Is the Molecular Composition of KATPChannels more Complex than Originally Thought?

Pountney, David; Sun, Zhuoqian; Porter, Lisa M.; Nitabach, Michael N.; Nakamura, Tomoe Y.; Holmes, Douglas; Rosner, Erika; Kaneko, Minoru; Manaris, Tom; Holmes, Todd C.; Coetzee, William A.

doi:10.1006/jmcc.2001.1407

Cited by 55 publications

(33 citation statements)

References 17 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This study favors the view that pore-forming KIR6.1 and KIR6.2 subunits can form heteromultimers in primary cultured rat cardiac myocytes and reconciles previously held differences in opinion on K ATP channel assembly in reconstitution studies and cardiomyocytes (12,19,28,32 (36). K ATP channel composition in atrial myocytes is not yet established, but the single channel conductance, at room temperature, has been shown to range from 58 pS in rats (8) to 73 pS in humans (44).…”

Section: Perspectivessupporting

confidence: 79%

“…Suggestive evidence that KIR6.1 can associate with KIR6.2 stems from channel reconstitution in heterologous expression systems. In HEK-293 cells expressing KIR6.1 and KIR6.2, immunoprecipitation revealed KIR6.1/KIR6.2 complexes (12,28). In COS7 cells, coexpression of the fusion protein KIR6.1-KIR6.2 with SUR2A resulted in functional K ATP channels that had a channel conductance of 59 pS, intermediate between KIR6.2 homotetramers (67 pS) and KIR6.1 homotetramers (34 pS) (19).…”

mentioning

confidence: 99%

See 1 more Smart Citation

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Bever¹,

Poitry²,

Faure³

et al. 2004

American Journal of Physiology-Heart and Circulatory Physiology

View full text Add to dashboard Cite

. Pore loop-mutated rat KIR6.1 and KIR6.2 suppress KATP current in rat cardiomyocytes. Am J Physiol Heart Circ Physiol 287: H850 -H859, 2004. First published March 25, 2004 10.1152/ajpheart.00054.2004.-Cardiomyocytes express mRNA for all major subunits of ATP-sensitive potassium (KATP) channels: KIR6.1, KIR6.2, SUR1A, SUR2A, and SUR2B. It has remained controversial as to whether KIR6.1 may associate with KIR6.2 to form the tetrameric pore of KATP channels in cardiomyocytes. To explore this possibility, cultured rat cardiomyocytes were examined for an inhibition of K ATP current by overexpression of pore loop-mutated (inactive) KIR6.x. Bicistronic plasmids were constructed encoding loop-mutated (AFA or SFG for GFG) rat KIR6.x followed by EGFP. In ventricular myocytes, the overexpression of KIR6.1SFG-pIRES2-EGFP or KIR6.2AFA-pIRES2-EGFP DNA caused, after 72 h, a major decrease of K ATP current density of 85.8% and 82.7%, respectively (P Ͻ 0.01), relative to EGFP controls (59 Ϯ 9 pA/pF). In atrial myocytes, overexpression of these poremutated KIR6.x by 6.0-fold and 10.6-fold, as assessed by quantitative immunohistochemistry, caused a decrease of KATP current density of 73.7% and 58.5%, respectively (P Ͻ 0.01). Expression of wild-type rat KIR6.2 increased the ventricular and atrial K ATP current density by 58.3% and 42.9%, respectively (P Ͻ 0.01), relative to corresponding EGFP controls, indicating a reserve of SUR to accommodate increased KIR6.x trafficking to the sarcolemma. The results favor the view that KIR6.1 may associate with KIR6.2 to form heterotetrameric pores of native K ATP channels in cardiomyocytes.

show abstract

Section: Perspectivessupporting

confidence: 79%

mentioning

confidence: 99%

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Bever¹,

Poitry²,

Faure³

et al. 2004

American Journal of Physiology-Heart and Circulatory Physiology

View full text Add to dashboard Cite

show abstract

“…Our immunocytochemistry data demonstrate a degree of Kir6.1 surface expression in immature myocytes (in addition to strong intracellular expression). Because Kir6.1 and Kir6.2 have the potential to interact with each other to form functional heteromultimers (18,20,21,(41)(42)(43), it is tempting to consider the possibility that heteromultimeric interaction between Kir6.1 and Kir6.2 subunits may occur to form the immature K ATP channels, similar to the situation in the vascular endothelium (21). This is a particularly tempting thought, considering that immature K ATP channels have a smaller unitary conductance (3), similar to that formed by heteromeric Kir6.1/Kir6.2 channels (41,44 -46).…”

Section: Discussionmentioning

confidence: 99%

“…We developed an anti-Kir6.1 antibody (NAF1) that we previously demonstrated specifically to detect Kir6.1 (and not Kir6.2 subunits) in Western blotting, with an electrophoretic mobility of the Kir6.1 protein corresponding to~44 kD (20,21). We also previously showed the Santa Cruz G-16 antibody specifically to detect the Kir6.2 protein as a 38 kD band in Western blotting (21).…”

Section: K Atp Channel Subunits and Maturationmentioning

confidence: 99%

Expression of ATP-Sensitive K+ Channel Subunits during Perinatal Maturation in the Mouse Heart

et al. 2005

View full text Add to dashboard Cite

Prevailing data suggest that sarcolemmal ATP-sensitive (K ATP ) channels in the adult heart consist of Kir6.2 and SUR2A subunits, but the expression of other K ATP channel subunits (including SUR1, SUR2B, and Kir6.1) is poorly defined. The situation is even less clear for the immature heart, which shows a remarkable resistance to hypoxia and metabolic stress. The hypoxia-induced action potential shortening and opening of sarcolemmal K ATP channels that occurs in adults is less prominent in the immature heart. This might be due in part to the different biophysical and pharmacological properties of K ATP channels of immature and adult K ATP channels. Because these properties are largely conferred by subunit composition, it is important to examine the relative expression levels of the various K ATP channel subunits during maturation. We therefore used RNAse protection assays, reverse transcription-PCR approaches, and Western blotting to characterize the mRNA and protein expression profiles of K ATP channel subunits in fetal, neonatal, and adult mouse heart. Our data indicate that each of the K ATP channel subunits (Kir6.1, Kir6.2, SUR1, SUR2A, and SUR2B) is expressed in the mouse heart at all of the developmental time points studied. However, the expression level of each of the subunits is low in the fetal heart and progressively increases with maturation. Each of the subunits seems to be expressed in ventricular myocytes with a subcellular expression pattern matching that found in the adult. Our data suggest that the K ATP channel composition may change during maturation, which has important implications for K ATP channel function in the developing heart. ATP-sensitive K ϩ (K ATP ) channels are robustly expressed in the immature heart (1,2). However, K ATP channels in the immature heart may differ in several respects to those of adult cardiac tissue. In immature rabbit ventricle, K ATP channel open probability is roughly the same as in the adult, but they exhibit a significantly smaller single-channel conductance (~55 pS) and channel density, as defined by the number of open channels per patch (3). In the immature rat heart, K ATP channel density is also high, but the single-channel conductance and the mean open time are the same as in the adult (2). Reports describing a smaller unitary conductance in neonatal rat atrium (4) suggested that there may be regional differences. The immature rat heart K ATP channel has a lower open probability, and the channels are much more sensitive to block by intracellular ATP (2,5). In addition to these biophysical changes, there are reports that immature K ATP channels may have a different pharmacological profile. The neonatal atrial K ATP channel shows a unique functional and pharmacological profile resembling the pancreatic ␤ cell channel for its unusually high affinity for glibenclamide and diazoxide (4,5). These characteristics suggest that neonatal K ATP channels may differ in their molecular composition relative to the mature heart. K ATP channels are believed to be hetero-...

show abstract

“…Kir6.1 and Kir6.2, being 64% identical, display overlapping tissue expressions and form heteromeric complexes when coexpressed (Babenko et al, 2000;Cui et al, 2001;Pountney et al, 2001;van Bever et al, 2004). On the contrary, based on the study from Giblin et al, it is generally accepted that SUR1 cannot coassemble with SUR2 to form K ATP channels (Giblin et al, 2002;Babenko, 2005;Neagoe and Schwappach, 2005;Tricarico et al, 2006), even though SUR1 and SUR2 share ‫%56ف‬ identity and are coexpressed in many cell types (Baron et al, 1999;Liss et al, 1999;Shi et al, 2005;Tricarico et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Sulfonylurea Receptors Type 1 and 2A Randomly Assemble to Form Heteromeric K_ATPChannels of Mixed Subunit Composition

Chan¹,

Wheeler²,

Csanády³

2008

J Cell Biol

View full text Add to dashboard Cite

ATP-sensitive potassium (K ATP ) channels play important roles in regulating insulin secretion, controlling vascular tone, and protecting cells against metabolic stresses. K ATP channels are heterooctamers of four pore-forming inwardly rectifying (Kir6.2) subunits and four sulfonylurea receptor (SUR) subunits. K ATP channels containing SUR1 (e.g. pancreatic) and SUR2A (e.g. cardiac) display distinct metabolic sensitivities and pharmacological profi les. The reported expression of both SUR1 and SUR2 together with Kir6.2 in some cells raises the possibility that heteromeric channels containing both SUR subtypes might exist. To test whether SUR1 can coassemble with SUR2A to form functional K ATP channels, we made tandem constructs by fusing SUR to either a wild-type (WT) or a mutant N160D Kir6.2 subunit. The latter mutation greatly increases the sensitivity of K ATP channels to block by intracellular spermine. We expressed, individually and in combinations, tandem constructs SUR1-Kir6.2 (S1-WT), SUR1-Kir6.2[N160D] (S1-ND), and SUR2A-Kir6.2[N160D] (S2-ND) in Xenopus oocytes, and studied the voltage dependence of spermine block in insideout macropatches over a range of spermine concentrations and RNA mixing ratios. Each tandem construct expressed alone supported macroscopic K + currents with pharmacological properties indistinguishable from those of the respective native channel types. Spermine sensitivity was low for S1-WT but high for S1-ND and S2-ND. Coexpression of S1-WT and S1-ND generated current components with intermediate spermine sensitivities indicating the presence of channel populations containing both types of Kir subunits at all possible stoichiometries. The relative abundances of these populations, determined by global fi tting over a range of conditions, followed binomial statistics, suggesting that WT and N160D Kir6.2 subunits coassemble indiscriminately. Coexpression of S1-WT with S2-ND also yielded current components with intermediate spermine sensitivities, suggesting that SUR1 and SUR2A randomly coassemble into functional K ATP channels. Further pharmacological characterization confi rmed coassembly of not only S1-WT and S2-ND, but also of coexpressed free SUR1, SUR2A, and Kir6.2 into functional heteromeric channels.

show abstract

Is the Molecular Composition of KATPChannels more Complex than Originally Thought?

Cited by 55 publications

References 17 publications

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Expression of ATP-Sensitive K+ Channel Subunits during Perinatal Maturation in the Mouse Heart

Sulfonylurea Receptors Type 1 and 2A Randomly Assemble to Form Heteromeric K_ATPChannels of Mixed Subunit Composition

Contact Info

Product

Resources

About

Is the Molecular Composition of KATPChannels more Complex than Originally Thought?

Cited by 55 publications

References 17 publications

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress KATPcurrent in rat cardiomyocytes

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress KATPcurrent in rat cardiomyocytes

Expression of ATP-Sensitive K+ Channel Subunits during Perinatal Maturation in the Mouse Heart

Sulfonylurea Receptors Type 1 and 2A Randomly Assemble to Form Heteromeric KATPChannels of Mixed Subunit Composition

Contact Info

Product

Resources

About

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Pore loop-mutated rat KIR6.1 and KIR6.2 suppress K_ATPcurrent in rat cardiomyocytes

Sulfonylurea Receptors Type 1 and 2A Randomly Assemble to Form Heteromeric K_ATPChannels of Mixed Subunit Composition