Affinage

STK39

STE20/SPS1-related proline-alanine-rich protein kinase · UniProt Q9UEW8

Length
545 aa
Mass
59.5 kDa
Annotated
2026-04-28
100 papers in source corpus 39 papers cited in narrative 39 extracted findings

Mechanistic narrative

Synthesis pass · prose summary of the discoveries below

STK39 (SPAK) is a STE20-family serine/threonine kinase that functions as the central effector of the WNK signaling cascade to regulate ion homeostasis in kidney, vasculature, and brain. WNK kinases phosphorylate SPAK at T-loop residue T243 and regulatory S383, and the scaffold MO25 further amplifies SPAK activity ~100-fold; activated SPAK then directly phosphorylates N-terminal threonines on Na⁺-coupled cation-chloride cotransporters NKCC1, NKCC2, and NCC to stimulate their activity, while simultaneously phosphorylating a conserved C-terminal threonine on KCC family members to inhibit them (PMID:16263722, PMID:21423148, PMID:24393035). Substrate recognition and upstream docking both require the conserved C-terminal CCT domain, which binds RFXV/I motifs on WNKs and cotransporters with nanomolar affinity (PMID:16669787, PMID:25994507). SPAK knockout or WNK-binding-deficient knock-in mice develop a Gitelman-like syndrome with reduced blood pressure, establishing SPAK as the primary in vivo regulator of renal salt reabsorption and vascular tone, with additional roles in epithelial HCO₃⁻ secretion via CFTR/NBCe1-B phosphorylation, T-cell AP-1 signaling via PKCθ, and cancer cell invasion via SNAI1 phosphorylation (PMID:20813865, PMID:20091762, PMID:21317537, PMID:14988727, PMID:34335956).

Mechanistic history

Synthesis pass · year-by-year structured walk · 14 steps
  1. 2000 Medium

    Establishing that SPAK is an active kinase capable of autophosphorylation and exogenous substrate phosphorylation resolved the basic question of whether STK39 encodes a functional kinase, and its activation of p38 MAPK placed it in a stress-signaling context.

    Evidence In vitro kinase assay and cotransfection reporter assays in mammalian cells

    PMID:10980603

    Open questions at the time
    • Physiological substrates unidentified
    • p38 activation mechanism unclear (direct vs. indirect)
    • Upstream activator unknown
  2. 2002 High

    Discovery that SPAK directly binds cation-chloride cotransporters NKCC1, NKCC2, and KCC3 via a C-terminal domain recognizing RFXV/I motifs identified the first physiological substrate class and the docking mechanism, fundamentally redirecting the field from stress-kinase signaling to ion transport regulation.

    Evidence Yeast two-hybrid, GST pulldown, co-immunoprecipitation from mouse brain

    PMID:12386165

    Open questions at the time
    • Whether SPAK phosphorylates these cotransporters was not yet shown
    • Identity of upstream activating kinase unknown
  3. 2004 High

    Identification of PKCθ as an upstream kinase phosphorylating SPAK at S311 and the demonstration that SPAK activates AP-1 in T cells established a non-ion-transport signaling role and the first upstream phosphorylation event on SPAK.

    Evidence Co-immunoprecipitation, in vitro kinase assay, RNAi, reporter assay in T cells

    PMID:14988727

    Open questions at the time
    • Whether this PKCθ-SPAK axis operates on cotransporters unknown
    • Relationship to WNK signaling unclear
  4. 2005 High

    The pivotal discovery that WNK1 phosphorylates SPAK and that SPAK/OSR1 in turn directly phosphorylate the N-terminal regulatory regions of NKCC1, NKCC2, and NCC established the WNK→SPAK→CCC phosphorylation cascade as a coherent signaling pathway governing cation-chloride cotransport.

    Evidence In vitro kinase assay with site-directed mutagenesis, cell-based phosphorylation assays

    PMID:16263722

    Open questions at the time
    • Specific phosphosites on cotransporters not yet mapped
    • In vivo relevance unconfirmed
  5. 2006 High

    Detailed characterization of the CCT domain's nanomolar-affinity RFXV-motif binding, identification of activation-loop residues T243/T247 as essential for catalysis, and mapping of NKCC1 phosphorylation sites (T203/T207/T212) provided the structural and biochemical framework for SPAK substrate recognition and activation.

    Evidence Surface plasmon resonance, X-ray-guided mutagenesis, 86Rb⁺ uptake in Xenopus oocytes, in vitro kinase assays

    PMID:16382158 PMID:16669787

    Open questions at the time
    • Full crystal structure of CCT-RFXV complex not yet solved
    • Regulation of SPAK by scaffolds other than WNK unknown
  6. 2008 High

    Extension of the WNK-SPAK cascade to NCC (mapping phosphosites T46/T55/T60) and demonstration that PKCδ also activates SPAK during osmotic stress broadened the upstream inputs and downstream substrates of the pathway beyond NKCC1.

    Evidence In vitro kinase assay with phospho-specific antibodies in kidney cells; siRNA and 86Rb⁺ flux in airway epithelia

    PMID:18270262 PMID:18550547

    Open questions at the time
    • In vivo significance of PKCδ-SPAK axis unconfirmed
    • Relative contributions of SPAK vs. OSR1 to NCC regulation unknown
  7. 2010 High

    SPAK knockout and WNK-binding-deficient knock-in mice provided definitive in vivo proof that SPAK is the essential kinase for NCC phosphorylation, regulates NKCC1-dependent vascular tone, and controls blood pressure, producing a Gitelman-like phenotype with hypotension.

    Evidence SPAK-null and knock-in mouse models with phospho-Western, blood pressure, and vascular contractility assays

    PMID:20091762 PMID:20813865

    Open questions at the time
    • Compensatory OSR1 effects not fully dissected
    • Tissue-specific isoform contributions unclear
  8. 2011 High

    Discovery that MO25 activates SPAK ~100-fold and that a kidney-specific truncated SPAK isoform acts as a dominant-negative inhibitor explained how SPAK activity is fine-tuned and why SPAK knockout has divergent effects on NCC vs. NKCC2 phosphorylation along the nephron.

    Evidence In vitro kinase assay with MO25, siRNA, SPAK knockout mouse with isoform-specific analysis

    PMID:21423148 PMID:21907141

    Open questions at the time
    • How MO25 binding is regulated in vivo unknown
    • Whether truncated isoform has independent functions unclear
  9. 2012 High

    Demonstration that SPAK/OSR1 phosphorylate KCC family members at a conserved C-terminal threonine to inhibit them established that SPAK reciprocally regulates Na⁺-importing (NKCC/NCC activated) and K⁺-exporting (KCC inhibited) cotransporters, unifying chloride homeostasis under one kinase.

    Evidence In vitro kinase assay, SPAK/OSR1 double-knockin ES cells, 86Rb⁺ uptake

    PMID:24393035

    Open questions at the time
    • In vivo KCC phosphorylation by SPAK not yet confirmed in genetic models
    • Contribution to neuronal Cl⁻ homeostasis inferred but not directly tested
  10. 2012 High

    Genetic epistasis in PI3K/Akt-hyperactive db/db mice showed that insulin signaling activates the WNK-SPAK-NCC cascade, and SPAK-inactivating knock-in completely rescued hypertension, linking metabolic syndrome to SPAK-dependent salt retention.

    Evidence SpakT243A/+ knock-in crossed with db/db mice, PI3K inhibitor, phospho-Western

    PMID:22949526

    Open questions at the time
    • Direct phosphorylation link between Akt and WNK not established
    • Translation to human metabolic hypertension unconfirmed
  11. 2015 High

    Crystal structures of the SPAK kinase domain revealed domain-swapped dimers that modulate substrate access rather than intrinsic catalysis, and CCT domain L502A knock-in mice confirmed that a single docking residue is essential for all RFXV-dependent interactions in vivo.

    Evidence X-ray crystallography at 2.5–3.1 Å resolution; L502A knock-in mice with Co-IP, phospho-Western, blood pressure

    PMID:25994507 PMID:26208601

    Open questions at the time
    • Full-length SPAK structure not available
    • Whether dimerization occurs in cells undemonstrated
  12. 2017 High

    DCT-specific constitutively active SPAK knock-in mice developed thiazide-sensitive hypertension with hyperkalemia and ASDN remodeling, proving that SPAK activation in a single nephron segment is sufficient to drive systemic blood pressure elevation and revealing DCT-ASDN structural coupling.

    Evidence Conditional CA-SPAK knock-in with DCT-specific Cre, immunofluorescence, thiazide challenge

    PMID:28442491

    Open questions at the time
    • Molecular signals mediating ASDN remodeling downstream of SPAK unknown
    • Whether CA-SPAK also alters KCC phosphorylation in vivo not tested
  13. 2020 High

    The SPAK inhibitor ZT-1a demonstrated that pharmacological SPAK inhibition simultaneously reduces NKCC1 phosphorylation and relieves KCC inhibition in vivo, attenuating cerebral edema and CSF hypersecretion, validating SPAK as a druggable target for neurological fluid disorders.

    Evidence In vitro kinase assay, rodent stroke and hydrocephalus models with CCC phosphorylation readouts

    PMID:31911626

    Open questions at the time
    • Off-target effects of ZT-1a not fully profiled
    • Long-term safety and specificity in vivo unknown
  14. 2021 Medium

    Identification of SNAI1 as a SPAK substrate phosphorylated at T203, promoting SNAI1 nuclear retention and EMT in breast cancer, extended SPAK's substrate repertoire beyond ion transport to transcription factor regulation and cancer metastasis.

    Evidence Co-immunoprecipitation, in vitro kinase assay, knockdown/overexpression, xenograft model

    PMID:34335956

    Open questions at the time
    • Whether SNAI1 phosphorylation depends on the CCT-RFXV docking mechanism is untested
    • Relevance to other cancer types unknown
    • Independent replication needed

Open questions

Synthesis pass · forward-looking unresolved questions
  • Key unresolved questions include the full-length SPAK structure with CCT domain and MO25, the molecular basis of SPAK dimerization in cells, the relative tissue-specific contributions of SPAK vs. OSR1, the mechanism by which SPAK regulates cell cycle and apoptosis independently of ion transport, and whether SPAK inhibitors can be developed for safe clinical use in hypertension and neurological disorders.
  • No full-length SPAK structure available
  • SPAK vs. OSR1 tissue-specific redundancy not genetically resolved in all organs
  • Cancer and cell-cycle roles lack mechanistic depth

Mechanism profile

Synthesis pass · controlled-vocabulary classification · explore literature graph →
Molecular activity
GO:0140096 catalytic activity, acting on a protein 10 GO:0098772 molecular function regulator activity 2
Localization
GO:0005829 cytosol 2 GO:0005886 plasma membrane 2 GO:0005634 nucleus 1
Pathway
R-HSA-382551 Transport of small molecules 8 R-HSA-162582 Signal Transduction 4 R-HSA-1643685 Disease 2 R-HSA-168256 Immune System 1
Partners
CAB39OXSR1PRKCQSLC12A1SLC12A2SLC12A3WNK1WNK4

Evidence

Reading pass · 39 per-paper findings extracted from the source corpus
Year Finding Method Journal Conf PMIDs
2000 SPAK (STK39) is a serine/threonine kinase that can autophosphorylate and phosphorylate exogenous substrates in vitro; it specifically activates the p38 MAPK pathway in cotransfection assays; full-length SPAK localizes to the cytoplasm while a caspase-cleaved form localizes predominantly to the nucleus. In vitro kinase assay, cotransfection/reporter assay, subcellular localization by immunofluorescence Oncogene Medium 10980603
2002 SPAK and OSR1 directly interact with cation-chloride cotransporters KCC3, NKCC1, and NKCC2 (but not KCC1 or KCC4) via a conserved C-terminal domain on the kinases that recognizes an (R/K)FX(V/I) binding motif on the cotransporters; co-immunoprecipitation confirmed the SPAK-NKCC1 interaction in mouse brain. Yeast two-hybrid, GST pulldown, co-immunoprecipitation, immunohistochemistry The Journal of biological chemistry High 12386165
2003 SPAK acts as a scaffolding protein at NKCC1: preventing SPAK binding to NKCC1 does not affect basal cotransporter function, but SPAK co-immunoprecipitates with p38 MAPK and NKCC1 in an activity-dependent manner, with p38 dissociating from the complex upon cellular stress while SPAK-NKCC1 interaction remains stable. 86Rb+ uptake assay in Xenopus oocytes, co-immunoprecipitation, Western blot The Journal of biological chemistry Medium 14563843
2004 PKCθ phosphorylates SPAK on Ser-311 in its kinase domain; SPAK interacts with PKCθ (but not PKCα) via its C-terminal 99 residues; TCR/CD28 costimulation enhances this association and SPAK kinase activity; SPAK synergizes with constitutively active PKCθ to activate AP-1 but not NF-κB in T cells. Co-immunoprecipitation, in vitro kinase assay, RNAi knockdown, reporter assay The EMBO journal High 14988727
2005 WNK1 phosphorylates the evolutionary conserved serine residue located outside the kinase domain of SPAK (and OSR1), and this phosphorylation influences SPAK/OSR1 kinase activity; SPAK and OSR1 directly phosphorylate the N-terminal regulatory regions of cation-chloride cotransporters NKCC1, NKCC2, and NCC; hypotonic stress activates SPAK/OSR1 and induces cotransporter phosphorylation. In vitro kinase assay, cell-based phosphorylation assay, site-directed mutagenesis The Journal of biological chemistry High 16263722
2006 SPAK activation loop residues T243 and T247 are required for kinase activity; mutation of T243A or T247A produces a dominant-negative effect on NKCC1 activity in Xenopus oocytes; OSR1 has similar kinase properties and activates NKCC1 when coexpressed with WNK4. Site-directed mutagenesis, 32P-ATP in vitro phosphorylation, 86Rb+ uptake in Xenopus oocytes Molecular and cellular biology High 16382158
2006 SPAK and OSR1 possess a conserved C-terminal CCT domain that interacts with nanomolar affinity with RFXV motifs in substrates (NKCC1) and upstream activators (WNK1/WNK4); specific residues within the CCT domain are required for RFXV binding; an intact CCT domain is required for WNK1 to efficiently phosphorylate and activate OSR1; SPAK/OSR1 phosphorylate NKCC1 at Thr203/Thr207/Thr212 (human) identified by in vitro assay. In vitro kinase assay, surface plasmon resonance, CCT domain mutagenesis, peptide competition The Biochemical journal High 16669787
2006 RELT (TNF receptor) binds SPAK via an RFRV motif and uses SPAK to mediate p38 and JNK activation; disruption of the SPAK binding motif in RELT or use of kinase-dead SPAK inhibits RELT-induced p38/JNK activation. Yeast two-hybrid, co-immunoprecipitation, reporter/kinase activity assay Biochemical and biophysical research communications Medium 16530727
2007 Physical docking of SPAK to NKCC1 via a single RFXV binding motif is necessary for cotransporter activation; mutation of the phenylalanine in the motif abolishes binding and activation; SPAK phosphorylates NKCC1 at T206 and T211 as major regulatory sites. Yeast two-hybrid, 32P-ATP in vitro phosphorylation, 86Rb+ uptake in Xenopus oocytes, site-directed mutagenesis Cellular physiology and biochemistry High 17595523
2007 AATYK1 scaffolds protein phosphatase 1 (PP1) via a PP1 docking motif and binds SPAK via RFXV motifs; AATYK1 inhibits NKCC1 activity by bringing PP1 into proximity with SPAK, thereby indirectly opposing SPAK/WNK4 activation of the cotransporter. Yeast two-hybrid, 86Rb+ uptake in Xenopus oocytes, site-directed mutagenesis American journal of physiology. Cell physiology Medium 17267545
2008 SPAK and OSR1 activated by WNK1 phosphorylate human NCC at Thr46, Thr55, and Thr60; efficient NCC phosphorylation requires a docking interaction between an RFXI motif in NCC and SPAK/OSR1; hypotonic low-chloride conditions activate the WNK1-SPAK/OSR1 pathway to phosphorylate NCC in kidney cells. In vitro kinase assay, phospho-specific antibodies, cell-based assay in HEK293 and mpkDCT cells, mutagenesis Journal of cell science High 18270262
2008 PKCδ acts upstream of SPAK to activate NKCC1 during hyperosmotic stress in airway epithelial cells; PKCδ directly binds SPAK and phosphorylates it to increase SPAK kinase activity; SPAK binds the amino terminus of NKCC1 directly and SPAK knockdown prevents NKCC1 phosphorylation and activation. siRNA knockdown, recombinant protein binding assay, in vitro kinase assay, 86Rb+ flux assay, co-immunoprecipitation The Journal of biological chemistry High 18550547
2009 STK39/SPAK interacts with WNK kinases and cation-chloride cotransporters in vivo; STK39 is expressed in the distal nephron where it may regulate renal Na+ excretion; an intronic conserved element shows allele-specific transcriptional activity, suggesting BP-associated variants increase STK39 expression. Cell-based co-immunoprecipitation, in vivo expression/localization, in vitro transcription reporter assay Proceedings of the National Academy of Sciences of the United States of America Medium 19114657
2009 AngII signaling increases NCC activity via a WNK4-SPAK-dependent pathway in Xenopus oocytes and mammalian cells; dominant-negative SPAK or elimination of the SPAK binding motif in NCC prevents AngII-mediated NCC activation; AngII increases phosphorylation of specific activation sites on SPAK and NCC. Xenopus oocyte expression, dominant-negative SPAK, phospho-specific antibodies in mpkDCT cells Proceedings of the National Academy of Sciences of the United States of America High 19240212
2009 Epigenetic silencing of STK39 in B-cell lymphoma inhibits caspase-3-dependent apoptosis from DNA double-strand breaks; SPAK knockdown by shRNA protects B cells from genotoxic stress-induced apoptosis but not osmotic/oxidative stress; c-Jun N-terminal kinase (JNK) is a potential downstream mediator of SPAK in this pathway. shRNA knockdown, apoptosis/caspase-3 assay, pharmacological JNK inhibition, DNA methylation analysis The American journal of pathology Medium 19717643
2010 SPAK knockout mice exhibit hypotension with Gitelman syndrome phenotype (hypokalemia, hypomagnesemia, hypocalciuria); NCC phosphorylation and expression are markedly reduced while NKCC2 phosphorylation is increased; NKCC1 phosphorylation in aortic tissue is decreased causing impaired vasoconstriction, establishing SPAK as an in vivo regulator of both renal NCC and vascular NKCC1. SPAK-null mouse model, phospho-specific Western blot, diuretic challenge, vascular contractility assay Journal of the American Society of Nephrology : JASN High 20813865
2010 SPAK knock-in mice in which SPAK cannot be activated by WNK kinases display markedly reduced phosphorylation and expression of NCC and NKCC2 cotransporters and significantly reduced blood pressure, establishing the WNK-SPAK axis as the key in vivo regulatory pathway for these transporters. Knock-in mouse model (WNK-binding site mutation), phospho-specific Western blot, blood pressure measurement EMBO molecular medicine High 20091762
2010 SPAK requires phosphorylation at T243 (catalytic domain) and S383 (regulatory domain) by WNK kinases for activation; mutating S383 to alanine or surrounding residues paradoxically renders SPAK constitutively active; a second catalytic-domain serine S321 can also be phosphorylated by WNK4; SPAK substrate recognition requires two threonines separated by four amino acids with a hydrophobic residue after the first. Site-directed mutagenesis, 86Rb+ uptake in Xenopus oocytes, in vitro kinase assay American journal of physiology. Cell physiology High 20463172
2010 PP1 dephosphorylates both SPAK and the N-terminal tail of NKCC1 directly; the PP1 binding motif on NKCC1 facilitates scaffolding of PP1 near SPAK, greatly enhancing PP1-mediated dephosphorylation of SPAK. In vitro dephosphorylation assay with recombinant proteins, 86Rb+ uptake in Xenopus oocytes, mutagenesis The Journal of biological chemistry Medium 20223824
2010 SORLA intracellular sorting receptor functionally interacts with SPAK and controls SPAK intracellular trafficking; SORLA deficiency results in missorting of SPAK and consequent failure to phosphorylate NKCC2 in the thick ascending limb. Co-immunoprecipitation, SORLA knockout mouse model, phospho-specific Western blot, immunofluorescence Molecular and cellular biology Medium 20385770
2011 MO25α/β bind SPAK/OSR1 and induce ~100-fold activation of their kinase activity, dramatically enhancing phosphorylation of ion cotransporters NKCC1, NKCC2, and NCC; siRNA-mediated MO25 reduction inhibits endogenous NKCC1 phosphorylation at SPAK/OSR1 sites. In vitro kinase assay, siRNA knockdown, phospho-specific Western blot, surface plasmon resonance The EMBO journal High 21423148
2011 WNK kinases act as scaffolds to recruit SPAK, which phosphorylates CFTR and NBCe1-B to reduce their cell surface expression in pancreatic ductal epithelium; IRBIT opposes WNK/SPAK effects by recruiting PP1 to dephosphorylate CFTR and NBCe1-B; silencing SPAK increases ductal secretion. siRNA knockdown in mouse pancreatic ducts, cell surface biotinylation, phosphorylation assay, rescue experiments The Journal of clinical investigation High 21317537
2011 A kidney-specific truncated SPAK isoform lacking the kinase domain inhibits full-length SPAK-mediated phosphorylation of NCC and NKCC2 in vitro; SPAK knockout has divergent effects along TAL (increased pNKCC2) and DCT (decreased pNCC), explained by differential isoform expression along the nephron. In vitro kinase assay, SPAK knockout mouse model, isoform-specific Western blot and immunofluorescence Cell metabolism High 21907141
2012 SPAK and OSR1 directly phosphorylate all KCC isoforms at a conserved C-terminal threonine (Site-2, Thr1048 in KCC3A) to promote their inhibition; WNK pathway inhibition suppresses this phosphorylation; cells lacking SPAK/OSR1 activity have elevated KCC3A activity; a Site-2 alanine KCC3A mutant shows increased activity. In vitro kinase assay with recombinant proteins, SPAK/OSR1 double-knockin ES cells, 86Rb+ uptake, WNK pathway inhibitor The Biochemical journal High 24393035
2012 SPAK/OSR1 double-knockin ES cells (where SPAK/OSR1 cannot be activated by WNK) show abolished NKCC1 phosphorylation and activation, providing genetic evidence that NKCC1 is strictly dependent on SPAK/OSR1 activity; WNK1 and WNK3 activity is markedly elevated in knockin cells, revealing a feedback where downstream SPAK/OSR1 influence upstream WNK activity. Double-knockin ES cell model, 86Rb+ uptake, phospho-specific Western blot The Biochemical journal High 22032326
2012 In the DCT, OSR1 is dependent on SPAK for apical membrane localization and activity; in SPAK knockout mice, OSR1 becomes largely inactive and redistributes from the apical membrane to cytoplasmic WNK1-containing puncta, causing loss of NCC phosphorylation specifically in DCT1. SPAK knockout mouse model, immunofluorescence, phospho-specific Western blot, subcellular fractionation The Journal of biological chemistry High 22977235
2012 PI3K/Akt signaling activates the WNK-OSR1/SPAK-NCC phosphorylation cascade in hyperinsulinemic db/db mice; this is evidenced by PI3K inhibitors correcting increased OSR1/SPAK and NCC phosphorylation, and by genetic knockin mice (SpakT243A/+ and Osr1T185A/+) completely correcting increased NCC phosphorylation and elevated blood pressure in db/db mice. Knockin mouse genetics, pharmacological PI3K/Akt inhibition, phospho-specific Western blot Hypertension High 22949526
2012 ASK3 interacts with WNK1 and suppresses the WNK1-SPAK/OSR1 signaling pathway; Ask3 knockout mice display hyperactivation of SPAK/OSR1 in renal tubules and hypertension. Co-immunoprecipitation, siRNA knockdown, Ask3 knockout mouse model, phospho-specific Western blot, blood pressure measurement Nature communications Medium 23250415
2013 SPAK phosphorylates NBCe1-B at Ser65 and IRBIT/PP1 at Thr49 to regulate the Na+-HCO3- cotransporter; IRBIT and PIP2 activate NBCe1-B by a convergent non-additive mechanism, with SPAK phosphorylation setting the inhibitory resting state. In vitro phosphorylation, mutagenesis, functional transport assay in Xenopus oocytes Proceedings of the National Academy of Sciences of the United States of America Medium 23431199
2013 Aldosterone acutely stimulates SPAK phosphorylation in the distal convoluted tubule, which increases NCC phosphorylation and activity without changing total NCC abundance; gene silencing of SPAK eliminates the aldosterone effect on NCC activity; the effect is also SGK1-dependent. siRNA knockdown of SPAK in mpkDCT cells, phospho-specific Western blot, 22Na+ uptake assay, adrenalectomized rodent model American journal of physiology. Renal physiology Medium 23739593
2014 WNK4 in association with the scaffold protein Cab39 can activate NKCC1 in a SPAK/OSR1-independent manner; WNK4 possesses a PF2-like domain homologous to the SPAK/OSR1 CCT domain that mediates direct interaction with NKCC1, allowing WNK4 to anchor to the N-terminal domain of NKCC1 and promote cotransporter activation. Xenopus oocyte expression, yeast two-hybrid, homology modeling, 86Rb+ uptake, mutagenesis The Journal of biological chemistry Medium 24811174
2015 The CCT domain Leu502 residue of SPAK is essential for interaction with RFXV/I motifs in WNK1, NCC, and NKCC2; CCT domain L502A knock-in mice abolish these co-immunoprecipitation interactions, show markedly reduced SPAK kinase activity and NCC/NKCC2 phosphorylation, and display Gitelman syndrome features with reduced blood pressure. Knock-in mouse genetics, co-immunoprecipitation, phospho-specific Western blot, blood pressure/electrolyte measurement Human molecular genetics High 25994507
2015 Crystal structures of SPAK kinase domain (SPAK 63-403 at 3.1 Å and T243D mutant at 2.5 Å) reveal domain-swapped dimer conformations; a monomeric SPAK mutant retains kinase activity and is activated by WNK1 but shows reduced phosphorylation of NKCC2, indicating domain swapping modulates substrate access rather than intrinsic catalysis. X-ray crystallography, site-directed mutagenesis, in vitro kinase assay Biochemistry High 26208601
2017 Constitutively active SPAK (kinase-activating mutation in Stk39) expressed specifically in the DCT causes thiazide-treatable hypertension and hyperkalemia with NCC hyperphosphorylation; CA-SPAK drives ASDN remodeling with reduced connecting tubule mass and decreased ENaC and ROMK apical expression, revealing a DCT-ASDN structural coupling mechanism. Conditional knock-in mouse with DCT-specific Cre, phospho-specific Western blot, immunofluorescence, thiazide challenge Journal of the American Society of Nephrology : JASN High 28442491
2017 Allosteric inhibitors (rafoxanide, closantel) bind a conserved pocket on the C-terminal domains of SPAK and OSR1, distinct from the ATP-binding site, and inhibit kinase activity by targeting this allosteric site. In silico screening, in vitro kinase assay, binding studies ChemMedChem Medium 28371477
2019 WNK4 is the primary active WNK isoform in WNK bodies (spherical cytoplasmic condensates in DCT) and catalyzes SPAK/OSR1 phosphorylation therein; phosphorylated SPAK/OSR1 is present both at the apical membrane and in WNK bodies during K+ deprivation; WNK body formation requires Kir4.1-dependent K+ sensing in the DCT. Immunofluorescence in WNK4-deficient and Kir4.1-conditional KO mice, phospho-specific antibodies, dietary manipulation American journal of physiology. Renal physiology Medium 31736353
2020 The SPAK inhibitor ZT-1a (5-chloro-N-(5-chloro-4-((4-chlorophenyl)(cyano)methyl)-2-methylphenyl)-2-hydroxybenzamide) decreases SPAK-dependent phosphorylation of NKCC1 and simultaneously stimulates KCCs by reducing their SPAK-dependent phosphorylation; intracerebroventricular ZT-1a reduces CSF hypersecretion in post-hemorrhagic hydrocephalus; systemic ZT-1a reduces ischemia-induced CCC phosphorylation and cerebral edema in stroke. In vitro kinase assay, rodent stroke/hydrocephalus models, CCC phosphorylation assay, pharmacological inhibition Nature communications High 31911626
2021 STK39 interacts with and phosphorylates SNAI1 at T203, promoting SNAI1 nuclear retention and stability in breast cancer cells; STK39 inhibition destabilizes SNAI1, impairs EMT, and reduces tumor cell migration, invasion, and metastasis in vitro and in vivo. Co-immunoprecipitation, in vitro kinase/phosphorylation assay, subcellular fractionation, knockdown/overexpression functional assays, xenograft model Theranostics Medium 34335956
2021 STK39 binds PLK1 (identified by mass spectrometry) and promotes HCC progression by activating the ERK signaling pathway in a PLK1-dependent manner; STK39 knockdown arrests cells in G2/M and promotes apoptosis. Mass spectrometry, co-immunoprecipitation, RNA-seq, siRNA knockdown, cell cycle and apoptosis assays Theranostics Medium 33500714

Source papers

Stage 0 corpus · 100 papers · ranked by NIH iCite citations
Year Title Journal Citations PMID
2005 WNK1 regulates phosphorylation of cation-chloride-coupled cotransporters via the STE20-related kinases, SPAK and OSR1. The Journal of biological chemistry 388 16263722
2002 Cation chloride cotransporters interact with the stress-related kinases Ste20-related proline-alanine-rich kinase (SPAK) and oxidative stress response 1 (OSR1). The Journal of biological chemistry 318 12386165
2008 Activation of the thiazide-sensitive Na+-Cl- cotransporter by the WNK-regulated kinases SPAK and OSR1. Journal of cell science 317 18270262
2006 Functional interactions of the SPAK/OSR1 kinases with their upstream activator WNK1 and downstream substrate NKCC1. The Biochemical journal 261 16669787
2008 The regulation of salt transport and blood pressure by the WNK-SPAK/OSR1 signalling pathway. Journal of cell science 249 18843116
2010 SPAK-knockout mice manifest Gitelman syndrome and impaired vasoconstriction. Journal of the American Society of Nephrology : JASN 239 20813865
2008 From the Cover: Whole-genome association study identifies STK39 as a hypertension susceptibility gene. Proceedings of the National Academy of Sciences of the United States of America 236 19114657
2010 Role of the WNK-activated SPAK kinase in regulating blood pressure. EMBO molecular medicine 227 20091762
2014 The WNK-SPAK/OSR1 pathway: master regulator of cation-chloride cotransporters. Science signaling 212 25028718
2009 Angiotensin II signaling increases activity of the renal Na-Cl cotransporter through a WNK4-SPAK-dependent pathway. Proceedings of the National Academy of Sciences of the United States of America 201 19240212
2008 Regulation of NKCC2 by a chloride-sensing mechanism involving the WNK3 and SPAK kinases. Proceedings of the National Academy of Sciences of the United States of America 194 18550832
2011 A SPAK isoform switch modulates renal salt transport and blood pressure. Cell metabolism 181 21907141
2014 The WNK-regulated SPAK/OSR1 kinases directly phosphorylate and inhibit the K+-Cl- co-transporters. The Biochemical journal 175 24393035
2003 Characterization of the interaction of the stress kinase SPAK with the Na+-K+-2Cl- cotransporter in the nervous system: evidence for a scaffolding role of the kinase. The Journal of biological chemistry 175 14563843
2008 Dietary salt regulates the phosphorylation of OSR1/SPAK kinases and the sodium chloride cotransporter through aldosterone. Kidney international 169 18800028
2008 SPAK and OSR1: STE20 kinases involved in the regulation of ion homoeostasis and volume control in mammalian cells. The Biochemical journal 163 18092945
2011 Regulation of the NKCC2 ion cotransporter by SPAK-OSR1-dependent and -independent pathways. Journal of cell science 161 21321328
2006 Characterization of SPAK and OSR1, regulatory kinases of the Na-K-2Cl cotransporter. Molecular and cellular biology 136 16382158
2000 SPAK, a STE20/SPS1-related kinase that activates the p38 pathway. Oncogene 122 10980603
2011 MO25 is a master regulator of SPAK/OSR1 and MST3/MST4/YSK1 protein kinases. The EMBO journal 121 21423148
2017 Constitutively Active SPAK Causes Hyperkalemia by Activating NCC and Remodeling Distal Tubules. Journal of the American Society of Nephrology : JASN 118 28442491
2012 SPAK isoforms and OSR1 regulate sodium-chloride co-transporters in a nephron-specific manner. The Journal of biological chemistry 118 22977235
2012 Molecular physiology of SPAK and OSR1: two Ste20-related protein kinases regulating ion transport. Physiological reviews 115 23073627
2012 SPAK/OSR1 regulate NKCC1 and WNK activity: analysis of WNK isoform interactions and activation by T-loop trans-autophosphorylation. The Biochemical journal 114 22032326
2020 Modulation of brain cation-Cl- cotransport via the SPAK kinase inhibitor ZT-1a. Nature communications 88 31911626
2013 SPAK differentially mediates vasopressin effects on sodium cotransporters. Journal of the American Society of Nephrology : JASN 87 23393317
2012 Phosphatidylinositol 3-kinase/Akt signaling pathway activates the WNK-OSR1/SPAK-NCC phosphorylation cascade in hyperinsulinemic db/db mice. Hypertension (Dallas, Tex. : 1979) 87 22949526
2011 IRBIT governs epithelial secretion in mice by antagonizing the WNK/SPAK kinase pathway. The Journal of clinical investigation 86 21317537
2005 Hypotonic shock mediation by p38 MAPK, JNK, PKC, FAK, OSR1 and SPAK in osmosensing chloride secreting cells of killifish opercular epithelium. The Journal of experimental biology 85 15767308
2014 WNK-SPAK-NCC cascade revisited: WNK1 stimulates the activity of the Na-Cl cotransporter via SPAK, an effect antagonized by WNK4. Hypertension (Dallas, Tex. : 1979) 81 25113964
2012 Aldosterone does not require angiotensin II to activate NCC through a WNK4-SPAK-dependent pathway. Pflugers Archiv : European journal of physiology 76 22549242
2004 SPAK kinase is a substrate and target of PKCtheta in T-cell receptor-induced AP-1 activation pathway. The EMBO journal 74 14988727
2013 Convergence of IRBIT, phosphatidylinositol (4,5) bisphosphate, and WNK/SPAK kinases in regulation of the Na+-HCO3- cotransporters family. Proceedings of the National Academy of Sciences of the United States of America 68 23431199
2014 Actions of the protein kinase WNK1 on endothelial cells are differentially mediated by its substrate kinases OSR1 and SPAK. Proceedings of the National Academy of Sciences of the United States of America 61 25362046
2012 ASK3 responds to osmotic stress and regulates blood pressure by suppressing WNK1-SPAK/OSR1 signaling in the kidney. Nature communications 61 23250415
2010 Effect of angiotensin II on the WNK-OSR1/SPAK-NCC phosphorylation cascade in cultured mpkDCT cells and in vivo mouse kidney. Biochemical and biophysical research communications 61 20175999
2014 Discovery of Novel SPAK Inhibitors That Block WNK Kinase Signaling to Cation Chloride Transporters. Journal of the American Society of Nephrology : JASN 60 25377078
2011 Phenotypes of pseudohypoaldosteronism type II caused by the WNK4 D561A missense mutation are dependent on the WNK-OSR1/SPAK kinase cascade. Journal of cell science 58 21486947
2015 SPAK-mediated NCC regulation in response to low-K+ diet. American journal of physiology. Renal physiology 56 25651563
2007 A single binding motif is required for SPAK activation of the Na-K-2Cl cotransporter. Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology 56 17595523
2006 Genome-wide analysis of SPAK/OSR1 binding motifs. Physiological genomics 55 17032814
2019 WNK bodies cluster WNK4 and SPAK/OSR1 to promote NCC activation in hypokalemia. American journal of physiology. Renal physiology 53 31736353
2016 SPAK and OSR1 play essential roles in potassium homeostasis through actions on the distal convoluted tubule. The Journal of physiology 52 27068441
2010 On the substrate recognition and negative regulation of SPAK, a kinase modulating Na+-K+-2Cl- cotransport activity. American journal of physiology. Cell physiology 51 20463172
2007 Apoptosis-associated tyrosine kinase scaffolding of protein phosphatase 1 and SPAK reveals a novel pathway for Na-K-2C1 cotransporter regulation. American journal of physiology. Cell physiology 51 17267545
2006 SPAK and OSR1, key kinases involved in the regulation of chloride transport. Acta physiologica (Oxford, England) 51 16734747
2013 Dietary salt intake regulates WNK3-SPAK-NKCC1 phosphorylation cascade in mouse aorta through angiotensin II. Hypertension (Dallas, Tex. : 1979) 50 24019400
2011 Role of SPAK and OSR1 signalling in the regulation of NaCl cotransporters. Current opinion in nephrology and hypertension 49 21610494
2010 Multiple pathways for protein phosphatase 1 (PP1) regulation of Na-K-2Cl cotransporter (NKCC1) function: the N-terminal tail of the Na-K-2Cl cotransporter serves as a regulatory scaffold for Ste20-related proline/alanine-rich kinase (SPAK) AND PP1. The Journal of biological chemistry 49 20223824
2013 Aldosterone acutely stimulates NCC activity via a SPAK-mediated pathway. American journal of physiology. Renal physiology 46 23739593
2010 SORLA/SORL1 functionally interacts with SPAK to control renal activation of Na(+)-K(+)-Cl(-) cotransporter 2. Molecular and cellular biology 46 20385770
2012 Effect of heterozygous deletion of WNK1 on the WNK-OSR1/ SPAK-NCC/NKCC1/NKCC2 signal cascade in the kidney and blood vessels. Clinical and experimental nephrology 43 22294159
2008 An analysis of candidate autism loci on chromosome 2q24-q33: evidence for association to the STK39 gene. American journal of medical genetics. Part B, Neuropsychiatric genetics : the official publication of the International Society of Psychiatric Genetics 43 18348195
2019 The WNK-SPAK/OSR1 Kinases and the Cation-Chloride Cotransporters as Therapeutic Targets for Neurological Diseases. Aging and disease 42 31165006
2020 Physiological Processes Modulated by the Chloride-Sensitive WNK-SPAK/OSR1 Kinase Signaling Pathway and the Cation-Coupled Chloride Cotransporters. Frontiers in physiology 41 33192599
2014 Hypotonicity stimulates potassium flux through the WNK-SPAK/OSR1 kinase cascade and the Ncc69 sodium-potassium-2-chloride cotransporter in the Drosophila renal tubule. The Journal of biological chemistry 40 25086033
2006 The TNF receptor, RELT, binds SPAK and uses it to mediate p38 and JNK activation. Biochemical and biophysical research communications 40 16530727
2017 Rafoxanide and Closantel Inhibit SPAK and OSR1 Kinases by Binding to a Highly Conserved Allosteric Site on Their C-terminal Domains. ChemMedChem 38 28371477
2012 Phosphorylation of Na-Cl cotransporter by OSR1 and SPAK kinases regulates its ubiquitination. Biochemical and biophysical research communications 36 22846565
2001 Androgens induce expression of SPAK, a STE20/SPS1-related kinase, in LNCaP human prostate cancer cells. Molecular and cellular endocrinology 36 11514053
2018 The Calcium-Sensing Receptor Increases Activity of the Renal NCC through the WNK4-SPAK Pathway. Journal of the American Society of Nephrology : JASN 35 29848507
2014 A novel Ste20-related proline/alanine-rich kinase (SPAK)-independent pathway involving calcium-binding protein 39 (Cab39) and serine threonine kinase with no lysine member 4 (WNK4) in the activation of Na-K-Cl cotransporters. The Journal of biological chemistry 35 24811174
2021 STK39 is a novel kinase contributing to the progression of hepatocellular carcinoma by the PLK1/ERK signaling pathway. Theranostics 34 33500714
2012 Enhanced FGF23 serum concentrations and phosphaturia in gene targeted mice expressing WNK-resistant SPAK. Kidney & blood pressure research 34 23235437
2011 SPAK and WNK kinases: a new target for blood pressure treatment? Current opinion in nephrology and hypertension 34 21088576
2016 Deletion of the WNK3-SPAK kinase complex in mice improves radiographic and clinical outcomes in malignant cerebral edema after ischemic stroke. Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism 33 26861815
2012 WNK-OSR1/SPAK-NCC signal cascade has circadian rhythm dependent on aldosterone. Biochemical and biophysical research communications 33 23044422
2015 Critical role of the SPAK protein kinase CCT domain in controlling blood pressure. Human molecular genetics 32 25994507
2012 The WNK/SPAK and IRBIT/PP1 pathways in epithelial fluid and electrolyte transport. Physiology (Bethesda, Md.) 31 23026752
2013 Loss of WNK3 is compensated for by the WNK1/SPAK axis in the kidney of the mouse. American journal of physiology. Renal physiology 30 23427142
2013 Knockout of Ste20-like proline/alanine-rich kinase (SPAK) attenuates intestinal inflammation in mice. The American journal of pathology 30 23499375
2012 WNK3-SPAK interaction is required for the modulation of NCC and other members of the SLC12 family. Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology 30 22415098
2008 PKCdelta acts upstream of SPAK in the activation of NKCC1 by hyperosmotic stress in human airway epithelial cells. The Journal of biological chemistry 29 18550547
2020 Activated WNK3 induced by intracerebral hemorrhage deteriorates brain injury maybe via WNK3/SPAK/NKCC1 pathway. Experimental neurology 27 32589890
2014 Novel mechanisms of Na+ retention in obesity: phosphorylation of NKCC2 and regulation of SPAK/OSR1 by AMPK. American journal of physiology. Renal physiology 27 24808538
2014 Negative regulation of the creatine transporter SLC6A8 by SPAK and OSR1. Kidney & blood pressure research 27 25531585
2012 Kinases SPAK and OSR1 are upregulated by estradiol and activate NKCC1 in the developing hypothalamus. The Journal of neuroscience : the official journal of the Society for Neuroscience 27 22238094
2009 Ste20-related proline/alanine-rich kinase (SPAK) regulated transcriptionally by hyperosmolarity is involved in intestinal barrier function. PloS one 26 19343169
2024 Modulation of Cerebrospinal Fluid Dysregulation via a SPAK and OSR1 Targeted Framework Nucleic Acid in Hydrocephalus. Advanced science (Weinheim, Baden-Wurttemberg, Germany) 25 38353402
2022 Role of SPAK-NKCC1 signaling cascade in the choroid plexus blood-CSF barrier damage after stroke. Journal of neuroinflammation 25 35413993
2021 Knockdown of long non-coding RNA CDKN2B-AS1 suppresses the progression of breast cancer by miR-122-5p/STK39 axis. Bioengineered 25 34374638
2017 STK39, overexpressed in osteosarcoma, regulates osteosarcoma cell invasion and proliferation. Oncology letters 25 28943960
2020 WNK-SPAK/OSR1-NCC kinase signaling pathway as a novel target for the treatment of salt-sensitive hypertension. Acta pharmacologica Sinica 24 32724175
2014 Regulation of ClC-2 activity by SPAK and OSR1. Kidney & blood pressure research 24 25323061
2019 The interplay of renal potassium and sodium handling in blood pressure regulation: critical role of the WNK-SPAK-NCC pathway. Journal of human hypertension 23 30723251
2014 SPAK and OSR1 dependent down-regulation of murine renal outer medullary K channel ROMK1. Kidney & blood pressure research 23 25322850
2014 SPAK dependent regulation of peptide transporters PEPT1 and PEPT2. Kidney & blood pressure research 23 25376088
2013 STK39 polymorphism is associated with essential hypertension: a systematic review and meta-analysis. PloS one 23 23527223
2022 NF-κB Signaling-Mediated Activation of WNK-SPAK-NKCC1 Cascade in Worsened Stroke Outcomes of Ang II-Hypertensive Mice. Stroke 22 35272484
2019 Suppression of WNK1-SPAK/OSR1 Attenuates Bone Cancer Pain by Regulating NKCC1 and KCC2. The journal of pain 22 31085334
2019 WNK4-SPAK modulates lipopolysaccharide-induced macrophage activation. Biochemical pharmacology 22 31786261
2018 WNK-SPAK/OSR1 signaling: lessons learned from an insect renal epithelium. American journal of physiology. Renal physiology 22 29923766
2014 STE20/SPS1-related proline/alanine-rich kinase (SPAK) is critical for sodium reabsorption in isolated, perfused thick ascending limb. American journal of physiology. Renal physiology 21 25477470
2021 Roles of WNK4 and SPAK in K+-mediated dephosphorylation of the NaCl cotransporter. American journal of physiology. Renal physiology 20 33719576
2009 Epigenetic silencing of Stk39 in B-cell lymphoma inhibits apoptosis from genotoxic stress. The American journal of pathology 20 19717643
2016 SPAK plays a pathogenic role in IgA nephropathy through the activation of NF-κB/MAPKs signaling pathway. Free radical biology & medicine 19 27519267
2015 Domain-Swapping Switch Point in Ste20 Protein Kinase SPAK. Biochemistry 19 26208601
2022 MiR-223-3p-loaded exosomes from bronchoalveolar lavage fluid promote alveolar macrophage autophagy and reduce acute lung injury by inhibiting the expression of STK39. Human cell 18 35932362
2021 STK39 promotes breast cancer invasion and metastasis by increasing SNAI1 activity upon phosphorylation. Theranostics 18 34335956
2009 Behavioral analysis of Ste20 kinase SPAK knockout mice. Behavioural brain research 18 20006650