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In 2001, with-no-lysine (WNK) kinases were identified as the genes responsible for the human hereditary hypertensive disease pseudohypoaldosteronism type II (PHAII). It took a further 6 years to clarify that WNK kinases participate in a signaling cascade with oxidative stress-responsive gene 1 (OSR1), Ste20-related proline-alanine-rich kinase (SPAK), and thiazide-sensitive NaCl cotransporter (NCC) in the kidney and the constitutive activation of this signaling cascade is the molecular basis of PHAII. Since this discovery, the WNK-OSR1/SPAK-NCC signaling cascade has been shown to be involved not only in PHAII but also in the regulation of blood pressure under normal and pathogenic conditions, such as hyperinsulinemia. However, the molecular mechanisms of WNK kinase regulation by dietary and hormonal factors and by PHAII-causing mutations remain poorly understood. In 2012, two additional genes responsible for PHAII, Kelch-like 3 ( KLHL3) and Cullin3, were identified. At the time of their discovery, the molecular mechanisms underlying the interaction between these genes and their involvement in PHAII were unknown. Here we review the pathophysiological roles of the WNK signaling cascade clarified to date and introduce a new mechanism of WNK kinase regulation by KLHL3 and Cullin3, which provides insight on previously unknown mechanisms of WNK kinase regulation. [ABSTRACT FROM AUTHOR]

Aquaporin‐2 (AQP2) water channels are proteins that are recycled between intracellular vesicles and the apical plasma membrane in renal collecting ducts. Lipopolysaccharide‐responsive beige‐like anchor protein (LRBA) is a protein kinase A (PKA) anchoring protein that creates compartmentalized PKA signalling responsible for AQP2 phosphorylation. In response to increased plasma osmolality, vasopressin/cyclic adenosine monophosphate (cAMP)/PKA signalling phosphorylates AQP2, promoting AQP2 trafficking into the apical plasma membrane and increasing water reabsorption from urine. However, the molecular mechanisms by which LRBA mediates vasopressin‐induced AQP2 phosphorylation remain unknown. To investigate AQP2 intracellular localization and phosphorylation status in vivo, a density gradient ultracentrifugation technique was combined with an in situ proximity ligation assay, super‐resolution structured illumination microscopy and immunoelectron microscopy. Most of the AQP2 was localized on the recycling endosome in the presence of tolvaptan, a vasopressin type 2 receptor (V2R) antagonist. Desmopressin, a V2R agonist, phosphorylated AQP2, translocating it from the recycling endosome to the apical plasma membrane. In contrast, LRBA was constitutively localized at the recycling endosome. Therefore, LRBA and AQP2 were well colocalized in the absence of vasopressin stimulation. The loss of LRBA/PKA signalling by Lrba knockout impaired vasopressin‐induced AQP2 phosphorylation, resulting in AQP2 retention at the recycling endosome. Defective AQP2 trafficking caused low urinary concentrating ability in Lrba−/− mice. The LRBA‐PKA complex created compartmentalized PKA signalling at the recycling endosome, which facilitated AQP2 phosphorylation in response to vasopressin. Key points: Membrane proteins are continuously internalized into the endosomal system via endocytosis, after which they are either recycled back to the plasma membrane or degraded at the lysosome.In T cells, lipopolysaccharide‐responsive beige‐like anchor protein (LRBA) binds directly to the cytotoxic T lymphocyte antigen 4 (CTLA‐4), a checkpoint immune molecule, to prevent CTLA‐4 lysosomal degradation and promote its vesicle recycling.LRBA has different physiological functions in renal collecting ducts. LRBA and aquaporin‐2 (AQP2) water channels were colocalized on the recycling endosome in vivo in the absence of the anti‐diuretic hormone vasopressin.LRBA promoted vasopressin‐induced AQP2 trafficking, increasing water reabsorption from urine via AQP2. LRBA determined renal responsiveness to vasopressin at recycling endosomes.LRBA is a ubiquitously expressed anchor protein. LRBA signalosomes might regulate membrane trafficking of several constitutively recycled proteins at recycling endosomes. [ABSTRACT FROM AUTHOR]

Avacopan is a novel C5a receptor antagonist recently approved for the treatment of microscopic polyangiitis and granulomatosis with polyangiitis. To our knowledge, thrombocytopenia induced by avacopan has not been reported. We report a case of a 78‐year‐old man with microscopic polyangiitis who developed rapidly progressive glomerulonephritis (RPGN) and vasculitis neuropathy. After developing RPGN, he was treated with prednisolone, which was ineffective. As the dosage of corticosteroids was decreased, he developed impaired dorsiflexion of the left ankle, tingling and numbness in his feet, consistent with vasculitis neuropathy. After a 3‐day administration of methylprednisolone, we started avacopan and prednisolone 20 mg/d to reduce the corticosteroid dosage. One week after starting avacopan, platelet counts began to decrease, eventually leading to the cessation of the drug. The possibility of thrombotic microangiopathy and heparin‐induced thrombocytopenia was considered unlikely given the clinical course and laboratory studies. After 3 weeks of avacopan cessation, platelet counts began to increase, suggesting avacopan as the most probable cause of thrombocytopenia. Our case highlights the importance of postmarketing surveillance of avacopan to identify its adverse events that were not reported in clinical trials to ensure its safe use. Clinicians should carefully monitor platelet counts when using avacopan. [ABSTRACT FROM AUTHOR]

AMP‐activated protein kinase (AMPK) inactivation in chronic kidney disease (CKD) leads to energy status deterioration in the kidney, constituting the vicious cycle of CKD exacerbation. Unc‐51‐like kinase 1 (ULK1) is considered a downstream molecule of AMPK; however, it was recently reported that the activity of AMPK could be regulated by ULK1 conversely. We demonstrated that AMPK and ULK1 activities were decreased in the kidneys of CKD mice. However, whether and how ULK1 is involved in the underlying mechanism of CKD exacerbation remains unknown. In this study, we investigated the ULK1 involvement in CKD, using ULK1 knockout mice. The CKD model of Ulk1−/− mice exhibited significantly exacerbated renal function and worsening renal fibrosis. In the kidneys of the CKD model of Ulk1−/− mice, reduced AMPK and its downstream β‐oxidation could be observed, leading to an energy deficit of increased AMP/ATP ratio. In addition, AMPK signaling in the kidney was reduced in control Ulk1−/− mice with normal renal function compared to control wild‐type mice, suggesting that ULK1 deficiency suppressed AMPK activity in the kidney. This study is the first to present ULK1 as a novel therapeutic target for CKD treatment, which regulates AMPK activity in the kidney. [ABSTRACT FROM AUTHOR]

Key Clinical Message: We report a patient with adult‐onset nephronophthisis (NPHP) that was identified a homozygous full gene deletion of NPHP1 and a heterozygous PKD1 mutation. We suggest that the PKD1 mutation may have epistatically ameliorated NPHP disease progression and that the screening of larger cohorts for similar possible epistatic effects is needed. We report a patient with adult‐onset nephronophthisis (NPHP) that was identified a homozygous full gene deletion of NPHP1 and a heterozygous PKD1 mutation. We suggest that the PKD1 mutation may have epistatically ameliorated NPHP disease progression and that the screening of larger cohorts for similar possible epistatic effects is needed. [ABSTRACT FROM AUTHOR]

R3551 --> 768.4 --> Pneumonia is the most common cause of the acute respiratory distress syndrome (ARDS), a potentially fatal lung disease characterized by hyperinflammation and endothelial barrier failure. Infectious and inflammatory stimuli can cause rapid downregulation of cystic fibrosis transmembrane conductance regulator (CFTR), and inhibition of CFTR has been proposed to increase lung microvascular endothelial permeability in vitro. Here, we identify loss of lung microvascular endothelial CFTR as important pathomechanism in lung barrier failure in pneumonia‐induced ARDS, delineate the molecular signaling pathway underlying this effect and identify CFTR potentiation as novel therapeutic strategy in ARDS. CFTR was downregulated in endothelial cells following Streptococcus pneumoniae(S.pn.) infection in human and murine lung tissue. Isolated perfused lungs revealed that CFTR inhibition increased endothelial permeability in parallel with intracellular Cl‐ and Ca2+concentrations ([Cl‐]i, [Ca2+]i). Inhibition of the Cl‐ sensitive with‐no‐lysine kinase 1 (WNK1) replicated the effect of CFTR inhibition on endothelial permeability and endothelial [Ca2+]i while WNK1 activation attenuated it. Endothelial [Ca2+]i transients and permeability in response to inhibition of either CFTR or WNK1 were prevented by inhibition of the cation channel transient receptor potential vanilloid 4 (TRPV4). Mice deficient in Trpv4 (Trpv4‐/‐) developed less lung edema and protein leak than their wild‐type littermates following infection with S. pn. Conversely, lungs of heterozygous Wnk1‐deficient mice (Wnk1+/‑) showed spontaneous leak. The CFTR potentiator ivacaftor prevented CFTR loss and reduced endothelial leak in response to pneumolysin (PLY), a key S.pn. virulence factor, or plasma from COVID‐19 patients in vitro, and prevented lung edema and protein leak after S. pn.infection in vivo. Lung infection causes rapid loss of CFTR that promotes lung edema formation through intracellular Cl‐accumulation, inhibition of WNK1 and subsequent disinhibition of TRPV4, resulting in endothelial Ca2+ influx and vascular barrier failure. Ivacaftor prevents CFTR loss and may thus present a promising therapeutic strategy in ARDS due to severe pneumonia including COVID‐19. [ABSTRACT FROM AUTHOR]

Aquaporin-7 (AQP7) is expressed in adipose tissue, permeated by water and glycerol, and is involved in lipid metabolism. AQP7-null mice develop obesity, insulin resistance, and adipocyte hypertrophy. Here, we show that AQP7 is expressed in adipocyte plasma membranes, and is re-localized to intracellular membranes in response to catecholamine in mouse white adipose tissue. We found that internalization of AQP7 was induced by PKA activation and comparative gene identification 58 (CGI-58). This relocation was confirmed by functional studies in 3T3-L1 adipocytes. Collectively, these results suggest that AQP7 makes several contributions to adipocyte metabolism, in both cortical and intracellular membranes. [ABSTRACT FROM AUTHOR]

Dendritic cells (DCs) have the ability to present antigen and play a critical role in the induction of the acquired immune response. Skin DCs uptake antigen and subsequently migrate to regional draining lymph nodes (LNs), where they activate naive T cells. Here we show that the water/glycerol channel protein aquaporin 7 (AQP7) is expressed on epidermal and dermal DCs and involved in the initiation of primary immune responses. AQP7-deficient DCs showed a decreased cellular uptake of low-molecular-mass compounds (fluorescein isothiocyanate and Lucifer yellow) and high-molecular-mass substances (ovalbumin and dextran), suggesting that AQP7 is involved in antigen uptake. AQP7-deficient DCs also exhibited reduced chemokine-dependent cell migration in comparison to wild-type DCs. Consistent with these in vitro results, AQP7-deficient mice demonstrated a reduced accumulation of antigen-retaining DCs in the LNs after antigen application to the skin, which could be attributed to decreased antigen uptake and migration. Coincidentally, AQP7-deficient mice had impaired antigen-induced sensitization in a contact hypersensitivity model. These observations suggested that AQP7 in skin DCs is primarily involved in antigen uptake and in the subsequent migration of DCs and is responsible for antigen presentation and the promotion of downstream immune responses. [ABSTRACT FROM AUTHOR]

ClC chloride channels (ClCs) can be classified into two groups in terms of their cellular localizations: ClCs present in the plasma membranes and those residing in intracellular organelles. Members of the latter group, including ClC-3, ClC-4, ClC-5, ClC-6, and ClC-7, are often co-expressed in a variety of cell types in many organs. Although the localization of individual channels within cells has been investigated, the degree of overlap between the locations of different ClCs in the same cell has not been clarified. To address this question, different combinations of ClCs, engineered to encode specific epitope tags (FLAG or HA), were either transiently or stably transfected into HEK293 cells, and we then compared the intracellular localization of the expressed channel proteins by immunofluorescence microscopy. Immunofluorescence images of the alternatively labeled channels clearly showed significant co-localization between all pair-wise combinations of ClCs. In particular, ClC-3, ClC-4, and ClC-5 showed a high degree of co-localization. As a significant degree of co-localization between ClCs was observed, we used co-immunoprecipitation to evaluate oligomer formation, and found that each ClC tested could form homo-oligomers, and that any pair-wise combination of ClC-3, ClC-4, and ClC-5 could also form hetero-oligomers. Neither ClC-6 nor ClC-7 was co-precipitated with any other channel protein. These results suggest that within cells ClC-3, ClC-4, and ClC-5 may have combinatorial functions, whereas ClC-6 and ClC-7 are more likely to function as homo-oligomers. © 2005 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

The article focuses on the renal pseudohypoaldosteronism type 1 (adPHA1), which is a rare condition of congenital resistance in regards to aldosterone. It discusses the adPHA1, as the mild phenotype for renal-limited mineralocorticoid resistance, elevation of plasma aldosterone level (PAL), sequencing analysis revealing a heterozygous nonsense mutation.

Background: There have been still few case reports of pseudohypoaldosteronism type II (PHA2), also known as Gordon's syndrome, genetically diagnosed, and this is the first report of familial PHA2 case in Japan with a novel D564N mutation in WNK4. Methods: A 29‐year‐old woman was admitted to our hospital due to hyperkalemia (serum potassium: 6.4 mmol/L). She had mild hypertension (135/91 mm Hg), a bicarbonate level at the lower limit of the normal range (HCO3: 22 mmol/L) with a normal anion gap, low plasma renin activity (0.2 ng ml‐1 hr‐1), and high urinary calcium excretion (505.4 mg/g Cre). A hereditary condition was suspected because her mother also had the same symptoms. We performed a comprehensive genetic analysis for major inherited kidney diseases with next‐generation sequencing including the genes responsible for PHA2 (WNK1, WNK4, KLHL3, and CUL3). Results: Genetic analysis revealed that the patient and her mother had a novel missense mutation (D564N) in the acidic motif in WNK4, which leads to the diagnosis of PHA2. Administration of trichlormethiazide (1 mg/day) effectively ameliorated her blood pressure (114/69 mm Hg), plasma bicarbonate (25 mmol/L), serum potassium (4.3 mmol/L), and urinary calcium excretion (27.2 mg/g Cre). Conclusion: We report the first Japanese familial case of PHA2 with WNK4 mutation. D564N mutation in WNK4 is a novel genetic cause of PHA2 with a relatively mild phenotype. [ABSTRACT FROM AUTHOR]