingman LV, Pang YP et al.: ABCC9 mutations identified in human dilated cardiomyopathy disrupt catalytic KATP channel gating. Nat Genet 2004, 36:382-387. 28. Jovanovic´ A, Jovanovic´ S, Lorenz E, Terzic A: Recombinant cardiac ATP-sensitive K+ channel subunits confer resistance towards chemical hypoxia-reoxygenation injury. Circulation 1998, 98:1548-1555. 29. Saks V, Kaambre T, Guzun R, Anmann T, Sikk P, Schlattner U, Wallimann T, Aliev M, Vendelin M: The creatine kinase phosphotransfer network: thermodynamic and kinetic considerations, the impact of the mitochondrial outer membrane and modelling approaches. Subcell Biochem 2007, 46:27-65.
Current Opinion in Pharmacology 2009, 9:189–193
SURA2 targeting for cardioprotection? Aleksandar Jovanovic´ and Sofija Jovanovic´ SUR2A is an ATP-binding protein known to serve as a regulatory subunit of metabolic-sensing, cardioprotective sarcolemmal ATP-sensitive K+ (KATP) channels. It has been recently found that a moderate increase in expression of SUR2A protects the heart against different types of metabolic stresses, including ischaemia/reperfusion and hypoxia. Although the sarcolemmal KATP channel is a multiprotein complex composed of many proteins in vivo, it seems that an increase in SUR2A levels is sufficient to increase the number of sarcolemmal KATP channels. This effect of SUR2A could be due to SUR2A being the rate-limiting factor in generating fully composed sarcolemmal KATP channels. An increased number of sarcolemmal KATP channels seems to protect the heart by regulating action membrane potential, inhibiting Ca2+ influx and preventing Ca2+ overload, although an additional yet to be recognised mechanism independent of KATP channels activity cannot be excluded. Address Division of Medical Sciences, Ninewells Hospital & Medical School, University of Dundee, Dundee DD1 9SY, UK Corresponding author: Jovanovic´, Aleksandar ([email protected]) and
Current Opinion in Pharmacology 2009, 9:189–193 This review comes from a themed issue on Cardiovascular and renal Edited by Michael Curtis Available online 10th December 2008 1471-4892/$ – see front matter # 2008 Elsevier Ltd. All rights reserved. DOI 10.1016/j.coph.2008.11.003
Introduction In 1986, Murry et al. discovered that brief periods of blood vessel occlusion and reperfusion administered before a sustained ischaemic episode lead to a reduction in myocardial infarct size [1]. This and subsequent studies have indicated that brief ischaemia or hypoxia are likely to switch on signalling pathways that prevent or delay lethal outcomes of stressed cardiac cells and this phenomenon is known as cardioprotection [2]. ATP-sensitive K+ (KATP) channels are present in different tissues and are combinations of an inwardly rectifying K+ channel, Kir6.1 or/and Kir6.2 and a regulatory subunit, the sulfonylurea receptor SUR1, SUR2A or SUR2B [3]. Physical association of the Kir6.2 and SUR2A isoforms www.sciencedirect.com
generates cardiac KATP channels that are expressed in high density at the sarcolemma [4]. More recent studies have suggested that the sarcolemmal KATP channel protein complex may be composed of more proteins than just Kir6.2 and SUR2A [5–10]. Since intracellular ATP regulates the activity of KATP channels, it is consensus view that this ion conductance couples metabolic state of the cell with the membrane excitability [4]. In numerous studies, it has been demonstrated that KATP channel openers, drugs that promote opening of KATP channels, decrease infarct size, mimic ischaemic preconditioning and improve functional and energetic recovery of cardiac muscle following ischaemic and hypoxic insults [11]. More recently, it has been shown that targeting specific subunits of sarcolemmal KATP channels protein complex might have significant consequences on myocardial resistance to different types of metabolic stresses, including ischaemia/reperfusion and hypoxia [12,13,14,15]. One of the subunits that can be targeted to manipulate with the cardiac resistance to metabolic stress is SUR2A. Molecular biology, biochemistry and physiology of SUR2A have been reviewed recently in great details [16] and readers can use this review to learn more about structural/physiological/functional aspects of SUR2A. Here, we will provide a brief outline of a relationship between SUR2A and cardioprotection and highlight how targeting SUR2A alone might be a potential strategy to protect the heart against metabolic stress.
Evidence that changes in SUR2A expression regulates myocardial resistance to ischaemia/ reperfusion and hypoxia The first evidence demonstrating that SUR2A level might be particularly important in myocardial resistance to ischaemia was presented in a study addressing genderspecific difference in sarcolemmal KATP channels. It has been shown that females have more assembled functional sarcolemmal KATP channels than males and this was associated with cardiac cells from females being more resistant to ischaemia reperfusion [17]. The most intriguing finding was that no gender-specific difference was observed in mRNA levels of any other KATP channel subunits apart from SUR2A [17]. Further studies have shown that any condition that increases the number of sarcolemmal KATP channels (young age, female gender, oestrogen treatment and chronic hypoxia) is associated with a sole increase in SUR2A mRNA and with increased resistance to ischaemia/reperfusion [17–20]. This has suggested that upregulation of SUR2A is sufficient to Current Opinion in Pharmacology 2009, 9:189–193
190 Cardiovascular and renal
Figure 1
Increase in SUR2A expression confers myocardial resistance against ischaemia/reperfusion. Typical photographs of myocardial slices from wild type and transgenic mice with increased SUR2A expression after exposure to 30 min-long ischaemia followed by 30 min-long reperfusion. Infarcted areas are pale/gray (indicated also with the blue arrows) whereas viable myocardium is dark/red. The average size of myocardial infarction was 21.2 5.1% (n = 17) in wild type and 5.3 1.2% (n = 6) in transgenic mice. This difference was statistically significant (P = 0.02). Modified from [13] with permission.
increase the resistance to ischaemia/reperfusion. This kind of conclusion was particularly supported by a study showing that chronic mild hypoxia upregulates SUR2A without affecting the expression of any other gene [20]. In a latter study, transgenic animals overexpressing SUR2A without changes of expression of other KATP channel forming subunits have been shown to acquire resistance against ischaemia reperfusion [13]. The hearts from this phenotype were significantly more resistant to ischaemia/ reperfusion and the isolated cells were also more resistant to hypoxia (Figure 1). Taken all together, it seems that increased expression of SUR2A is cardioprotective. However, it should also be pointed out that there is a recent report suggesting that a phenotype lacking SUR2A in the heart also confers cardioprotection [14]. From a standpoint of what is known about KATP channels up to date it is difficult to explain this outcome of knocking out SUR2 gene. As an example, a lack of SUR2A produces a phenotype lacking sarcolemmal KATP channels [14] that is similar to a Kir6.2 knockout phenotype that also lack sarcolemmal KATP channels [12]. A lack of sarcolemmal KATP channels as a consequence of knocking Kir6.2 out was associated with unchanged response to ischaemia/ reperfusion under non-preconditioned conditions and abolishment of preconditioning [12]. Thus, it is difficult to explain the observed difference between phenotypes lacking Kir6.2 and SUR2A. One possibility is that this difference could be associated with a lack of SUR2B in SUR2 knockout phenotype. SUR2B is expressed in the heart, but its role is yet to be understood, as this subunit does not seem to be a part of sarcolemmal KATP channel Current Opinion in Pharmacology 2009, 9:189–193
protein complex in the myocardium [11]. The KATP channel subunits that do not form sarcolemmal KATP channels in the heart could affect the heart resistance to metabolic stress is demonstrated by a study showing that a lack of SUR1, another KATP channel subunit with unknown function in the heart, decreases myocardial susceptibility to ischaemia reperfusion [15]. On the other hand, mice overexpressing SUR2A did not have affected expression of any other KATP channel subunit suggesting that the observed increase in myocardial resistance to ischaemia/reperfusion is solely due to upregulation of SUR2A [13].
The mechanism underlying SUR2A-mediated cardioprotection How increase in SUR2A alone increases the heart resistance to ischemia is still a matter of vigourous investigation. It has been suggested that an increase in SUR2A increases the level of fully assembled KATP channel complexes and number of sarcolemmal KATP channels in the heart [17–20]. The study of transgenic phenotype overexpressing SUR2A has demonstrated that an increase in SUR2A expression is associated with the increased number of sarcolemmal KATP channels [13]. How an increase of a single subunit would increase a number of fully assembled channels is still not certain, but we have proposed a theory based on our finding that SUR2A is probably the least expressed KATP channel forming subunit (Figure 2; [13]). More specifically, we have found that mRNA levels of SUR2A in the heart are 30 times lower than those Kir6.2 levels [13]. How this transposes www.sciencedirect.com
SURA2 targeting for cardioprotection? Jovanovic and Jovanovic 191
Figure 2
SUR2A is less expressed than Kir6.2 in the heart. Representative progress curves done in duplicate for the real-time PCR amplification of SUR2A and Kir6.2 cDNA (dilution 1:25 ng of template; dilution 2:5 ng of template; dilution 3:1 ng of template). Average cycle threshold was 21.9 0.3 and 28.4 0.8 at dilution 1 for Kir6.2 and SUR2A, respectively, n = 5 for each, P < 0.0001. Modified from [13] with permission.
into difference at protein level is not yet known, but it is likely that some difference between protein levels also exists. If this is the case, it would mean that changes in Kir6.2 levels, unless it is an extreme decrease, would not affect the number of fully assembled KATP channels (as this subunit is anyway in surplus); by contrast, any change in SUR2A amounts would change the number of functional KATP channels in sarcolemma as this is the subunit that is in deficit when KATP channels are to be assembled. However, there is an experimental model of transgenic expression of SUR2A that did not result in increased number of sarcolemmal KATP channels; on the contrary, a decreased number of functional KATP channels was reported [21]. In this particular phenotype, the expression of SUR2A was controlled by aMHC promoter and the SUR2A mRNA levels were 50 times increased when compared with the wild type. On the contrary, our phenotype had only 6.5 times more SUR2A mRNA than the wild type (unpublished observation), but the level of SUR2A protein and number of functional sarcolemmal KATP channels was increased [13]. It is difficult to explain an apparent difference between these two phenotypes. It is possible that a moderate increase in SUR2A levels stimulates formation of fully assembled KATP channels, while a more dramatic increase of SUR2A decreases the number of functional channels. As it was mentioned, cardiac Kir6.2 mRNA levels are physiologically 30 times higher than those values of SUR2A. It is possible that stimulating expression of SUR2A to surpass levels of Kir6.2 would suppress the functional expression of KATP channels from optimized dimeric SUR2A/Kir6.2. A similar notion has been previously raised by [21] to explain why the phenotype expressing SUR2A 50 times more than the wild type would actually have less fully assemble KATP channels. Thus, it seems that a moderate increase in SUR2A expression increases the number of sarcolemmal KATP channels while a more drastic increase of SUR2A has an opposite effect. www.sciencedirect.com
It has been shown that acute increase in the number of sarcolemmal KATP channels mediates ischaemic preconditioning and is associated with preconditioning-induced activation of sarcolemmal KATP channels [22,23]. An activation of sarcolemmal KATP channels induced shortening of action membrane potential, which, in turn, decreased influx of Ca2+ preventing Ca2+ overload, which is the main cause of cell death during ischaemia. The response to hypoxia in cardiac cells with moderately increased SUR2A resembles a response in preconditioned wild type cardiac cells. In cells with moderate increase in SUR2A, hypoxia activates KATP channels much earlier than in cells from the wild type, which could explain how more SUR2A protects against metabolic stress in the heart. What activates KATP channels in hypoxia and ischaemia is still a matter of discussion. Apart from decrease of intracellular ATP, it has been suggested that a decrease of intracellular pH, disruption of cytoskeleton and changes in intracellular levels of other nucleotides stimulate the activation of sarcolemmal KATP channels [24]. We and others have shown in the past few years that KATP channels in the heart exists in vivo as a multiprotein protein complex. It has been suggested that the sarcolemmal KATP channel protein complex in its natural environment may be composed of more proteins than just Kir6.2 and SUR2A. More specifically, it has been shown that Kir6.1, a KATP channel pore-forming subunit predominant in some non-cardiac tissues, and some enzymes regulating intracellular ATP levels and glycolysis (adenylate kinase (AK), creatine kinase (CK), muscle form of lactate dehydrogenase (M-LDH), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), triose-phosphate isomerase (TPI) and pyruvate kinase (PK)) are integral parts of sarcolemmal KATP channel protein complex [5– 10]. Accessory proteins could be essential for sarcolemmal KATP channel regulation and function. It has been suggested that products of catalytic activity of adenylate kinase (ADP), GAPDH (1,3-bisphosphoglycerate) and Current Opinion in Pharmacology 2009, 9:189–193
192 Cardiovascular and renal
M-LDH (lactate) activate KATP channels even in the presence of millimolar ATP [8,9,25,26]. It is therefore possible that more fully assembled KATP channels would produce more of the channel opening compounds, which, in turn, would activate KATP channels. This would be crucial for shortening of action membrane potential, decrease influx of Ca2+ and Ca2+ overload. There are also some additional aspects in KATP channelsmediated cardioprotection; some reports suggest that KATP channels might play a role in cardioprotection independently of their activity. As an example, patients with heart failure of unknown aetiology were discovered to have mutations in ABCC9, the gene encoding the SUR2A protein. This suggests that SUR2A is important for heart well being even in hearts that were not exposed to ischaemia [27]. The protective effect of the sarcolemmal KATP channels is also seen when these channels are expressed and activated in cells that do not generate action membrane potential, including quiescent cardiomyocytes [8,28]. It has been also shown that inhibition of increase in the number of sarcolemmal KATP channels inhibits preconditioninginduced cardioprotection [22]. An enzyme that is responsible for 95% of ATP production in the heart [29], creatine kinase (CK) is a physical part of sarcolemmal KATP channel protein complex [7], meaning that ATP can be produced in subsarcolemmal regions where it is needed to be utilized by non-KATP channel-related processes (active transports, ion homeostasis and so on). Taking these into consideration, it is possible that increase in number of sarcolemmal KATP channels protein complexes results in more subsarcolemmal (even total) ATP production, which is certainly beneficial under conditions of ischaemia and hypoxia. Thus, taken all together, it is possible that some nonchannel properties of KATP channels that are yet to be fully understood mediate cardioprotection afforded by moderately increased levels of SUR2A.
Conclusion A moderate increase in SUR2A expression generates a cardiac phenotype that acquires resistance to different types of metabolic stresses, including ischaemia/reperfusion and hypoxia. SUR2A seems to regulate the number of KATP channels in sarcolemma by being the least expressed KATP channel forming protein. An increased number of sarcolemmal KATP channels protect the heart by regulating duration of action membrane potential and possibly by mechanism that is yet to be fully understood.
References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as: of special interest of outstanding interest 1.
Murry CE, Jennings RB, Reimer KA: Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation 1986, 74:1124-1136.
Current Opinion in Pharmacology 2009, 9:189–193
2.
Murphy E, Steenbergen C: Mechanisms underlying acute protection from cardiac ischemia-reperfusion injury. Physiol Rev 2008, 88:581-609.
3.
Seino S: ATP-sensitive potassium channels: a model of heteromultimeric potassium channel/receptor assemblies. Annu Rev Physiol 1999, 61:337-362.
4.
Zingman LV, Alekseeve AE, Hodgson-Zingman DM, Terzic A: ATP-sensitive potassium channels: metabolic sensing and cardioprotection. J Appl Physiol 2007, 103:1888-1893. A comprehensive review about regulation and role of sarcolemmal KATP channels in the heart. 5.
Cui Y, Giblin JP, Clapp LH, Tinker A: A mechanism for ATPsensitive potassium channel diversity: Functional coassembly of two pore-forming subunits. Proc Natl Acad Sci U S A 2001, 98:729-734.
6.
Carrasco AJ, Dzeja PP, Alekseev AE, Pucar D, Zingman LV, Abraham MR, Hodgson D, Bienengraeber M, Puceat M, Janssen E, Wieringa B et al.: Adenylate kinase phosphotransfer communicates cellular energetic signals to ATP-sensitive potassium channels. Proc Natl Acad Sci U S A 2001, 98:7623-7628.
7.
Crawford RM, Ranki HJ, Botting CH, Budas GR, Jovanovic´ A: Creatine kinase is physically associated with the cardiac ATPsensitive K+ channel in vivo. FASEB J 2002, 16:102-104.
8.
Crawford RM, Budas GR, Jovanovic´ S, Ranki HJ, Wilson TJ, Davies AM, Jovanovic´ A: M-LDH serves as a sarcolemmal K(ATP) channel subunit essential for cell protection against ischemia. EMBO J 2002, 21:3936-3948.
9.
Jovanovic´ S, Du Q, Crawford RM, Budas GR, Stagljar I, Jovanovic´ A: Glyceraldehyde 3-phosphate dehydrogenase serves as an accessory protein of the cardiac sarcolemmal K(ATP) channel. EMBO Rep 2005, 6:848-852.
10. Dhar-Chowdhury P, Harrell MD, Han SY, Jankowska D, Parachuru L, Morrissey A, Srivastava S, Liu W, Malester B, Yoshida H, Coetzee WA: The glycolytic enzymes, glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, and pyruvate kinase are components of the K(ATP) channel macromolecular complex and regulate its function. J Biol Chem 2005, 281:38464-38470. 11. Kane GC, Liu XK, Yamada S, Olson TM, Terzic A: Cardiac KATP channels in health and disease. J Mol Cell Cardiol 2005, 38:937-943. 12. Suzuki M, Sasaki N, Miki T, Sakamoto N, Ohmoto-Sekine Y, Tamagawa M, Seino S, Marba´n E, Nakaya H: Role of sarcolemmal K(ATP) channels in cardioprotection against ischemia/reperfusion injury in mice. J Clin Invest 2002, 109:509-516. 13. Du Q, Jovanovic´ S, Clelland A, Sukhodub A, Budas GR, Phelan K, Murray-Tait V, Malone L, Jovanovic´ A: Overexpression of SUR2A generates a cardiac phenotype resistant to ischaemia. FASEB J 2006, 20:1131-1141. The study providing most of the results described in this review. 14. Stoller D, Kakkar R, Smelley M, Chalupsky K, Earley JU, Shi NQ, Makielski JC, McNally EM: Mice lacking sulfonylurea receptor 2 (SUR2) ATP-sensitive potassium channels are resistant to acute cardiovascular stress. J Mol Cell Cardiol 2007, 43:445-454. A study suggesting that cardioprotection is conferred by a lack of SUR2 in the heart. 15. Elrod JW, Harrell M, Flagg TP, Gundewar S, Magnuson MA, Nichols CG, Coetzee WA, Lefer DJ: Role of sulfonylurea receptor type 1 subunits of ATP-sensitive potassium channels in myocardial ischemia/reperfusion injury. Circulation 2008, 117:1405-1413. 16. Burke MA, Mutharasan RK, Ardehali H: The sulfonylurea receptor, an atypical ATP-binding cassette protein, and its regulation of the KATP channel. Circ Res 2008, 102:164-176. A comprehensive review about SUR2A in the heart. 17. Ranki HJ, Budas GR, Crawford RM, Jovanovic´ A: Genderspecific difference in cardiac ATP-sensitive K+ channels. J Am Coll Cardiol 2001, 38:906-915. www.sciencedirect.com
SURA2 targeting for cardioprotection? Jovanovic and Jovanovic 193
18. Ranki HJ, Crawford RM, Budas GR, Jovanovic´ A: Ageing is associated with decrease in number of sarcolemmal ATPsensitive K+ channels in a gender-dependent manner. Mech Ageing Dev 2002, 123:695-705.
24. Alekseev AE, Hodgson DM, Karger AB, Park S, Zingman LV, Terzic A: ATP-sensitive K+ channel channel/enzyme multimer: metabolic gating in the heart. J Mol Cell Cardiol 2005, 38:895-905.
19. Ranki HJ, Budas GR, Crawford RM, Davies AM, Jovanovic´ A: 17bestradiol regulates expression of KATP channels in heartderived H9c2 cells. J Am Coll Cardiol 2002, 40:367-374.
25. Elvir-Mairena JR, Jovanovic´ A, Gomez LA, Alekseev AE, Terzic A: Reversal of the ATP-liganded state of ATP-sensitive K+ channels by adenylate kinase activity. J Biol Chem 1996, 271:31903-31908.
20. Crawford RM, Jovanovic´ S, Budas GR, Davies AM, Lad H, Wenger RH, Robertson KA, Roy DJ, Ranki HJ, Jovanovic´ A: Chronic mild hypoxia protects heart-derived H9c2 cells against acute hypoxia/reoxygenation by regulating expression of the SUR2A subunit of the ATP-sensitive K+ channels. J Biol Chem 2003, 278:31444-31455. 21. Flagg TP, Remedi MS, Masia R, Gomes J, McLerie M, Lopatin AN, Nichols CG: Transgenic overexpression of SUR1 in the heart suppresses sarcolemmal K(ATP). J Mol Cell Cardiol 2005, 39:647-656. A study suggesting that a dramatic increase of SUR2A expression decreases the number of functional KATP channels. 22. Budas GR, Jovanovic´ S, Crawford RM, Jovanovic´ A: Hypoxiainduced preconditioning in adult stimulated cardiomyocytes is mediated by the opening and trafficking of sarcolemmal KATP channels. FASEB J 2004, 18:1046-1048. 23. Sukhodub A, Jovanovic´ S, Du Q, Budas GR, Clelland A, Shen M, Sakamoto K, Tian R, Jovanovic´ A: AMP-activated protein kinase mediates preconditioning in cardiomyocytes by regulating activity and trafficking of sarcolemmal ATP-sensitive K+ channels. J Cell Physiol 2007, 210:224-236.
www.sciencedirect.com
26. Jovanovic´ S, Jovanovic´ A: High glucose regulates the activity of cardiac sarcolemmal ATP-sensitive K+ channels via 1,3-bisphosphoglycerate: a novel link between cardiac membrane excitability and glucose metabolism. Diabetes 2005, 54:383-393. 27. Bienengraeber M, Olson TM, Selivanov VA, Kathmann EC, O’Cochlain F, Gao F, Karger AB, Ballew JD, Hodgson DM, Zingman LV, Pang YP et al.: ABCC9 mutations identified in human dilated cardiomyopathy disrupt catalytic KATP channel gating. Nat Genet 2004, 36:382-387. 28. Jovanovic´ A, Jovanovic´ S, Lorenz E, Terzic A: Recombinant cardiac ATP-sensitive K+ channel subunits confer resistance towards chemical hypoxia-reoxygenation injury. Circulation 1998, 98:1548-1555. 29. Saks V, Kaambre T, Guzun R, Anmann T, Sikk P, Schlattner U, Wallimann T, Aliev M, Vendelin M: The creatine kinase phosphotransfer network: thermodynamic and kinetic considerations, the impact of the mitochondrial outer membrane and modelling approaches. Subcell Biochem 2007, 46:27-65.
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