Structure of the human cation–chloride cotransporter NKCC1 determined by single-particle electron cryo-microscopy

Yang, Xiaoyong; Wang, Qinzhe; Cao, Erhu

doi:10.1038/s41467-020-14790-3

Cited by 57 publications

(103 citation statements)

References 70 publications

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“…The observation of cell-type specific action of these agents is in line with different reversal potentials in mature neuronal populations [115] and calls for comprehensive neuron subtype-specific phospho analysis. The recently reported structural data of CCCs [116][117][118][119] finally lay the foundation to analyze jointly the physiological role of phosphorylation and underlying structural changes to obtain an integrative and mechanistic view of the action of phosphorylation.…”

Section: Discussionmentioning

confidence: 99%

Staurosporine and NEM mainly impair WNK-SPAK/OSR1 mediated phosphorylation of KCC2 and NKCC1

et al. 2020

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The pivotal role of KCC2 and NKCC1 in development and maintenance of fast inhibitory neurotransmission and their implication in severe human diseases arouse interest in posttranscriptional regulatory mechanisms such as (de)phosphorylation. Staurosporine (broad kinase inhibitor) and N-ethylmalemide (NEM) that modulate kinase and phosphatase activities enhance KCC2 and decrease NKCC1 activity. Here, we investigated the regulatory mechanism for this reciprocal regulation by mass spectrometry and immunoblot analyses using phospho-specific antibodies. Our analyses revealed that application of staurosporine or NEM dephosphorylates Thr 1007 of KCC2, and Thr 203 , Thr 207 and Thr 212 of NKCC1. Dephosphorylation of Thr 1007 of KCC2, and Thr 207 and Thr 212 of NKCC1 were previously demonstrated to activate KCC2 and to inactivate NKCC1. In addition, application of the two agents resulted in dephosphorylation of the T-loop and S-loop phosphorylation sites Thr 233 and Ser 373 of SPAK, a critical kinase in the WNK-SPAK/OSR1 signaling module mediating phosphorylation of KCC2 and NKCC1. Taken together, these results suggest that reciprocal regulation of KCC2 and NKCC1 via staurosporine and NEM is based on WNK-SPAK/OSR1 signaling. The key regulatory phospho-site Ser 940 of KCC2 is not critically involved in the enhanced activation of KCC2 upon staurosporine and NEM treatment, as both agents have opposite effects on its phosphorylation status. Finally, NEM acts in a tissue-specific manner on Ser 940 , as shown by comparative analysis in HEK293 cells and immature cultured hippocampal neurons. In summary, our analyses identified phospho-sites that are responsive to staurosporine or NEM application. This provides important information towards a better understanding of the cooperative interactions of different phospho-sites.

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Section: Discussionmentioning

confidence: 99%

Staurosporine and NEM mainly impair WNK-SPAK/OSR1 mediated phosphorylation of KCC2 and NKCC1

et al. 2020

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“…21,22 SLC12A2 encodes for the protein NKCC1 which regulates transport of chloride, potassium and sodium across cell membranes, and is key in modulating ion movement across the epithelium, chloride homeostasis, volume of cells, macrophage activation, and anti-microbial activity. 23,24 Through access to published gene expression studies we observed that across several independent studies, the minor allele (T) of the lead SNP in LINC01184/SLC12A2, rs3749748, was particularly associated with reduced gene expression of SLC12A2 and LINC01184 in monocytes and macrophages. Macrophages are the most abundant resident immune cell type in the skin, and are critical for the resistance against invading microbes and tissue repair.…”

Section: Discussionmentioning

confidence: 99%

“…Many of these traits are linked to cell-volume regulation, which is a key role of NKCC1. 23 While it is unclear whether any of these traits may themselves be causally linked to SSTI susceptibility, cell-volume regulation may be. By changing the intracellular volume of cells, mechanical barrier function may be regulated and affect immune cell and bacterial migration.…”

Section: Discussionmentioning

confidence: 99%

Genome-wide association study of skin and soft tissue infection susceptibility

Rogne¹,

Liyanarachi²,

Rasheed³

et al. 2020

Preprint

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Background: Skin and soft tissue infections (SSTIs) are common worldwide, but little is known about the genetic susceptibility and the causal effect of cardiometabolic risk factors. We therefore conducted the first genome-wide association study (GWAS) of SSTIs, with downstream analyses including Mendelian randomization analyses. Methods: The GWAS was conducted using the UK Biobank as discovery cohort, with 6,107 cases and 399,239 controls, and the Trondelag Health Study (HUNT) as replication cohort with 1,657 cases and 67,522 controls. Cases and controls were those who had or had not been hospitalized with an SSTI diagnosis, respectively. Findings: One locus, lead single-nucleotide polymorphism rs3749748 in LINC01184/SLC12A2, was associated with SSTIs in the UK Biobank (odds ratio [OR] 1.19, p-value = 7.6e-16) and replicated in HUNT (OR 1.15, p-value = 6.3e-4). Meta-analysis confirmed the lead variant (OR 1.18, p-value = 4.4e-18), as well as suggested two additional loci close to genome-wide significance (rs2007361 in PSMA1, OR 0.91, p-value = 5.1e-8; and rs78625038 in GAN, OR 1.86, p-value = 5.9e-8). Gene-based association tests identified four genes linked to SSTIs: SLC12A2, PSMA1, GAN, and IL6R. The minor allele of rs3749748 reduced the gene expression of SLC12A2 primarily in monocytes and macrophages. Mendelian randomization analyses showed that increasing body mass index and lifetime smoking habits increased risk of SSTIs. Interpretation: LINC0118/SLC12A2 was robustly associated with SSTI incidence and may exert its effect through reduced gene expression in monocytes and macrophages. Reducing tobacco smoking, overweight and obesity in the population may reduce the incidence of SSTIs.

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“…The number and position of phospho-sites vary between different family members as they are not evolutionary conserved among isoforms and paralogs [23]. Phosphatases such as protein phosphatase 1 (PP1) counterbalance the act of the kinases [28].…”

Section: Functional Regulation Of the Cation Chloride Cotransporter Fmentioning

confidence: 99%

Role of the Cation-chloride-cotransporters in Cardiovascular Disease

Azlan

Zhang

2020

Preprint

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The SLC12 family of cation-chloride-cotransporters (CCCs), comprising potassium chloride cotransporters (KCCs)-mediated Cl- extrusion relative to sodium chloride cotransporters (NKCCs)-mediated Cl- loading, play vital roles in cell volume regulation and ion homeostasis. These functions of the CCCs influence a variety of physiological processes, many of which overlap with the pathophysiology of cardiovascular disease. Although not all of the cotransporters have been linked to Mendelian genetic disorders, recent studies have provided new insights into their functional role in vascular and renal cells along with their contribution to cardiovascular diseases. Particularly, an imbalance in potassium levels promote the pathogenesis of atherosclerosis and disturbances in sodium homeostasis are one of the causes of hypertension. Recent findings even suggest hypothalamic signalling as a key signalling pathway in the pathophysiology of hypertension. In this review, we summarize and discuss the role of CCCs in cardiovascular disease with particular emphasis on knowledge gained in recent years on NKCCs and KCCs.

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Structure of the human cation–chloride cotransporter NKCC1 determined by single-particle electron cryo-microscopy

Cited by 57 publications

References 70 publications

Staurosporine and NEM mainly impair WNK-SPAK/OSR1 mediated phosphorylation of KCC2 and NKCC1

Staurosporine and NEM mainly impair WNK-SPAK/OSR1 mediated phosphorylation of KCC2 and NKCC1

Genome-wide association study of skin and soft tissue infection susceptibility

Role of the Cation-chloride-cotransporters in Cardiovascular Disease

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