Structural and Biochemical Insights into the Activation Mechanisms of Germinal Center Kinase OSR1

Li, Chuanchuan; Mao, Feng; Shi, Zhubing; Hao, Qian; Song, Xiaomin; Wang, Wenjia; Zhao, Yun; Shi, Jianzhong; Zhou, Zhaocai

doi:10.1074/jbc.m114.592097

Cited by 9 publications

(11 citation statements)

References 43 publications

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“…αAL helices occur in diverse classes of kinases, e.g., EGF receptor (51), and are known to be inhibitory by sterically blocking the inward movement of αC to form the K-E ion pair. Recent work has shown that deletion of the αAL helix in OSR1 leads to enhanced kinase activity (52). Interestingly, the positions of SPAK WT and OSR1 WT αAL helices are incompatible with the position of the SPAK T243D activation loop (Fig.…”

Section: Resultsmentioning

confidence: 99%

Domain-Swapping Switch Point in Ste20 Protein Kinase SPAK

et al. 2015

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The related protein kinases SPAK and OSR1 regulate ion homeostasis in part by phosphorylating cation cotransporter family members. The structure of the kinase domain of OSR1 was solved in the unphosphorylated inactive form, and like some other Ste20 kinases, exhibited a domain-swapped activation loop. To further probe the role of domain swapping in SPAK/OSR1, we have determined the crystal structures of SPAK 63–403 at 3.1 Å and SPAK 63–390 T243D at 2.5 Å resolutions. These structures encompass the kinase domain and different portions of the C-terminal tail, the longer without, and the shorter with an activating point mutation T243D. The structure of the T243D protein reveals significant conformational differences relative to unphosphorylated SPAK and OSR1, but also has some features of an inactive kinase. Both structures are domain-swapped dimers. Sequences involved in domain swapping were identified and mutated to create a SPAK monomeric mutant with kinase activity, indicating that monomeric forms are active. The monomeric mutant is activated by WNK1, but has reduced activity toward its substrate NKCC2, suggesting regulatory roles for domain swapping. The structure of the partially active SPAK T243D is consistent with a multi-stage activation process in which phosphorylation induces a SPAK conformation that requires further remodeling to build the active structure.

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Section: Resultsmentioning

confidence: 99%

Domain-Swapping Switch Point in Ste20 Protein Kinase SPAK

et al. 2015

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“…The scaffold protein Mo25/calcium binding protein 39 stimulates SPAK/OSR1 activity in vitro and seems to work synergistically with WNKs. 9 Amino acids important for SPAK/ OSR1 interactions with Mo25, including sites A-D Glu, Val, Tyr, and Trp/Glu/Trp in SPAK/OSR1 and Met260 in Mo25, are conserved in Fray and DmMo25, 9,36 and Fray and DmMo25 interact in Drosophila developmental processes. 37 We examined whether DmMo25 stimulates Fray activity by performing in vitro kinase assays using purified Drosophila proteins, DmMo25, Fray, and the N terminus of NKCC (encoded by Ncc69).…”

Section: Mo25 Is Required For Maximal Transepithelial Ion Transportmentioning

confidence: 99%

Intracellular Chloride and Scaffold Protein Mo25 Cooperatively Regulate Transepithelial Ion Transport through WNK Signaling in the Malpighian Tubule

Sun¹,

Wu²,

Jonusaite

et al. 2018

JASN

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With No Lysine kinase (WNK) signaling regulates mammalian renal epithelial ion transport to maintain electrolyte and BP homeostasis. Our previous studies showed a conserved role for WNK in the regulation of transepithelial ion transport in the Malpighian tubule. Using assays and transgenic lines, we examined two potential WNK regulators, chloride ion and the scaffold protein mouse protein 25 (Mo25), in the stimulation of transepithelial ion flux., autophosphorylation of purified WNK decreased as chloride concentration increased. In conditions in which tubule intracellular chloride concentration decreased from 30 to 15 mM as measured using a transgenic sensor, WNK activity acutely increased. WNK activity in tubules also increased or decreased when bath potassium concentration decreased or increased, respectively. However, a mutation that reduces chloride sensitivity of WNK failed to alter transepithelial ion transport in 30 mM chloride. We, therefore, examined a role for Mo25. In kinase assays, Mo25 enhanced the activity of the WNK downstream kinase Fray, the fly homolog of mammalian Ste20-related proline/alanine-rich kinase (SPAK), and oxidative stress-responsive 1 protein (OSR1). Knockdown of in the Malpighian tubule decreased transepithelial ion flux under stimulated but not basal conditions. Finally, whereas overexpression of wild-type , with or without, did not affect transepithelial ion transport, overexpressed with chloride-insensitive increased ion flux. Cooperative interactions between chloride and Mo25 regulate WNK signaling in a transporting renal epithelium.

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“…In this study, we were the rst to demonstrate that the common allele homozygotes of rs1384006 C > T of the OXSR1 gene were signi cantly associated with a higher exacerbation rate and the risk of FE in the nonsmoking asthmatics. OXSR1 has rarely been studied with regard to respiratory diseases, although it plays a role as a salt transportation, and cell volume control through ionic mechanisms [33,34] and ion transport by bronchial epithelial cells is essential for healthy airways. Imbalance of the transport system is closely related to the pathophysiology of asthma such as dysfunction of epithelial cells and smooth muscles.…”

Section: Discussionmentioning

confidence: 99%

“…OXSR1 is also involved in the regulation of immune responses by interacting with TNF receptor protein kinase C-θ (PKCθ), which is expressed by lymphoid tissues. [25,34] OXSR1 and WNK1 kinase, an upstream activator of OXSR1, are hardly detectable at basal activity, which may mean that WNK-OXSR1 signaling is regulated tightly in normal physiological conditions. [34] Interestingly, this genetic effect of rs1384006 C > T was not found in the smoker asthmatics.…”

Section: Discussionmentioning

confidence: 99%

“…[25,34] OXSR1 and WNK1 kinase, an upstream activator of OXSR1, are hardly detectable at basal activity, which may mean that WNK-OXSR1 signaling is regulated tightly in normal physiological conditions. [34] Interestingly, this genetic effect of rs1384006 C > T was not found in the smoker asthmatics. The reason for this nding could be explained by smoking itself being a strong inducer to exacerbate asthma.…”

Section: Discussionmentioning

confidence: 99%

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Association of Genetic Variants of Oxidative Stress Responsive Kinase 1 (OXSR1) with Asthma Exacerbations in Non-Smoking Asthmatics

Kim1

Chang

Lee

et al. 2021

Preprint

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Background: Asthma exacerbation threatens patient's life. Several genetic studies have been conducted to determine the risk factors for asthma exacerbation, but this information is still lacking. We aimed to determine whether genetic variants of Oxidative Stress Responsive Kinase 1(OXSR1), a gene with functions of salt transport, immune response, and oxidative stress, are associated with exacerbation of asthma.Methods: Clinical data were obtained from 1,454 asthmatics and single nucleotide polymorphisms (SNPs) of OXSR1 were genotyped. Genetic associations with annual exacerbation rate were analyzed depending on smoking status. Results: Eleven SNPs were selected using Asian data in the International HapMap database. The common allele of rs1384006 C>T of OXSR1 showed a significantly higher annual exacerbation rate than the rare allele in non-smoking asthmatics (CC vs. CT vs. TT: 0.43 ± 0.04 vs. 0.28 ± 0.03 vs. 0.31 ± 0.09, P=0.004, Pcorr=0.039). The frequent exacerbators had a significantly higher frequency of the common allele of rs1384006 C>T than did the infrequent exacerbators (74.4% vs. 55.2%, P=0.004, P corr=0.038). Conclusion: The common allele of rs1384006 C>T of OXSR1 was associated with the asthma exacerbation rate and a higher risk of being a frequent exacerbator, indicating that non-smoking asthmatics who carry common alleles may be vulnerable to asthma exacerbations.

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Structural and Biochemical Insights into the Activation Mechanisms of Germinal Center Kinase OSR1

Cited by 9 publications

References 43 publications

Domain-Swapping Switch Point in Ste20 Protein Kinase SPAK

Domain-Swapping Switch Point in Ste20 Protein Kinase SPAK

Intracellular Chloride and Scaffold Protein Mo25 Cooperatively Regulate Transepithelial Ion Transport through WNK Signaling in the Malpighian Tubule

Association of Genetic Variants of Oxidative Stress Responsive Kinase 1 (OXSR1) with Asthma Exacerbations in Non-Smoking Asthmatics

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