Zhao Yongxiang, Vidossich Pietro, Forbush Biff, Ma Junfeng, Rinehart Jesse, De Vivo Marco, Cao Erhu
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, UT, 84112-5650, USA.
Key Laboratory of Magnetic Resonance in Biological Systems, State Key Laboratory of Magnetic Resonance and Atomic and Molecular Physics, National Center for Magnetic Resonance in Wuhan, Wuhan Institute of Physics and Mathematics, Innovation Academy for Precision Measurement Science and Technology, Chinese Academy of Sciences, 430071, Wuhan, P. R. China.
EMBO J. 2025 Mar;44(5):1540-1562. doi: 10.1038/s44318-025-00368-6. Epub 2025 Jan 28.
Na-K-Cl cotransporters functions as an anion importers, regulating trans-epithelial chloride secretion, cell volume, and renal salt reabsorption. Loop diuretics, including furosemide, bumetanide, and torsemide, antagonize both NKCC1 and NKCC2, and are first-line medicines for the treatment of edema and hypertension. NKCC1 activation by the molecular crowding sensing WNK kinases is critical if cells are to combat shrinkage during hypertonic stress; however, how phosphorylation accelerates NKCC1 ion transport remains unclear. Here, we present co-structures of phospho-activated NKCC1 bound with furosemide, bumetanide, or torsemide showing that furosemide and bumetanide utilize a carboxyl group to coordinate and co-occlude a K, whereas torsemide encroaches and expels the K from the site. We also found that an amino-terminal segment of NKCC1, once phosphorylated, interacts with the carboxyl-terminal domain, and together, they engage with intracellular ion exit and appear to be poised to facilitate rapid ion translocation. Together, these findings enhance our understanding of NKCC-mediated epithelial ion transport and the molecular mechanisms of its inhibition by loop diuretics.
钠-钾-氯共转运体作为阴离子导入蛋白发挥作用，调节跨上皮氯化物分泌、细胞体积和肾脏盐重吸收。包括呋塞米、布美他尼和托拉塞米在内的袢利尿剂可拮抗NKCC1和NKCC2，是治疗水肿和高血压的一线药物。如果细胞要在高渗应激期间对抗细胞萎缩，由分子拥挤感应WNK激酶激活NKCC1至关重要；然而，磷酸化如何加速NKCC1离子转运仍不清楚。在这里，我们展示了与呋塞米、布美他尼或托拉塞米结合的磷酸化激活型NKCC1的共结构，表明呋塞米和布美他尼利用一个羧基来配位并共同封闭一个钾离子，而托拉塞米侵入并将钾离子从该位点排出。我们还发现，NKCC1的氨基末端片段一旦磷酸化，就会与羧基末端结构域相互作用，并且它们共同与细胞内离子出口结合，似乎准备好促进快速离子转运。总之，这些发现加深了我们对NKCC介导的上皮离子转运及其被袢利尿剂抑制的分子机制的理解。
