WNK3

With‑no‑lysine kinase 3 (WNK3) is a serine/threonine kinase that functions by regulating downstream signaling molecules. WNK3 mainly regulates intracellular and extracellular Na+, Cl‑ and K+ levels by regulating downstream ion transporters, the disruption of which has been associated with cerebral ischemia, epilepsy, glioma and other diseases. In addition, WNK3 was demonstrated to regulate neuronal splicing factor RNA binding fox‑1 homolog‑1 to influence autism. Over the past 20 years, accumulating evidence has reported that dysfunctional WNK3 signaling was involved in the pathologies of various neurological disorders; therefore, WNK3 has become a promising therapeutic target for ameliorating the corresponding symptoms of such disorders...

WNK kinases regulate electro-neutral cotransporters that are controlled by osmotic stress and chloride. We showed previously that autophosphorylation of WNK1 is inhibited by chloride, raising the possibility that WNKs are activated by osmotic stress. Here we demonstrate that unphosphorylated WNK isoforms 3 and 1 autophosphorylate in response to osmotic pressure in vitro, applied with the crowding agent polyethylene glycol 400 or osmolyte ethylene glycol, and that this activation is opposed by chloride. Small Angle X-ray Scattering of WNK3 in the presence and absence of PEG400, static light scattering in ethylene glycol, and crystallography of WNK1 were used to understand mechanism...

WNK (With No Lysine (K)) kinases regulate epithelial ion transport in the kidney to maintain homeostasis of electrolyte concentrations and blood pressure. Chloride ion directly binds WNK kinases to inhibit autophosphorylation and activation. Changes in extracellular potassium are thought to regulate WNKs through changes in intracellular chloride. Prior studies demonstrate that in some distal nephron epithelial cells, intracellular potassium changes with chronic low or high potassium diet. We therefore investigated whether potassium regulates WNK activity independent of chloride...

The with-no-lysine (WNK) kinase causes pseudohypoaldosteronism type II, a genetic form of hypertension. Due to ∼80% similarity among four isoforms (WNK1/2/3/4) of the WNK protein family, the discovery of an ATP-competitive inhibitor renders a significant challenge. Here, we combined molecular modeling and molecular dynamics simulations to study the structural and conformational properties of the WNK kinase isoforms bound to an ATP competitive inhibitor (WNK463). We have also investigated the effect of phosphorylation on the conformational properties of each isoform...

BACKGROUND: Intracerebral hemorrhage (ICH) is the common brain diseases in middle-aged and elderly people, with high disability and/or mortality rate, and is a serious public health concern. Both WNK3 kinase and the WNK3/SPAK/NKCC1 signaling pathway play an integral role in maintaining normal cell homeostasis. However, their role and underlying mechanisms in ICH-induced secondary brain injury (SBI) have yet to be elucidated. METHODS: We established an ICH model using male Sprague-Dawley (SD) rats by injecting autologous arterial blood into the unilateral basal ganglia...

Cation-coupled chloride cotransporters (CCC) play a role in modulating intracellular chloride concentration ([Cl- ]i ) and cell volume. Cell shrinkage and cell swelling are accompanied by an increase or decrease in [Cl- ]i , respectively. Cell shrinkage and a decrease in [Cl- ]i increases the activity of NKCCs (Na-K-Cl cotransporters: NKCC1, NKCC2 and Na-Cl) and inhibits the activity of KCCs (K-Cl cotransporters: KCC1 to KCC4), while cell swelling and an increase in [Cl- ]i activates KCCs and inhibits NKCCs; thus, it is unlikely that the same kinase is responsible for both effects...

Anaplastic lymphoma kinase (ALK) fusions have been recognized as a therapeutic target in non-small cell lung cancer (NSCLC). However, molecular signatures and clinical characteristics of the Chinese population with ALK-rearranged NSCLC are not well elucidated. In the present study, we carried out targeted next-generation sequencing on tissue and plasma ctDNA samples in 1688 patients with NSCLC. Overall, ALK fusions were detected in 70 patients (4.1%), and the frequencies of ALK fusions detected in tissue and plasma samples were 5...

Background: Liver is the most common metastatic site in advanced colorectal cancer. Most patients with colorectal cancer liver metastasis (CRLM) do not benefit from current treatment. Patient-derived xenografts (PDXs) with defined molecular signatures are attractive models for preclinical studies. Methods: Successfully established PDXs were evaluated to elucidate their fidelity of patients' biologic characteristics (pathologic, genetic and protein properties, together with chemosensitivity). The genomic variations of PDXs were analyzed by next-generation sequencing to explore the underlying molecular mechanism of metastasis and potential therapeutic targets...

The With-No-Lysine (WNK) kinase family plays a significant role in regulating cation-chloride cotransporters, blood pressure and body fluid homeostasis. Mutations in the gene of WNK family, especially in WNK1 and WNK4 are responsible for pseudohypoaldosteronism type II (PHAII), characterized by hypertension. The selective inhibition of WNK1 over other isoforms has created an immense challenge in the design of an ATP competitive inhibitor due to their high conservatism. In this work, we have compared the selectivity of the inhibitor WNK463, which was designed for WNK1 with other WNK family isoforms by comprehensive molecular modeling, docking and molecular dynamics simulations in conjunction with the Molecular Mechanics Poisson-Boltzmann Surface Area method...

Granule cell dispersion (GCD) is a common neuropathological feature of hippocampal sclerosis (HS) in patients with temporal lobe epilepsy (TLE). However, the underlying molecular mechanism of GCD formation remains unclear. The present study aimed to investigate the expressional changes of With No Lysine protein kinase subtype 3 (WNK3), a molecule upstream of cation-chloride cotransporters with reciprocal expression in sclerosed hippocampus of TLE patients. Using immunofluorescence staining, we analyzed WNK3 immunoreactivity in hippocampal specimens from histologically normal controls and TLE patients with HS...

Ion Transport across the cell membrane is required to maintain cell volume homeostasis. In response to changes in extracellular osmolarity, most cells activate specific metabolic or membrane-transport pathways to respond to cell swelling or shrinkage and return their volume to its normal resting state. This process involves the rapid adjustment of the activities of channels and transporters that mediate flux of K+ , Na+ , Cl- , and small organic osmolytes. Cation chloride cotransporters (CCCs) NKCCs and KCCs are a family of membrane proteins modulated by changes in cell volume and/or in the intracellular chloride concentration ([Cl- ]i )...

PURPOSE: Nonobstructive azoospermia (NOA) affects 1% of the male population; however, despite state-of-the-art clinical assessment, for most patients the cause is unknown. We capitalized on an analysis of multiplex families in the Middle East to identify highly penetrant genetic causes. METHODS: We used whole-exome sequencing (WES) in 8 consanguineous families and combined newly discovered genes with previously reported ones to create a NOA gene panel, which was used to identify additional variants in 75 unrelated idiopathic NOA subjects and 74 fertile controls...

Serotonergic neurons in the dorsal raphe nucleus (DRN) act as wake-inducing neurons in the sleep-wake cycle and are controlled by gamma-aminobutyric acid (GABA) synaptic inputs. We investigated daily changes in GABAergic inhibition of the rat DRN neurons and the role of nitric oxide (NO) and cation-chloride co-transporters in the GABAergic action. Neuronal NO synthase (nNOS) was co-expressed in 74% of serotonergic DRN neurons and nNOS expression was higher during daytime (the sleep cycle) than that during nighttime (the wake cycle)...

Neuronal regeneration and axonal regrowth mechanisms in the injured mammalian central nervous system are largely unknown. As part of a major pathway for inhibiting axonal regeneration, activated neuronal glycosylphosphatidylinositol-anchored Nogo receptor (NgR) interacts with LINGO-1 and p75NTR to form a complex at the cell surface. However, it was found in our previous report that upregulation of NgR stimulated by injury plays a key role in neuronal regeneration in the neonatal cortex freeze-lesion model, but its downstream signalling remains elusive...

Recurrent microduplications/microdeletions of 1q21.1 are characterized by variable phenotypes ranging from normal development to developmental delay (DD) and congenital anomalies. Their interpretation is challenging especially in families with affected and unaffected carriers. We used whole exome sequencing (WES) to look for sequence variants in two male probands with inherited 1q21.1 CNVs that could explain their more severe phenotypes. One proband had a 1q21.1 deletion transmitted from maternal grandmother, while the other had a paternal duplication...

With-no-lysine kinase (WNK) and Na+ -K+ -2Cl- cotransporter 1 (NKCC1) are involved in the pathogenesis of hypertension. In this study, we investigated the WNK-NKCC1 signaling pathway in spontaneously hypertensive rats (SHR) and their associated susceptibility to stroke injury. Basal NKCC1 protein levels were higher in SHR than in normotensive Wistar Kyoto (WKY) rat brains. After inducing ischemic stroke, adult male WKY and SHR received either saline or NKCC1 inhibitor bumetanide (10 mg/kg/day, i.p.) starting at 3-h post-reperfusion...

Cell volume homeostasis requires the dynamically regulated transport of ions across the plasmalemma. While the ensemble of ion transport proteins involved in cell volume regulation is well established, the molecular coordinators of their activities remain poorly characterized. We utilized a functional kinomics approach including a kinome-wide siRNA-phosphoproteomic screen, a high-content kinase inhibitor screen, and a kinase trapping-Orbitrap mass spectroscopy screen to systematically identify essential kinase regulators of KCC3 Thr991 /Thr1048 phosphorylation - a key signaling event in cell swelling-induced regulatory volume decrease (RVD)...

BACKGROUND: WNK kinase is a serine/threonine kinase that plays an important role in normal blood pressure homeostasis. WNK3 was previously found to enhance the activity of sodium chloride cotransporter (NCC) in Xenopus oocyte. However, the mechanism through which it works remains unclear. METHODS: Using overexpression and siRNA knock-down techniques, the effects of WNK3 on NCC in both Cos-7 and mouse distal convoluted cells were analyzed by Western blot. RESULTS: We found that WNK3 significantly increased NCC protein expression in a dose-dependent manner...

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Members of With-no-lysine (WNK) family of serine-threonine kinase are key regulators of chloride ion transport in diverse cell types, controlling the activity and the surface expression of cation-chloride (Na(+)/K(+)-Cl(-)) co-transporters. Mutations in WNK1 and WNK4 are linked to a hereditary form of hypertension, and WNKs have been extensively investigated pertaining to their roles in renal epithelial ion homeostasis. However, some members of the WNK family and their splice isoforms are also expressed in the mammalian brain, and have been implicated in aspects of hereditary neuropathy as well as neuronal and glial survival...