Syntaxin-1A inhibits KATP channels by interacting with specific conserved motifs within sulfonylurea receptor 2A |
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Authors: | Chao Christin Liang Tao Kang Youhou Lin Xianguang Xie Huanli Feng Zhong-Ping Gaisano Herbert Y |
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Affiliation: | aDepartment of Medicine, University of Toronto, Room 7368, Medical Sciences Building, Toronto, ON, Canada M5S 1A8;bDepartment of Physiology, University of Toronto, Room 3306, Medical Sciences Building, Toronto, ON, Canada M5S 1A8 |
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Abstract: | We previously demonstrated that syntaxin (Syn)-1A is present in the sarcolemma of rat cardiomyocytes and binds sulfonylurea receptor (SUR) 2A nucleotide binding folds (NBFs) to inhibit ATP-sensitive potassium (KATP) channel. Here, we examined for the precise domains within the NBFs of SUR2A that may interact with Syn-1A. Specifically, we tested truncated NBF protein segments encompassing the conserved motifs Walker A (WA), signature/Linker (L), and Walker B (WB). In vitro binding results indicate that the domains encompassing WA and L of NBF-1 and all three conserved motifs of NBF-2 bound Syn-1A. Electrophysiological studies, employing inside-out patch-clamp recordings from SUR2A/Kir6.2 expressing HEK cells and mouse cardiomyocytes, show that WB and L of NBF-1 and all three NBF-2 truncated protein segments reduced Syn-1A inhibition of SUR2A/KATP channels. Remarkably, these same NBF-1 and -2 truncated proteins could independently disrupt the intimate FRET interactions of full length SUR2A (− mCherry) and Syn-1A (− EGFP). These results taken together indicate that Syn-1A possibly maintains inhibition of cardiac ventricular KATP channels by binding to large regions of NBF-1 and NBF-2 to stabilize the NBF-1-NBF-2 heterodimer formation and prevent ATP-binding and ATP hydrolysis. Since KATP channels are closely coupled to metabolic states, we postulate that these very intimate Syn-1A-SUR2A interactions are critically important for myocardial protection during stress, in which profound changes in metabolic factors (pH, ATP) could modulate these Syn-1A-SUR2A interactions. |
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Keywords: | Syntaxin-1A SNARE ATP-sensitive potassium channel SUR2A Kir6.2 |
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