Department of Physiology and Neurobiology, University of Connecticut, 75 N Eagleville Rd Unit 3156, Storrs-Mansfield, CT 06269-9011, USA.
J Physiol. 2009 Dec 15;587(Pt 24):5819-30. doi: 10.1113/jphysiol.2009.180372.
AMP-activated protein kinase (AMPK) is a serine/threonine kinase activated by conditions that increase the AMP : ATP ratio. In carotid body glomus cells, AMPK is thought to link changes in arterial O(2) with activation of glomus cells by inhibition of unidentified background K(+) channels. Modulation by AMPK of individual background K(+) channels has not been described. Here, we characterize effects of activated AMPK on recombinant TASK-1, TASK-3, TREK-1 and TREK-2 background K(+) channels expressed in HEK293 cells. We found that TREK-1 and TREK-2 channels but not TASK-1 or TASK-3 channels are inhibited by AMPK. AMPK-mediated inhibition of TREK involves key serine residues in the C-terminus that are also known to be important for PKA and PKC channel modulation; inhibition of TREK-1 requires Ser-300 and Ser-333 and inhibition of TREK-2 requires Ser-326 and Ser-359. Metabolic inhibition by sodium azide can also inhibit both TREK and TASK channels. The effects of azide on TREK occlude subsequent channel inhibition by AMPK and are attenuated by expression of a dominant negative catalytic subunit of AMPK (dnAMPK), suggesting that metabolic stress modulates TREK channels by an AMPK mechanism. By contrast, inhibition of TASK channels by azide was unaffected by expression of dnAMPK, suggesting an AMPK-independent mechanism. In addition, prolonged exposure (6-7 min) to hypoxia ( = 11 +/- 1 mmHg) inhibits TREK channels and this response was blocked by expression of dnAMPK. Our results identify a novel modulation of TREK channels by AMPK and indicate that select residues in the C-terminus of TREK are points of convergence for multiple signalling cascades including AMPK, PKA and PKC. To the extent that carotid body O(2) sensitivity is dependent on AMPK, our finding that TREK-1 and TREK-2 channels are inhibited by AMPK suggests that TREK channels may represent the AMPK-inhibited background K(+) channels that mediate activation of glomus cells by hypoxia.
AMP 激活的蛋白激酶 (AMPK) 是一种丝氨酸/苏氨酸激酶，可被增加 AMP:ATP 比值的条件激活。在颈动脉体球细胞中，AMPK 被认为通过抑制未鉴定的背景 K(+)通道将动脉 O(2)的变化与球细胞的激活联系起来。AMPK 对单个背景 K(+)通道的调制尚未被描述。在这里，我们描述了激活的 AMPK 对在 HEK293 细胞中表达的重组 TASK-1、TASK-3、TREK-1 和 TREK-2 背景 K(+)通道的影响。我们发现 TREK-1 和 TREK-2 通道但不是 TASK-1 或 TASK-3 通道被 AMPK 抑制。AMPK 介导的 TREK 抑制涉及 C 末端的关键丝氨酸残基，这些残基也被认为对 PKA 和 PKC 通道调节很重要；TREK-1 的抑制需要 Ser-300 和 Ser-333，而 TREK-2 的抑制需要 Ser-326 和 Ser-359。NaN3 引起的代谢抑制也可以抑制 TREK 和 TASK 通道。NaN3 对 TREK 的作用阻断了 AMPK 对通道的后续抑制，并且被 AMPK 的显性负催化亚基 (dnAMPK) 的表达减弱，表明代谢应激通过 AMPK 机制调节 TREK 通道。相比之下，NaN3 对 TASK 通道的抑制作用不受 dnAMPK 表达的影响，表明存在 AMPK 独立的机制。此外，长时间（6-7 分钟）暴露于低氧（=11 +/- 1 mmHg）抑制 TREK 通道，并且这种反应被 dnAMPK 的表达阻断。我们的结果确定了 AMPK 对 TREK 通道的一种新的调节作用，并表明 TREK 中的几个 C 末端残基是包括 AMPK、PKA 和 PKC 在内的多个信号级联的汇聚点。在一定程度上，颈动脉体 O(2)敏感性依赖于 AMPK，我们发现 TREK-1 和 TREK-2 通道被 AMPK 抑制表明 TREK 通道可能代表 AMPK 抑制的背景 K(+) 通道，介导缺氧对球细胞的激活。
