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Bouley, Richard

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Bouley

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Richard

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Bouley, Richard
Author's Publications: search.filters.isAuthorOfPublication.2d1dbdaa-3680-49fd-a77f-3e4984fa7649

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    Identification of ROCK1 kinase as a critical regulator of Beclin1 mediated autophagy during metabolic stress
    (2013) Gurkar, Aditi U.; Chu, Kiki; Raj, Lakshmi; Bouley, Richard; Lee, Seung-Hwan; Kim, Young-Bum; Dunn, Sandra E.; Mandinova, Anna; Lee, Sam
    The Ser/Thr Rho kinase 1 (ROCK1) is known to play major roles in a wide range of cellular activities, including those involved in tumor metastasis and apoptosis. Here we identify an indispensable function of ROCK1 in metabolic stress-induced autophagy. Applying a proteomics approach, we characterize Beclin1, a proximal component of the PI(3)kinase class III lipid-kinase complex that induces autophagy, as an interacting partner of ROCK1. Upon nutrient deprivation, activated ROCK1 promotes autophagy by binding and phosphorylating Beclin1 at Thr119. This results in the specific dissociation of the Beclin1-Bcl-2 complex, without affecting the Beclin1-UVRAG interaction. Conversely, inhibition of ROCK1 activity increases Beclin1-Bcl-2 association, thus reducing nutritional stress-mediated autophagy. Genetic knockout of ROCK1 function in mice also leads to impaired autophagy as evidenced by reduced autophagosome formation. These results show that ROCK1 acts as a prominent upstream regulator of Beclin1-mediated autophagy and maintains a homeostatic balance between apoptosis and autophagy.
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    Renal Intercalated Cells Sense and Mediate Inflammation via the P2Y14 Receptor
    (Public Library of Science, 2015) Azroyan, Anie; Cortez-Retamozo, Virna; Bouley, Richard; Liberman, Rachel; Ruan, Ye Chun; Kiselev, Evgeny; Jacobson, Kenneth A.; Pittet, Mikael; Brown, Dennis; Breton, Sylvie
    Uncontrolled inflammation is one of the leading causes of kidney failure. Pro-inflammatory responses can occur in the absence of infection, a process called sterile inflammation. Here we show that the purinergic receptor P2Y14 (GPR105) is specifically and highly expressed in collecting duct intercalated cells (ICs) and mediates sterile inflammation in the kidney. P2Y14 is activated by UDP-glucose, a damage-associated molecular pattern molecule (DAMP) released by injured cells. We found that UDP-glucose increases pro-inflammatory chemokine expression in ICs as well as MDCK-C11 cells, and UDP-glucose activates the MEK1/2-ERK1/2 pathway in MDCK-C11 cells. These effects were prevented following inhibition of P2Y14 with the small molecule PPTN. Tail vein injection of mice with UDP-glucose induced the recruitment of neutrophils to the renal medulla. This study identifies ICs as novel sensors, mediators and effectors of inflammation in the kidney via P2Y14.
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    AQP2 is Necessary for Vasopressin- and Forskolin-Mediated Filamentous Actin Depolymerization in Renal Epithelial Cells
    (The Company of Biologists, 2011) Yui, Naofumi; Lu, Hua; Bouley, Richard; Brown, Dennis
    Remodeling of the actin cytoskeleton is required for vasopressin (VP)‐induced aquaporin 2 (AQP2) trafficking. Here, we asked whether VP and forskolin (FK)‐mediated F‐actin depolymerization depends on AQP2 expression. Using various MDCK and LLC‐PK1 cell lines with different AQP2 expression levels, we performed F‐actin quantification and immunofluorescence staining after VP/FK treatment. In MDCK cells, in which AQP2 is delivered apically, VP/FK mediated F‐actin depolymerization was significantly correlated with AQP2 expression levels. A decrease of apical membrane associated F‐actin was observed upon VP/FK treatment in AQP2 transfected, but not in untransfected cells. There was no change in basolateral actin staining under these conditions. In LLC‐PK1 cells, which deliver AQP2 basolaterally, a significant VP/FK mediated decrease in F‐actin was also detected only in AQP2 transfected cells. This depolymerization response to VP/FK was significantly reduced by siRNA knockdown of AQP2. By immunofluorescence, an inverse relationship between plasma membrane AQP2 and membrane‐associated F‐actin was observed after VP/FK treatment again only in AQP2 transfected cells. This is the first report showing that VP/FK mediated F‐actin depolymerization is dependent on AQP2 protein expression in renal epithelial cells, and that this is not dependent on the polarity of AQP2 membrane insertion.
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    Impaired AQP2 trafficking in Fxyd1 knockout mice: A role for FXYD1 in regulated vesicular transport
    (Public Library of Science, 2017) Arystarkhova, Elena; Bouley, Richard; Liu, Yi; Sweadner, Kathleen
    The final adjustment of urine volume occurs in the inner medullary collecting duct (IMCD), chiefly mediated by the water channel aquaporin 2 (AQP2). With vasopressin stimulation, AQP2 accumulation in the apical plasma membrane of principal cells allows water reabsorption from the lumen. We report that FXYD1 (phospholemman), better known as a regulator of Na,K-ATPase, has a role in AQP2 trafficking. Daytime urine of Fxyd1 knockout mice was more dilute than WT despite similar serum vasopressin, but both genotypes could concentrate urine during water deprivation. FXYD1 was found in IMCD. In WT mice, phosphorylated FXYD1 was detected intracellularly, and vasopressin induced its dephosphorylation. We tested the hypothesis that the dilute urine in knockouts was caused by alteration of AQP2 trafficking. In WT mice at baseline, FXYD1 and AQP2 were not strongly co-localized, but elevation of vasopressin produced translocation of both FXYD1 and AQP2 to the apical plasma membrane. In kidney slices, baseline AQP2 distribution was more scattered in the Fxyd1 knockout than in WT. Apical recruitment of AQP2 occurred in vasopressin-treated Fxyd1 knockout slices, but upon vasopressin washout, there was more rapid reversal of apical AQP2 localization and more heterogeneous cytoplasmic distribution of AQP2. Notably, in sucrose gradients, AQP2 was present in a detergent-resistant membrane domain that had lower sedimentation density in the knockout than in WT, and vasopressin treatment normalized its density. We propose that FXYD1 plays a role in regulating AQP2 retention in apical membrane, and that this involves transfers between raft-like membrane domains in endosomes and plasma membranes.
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    Overexpression of angiotensinogen downregulates aquaporin 1 expression via modulation of Nrf2–HO-1 pathway in renal proximal tubular cells of transgenic mice
    (SAGE Publications, 2016) Chang, Shiao-Ying; Lo, Chao-Sheng; Zhao, Xin-Ping; Liao, Min-Chun; Chenier, Isabelle; Bouley, Richard; Ingelfinger, Julie; Chan, John SD; Zhang, Shao-Ling
    Introduction: We aimed to examine the regulation of aquaporin 1 expression in an angiotensinogen transgenic mouse model, focusing on underlying mechanisms. Methods: Male transgenic mice overexpressing rat angiotensinogen in their renal proximal tubular cells (RPTCs) and rat immortalised RPTCs stably transfected with rat angiotensinogen cDNA were used. Results: Angiotensinogen-transgenic mice developed hypertension and nephropathy, changes that were either partially or completely attenuated by treatment with losartan or dual renin–angiotensin system blockade (losartan and perindopril), respectively, while hydralazine prevented hypertension but not nephropathy. Decreased expression of aquaporin 1 and heme oxygenase-1 and increased expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and sodium–hydrogen exchanger 3 were observed in RPTCs of angiotensinogen-transgenic mice and in angiotensinogen-transfected immortalised RPTCs. These parameters were normalised by dual renin–angiotensin system blockade. Both in vivo and in vitro studies identified a novel mechanism in which angiotensinogen overexpression in RPTCs enhances the cytosolic accumulation of Nrf2 via the phosphorylation of pGSK3β Y216. Consequently, lower intranuclear Nrf2 levels are less efficient to trigger heme oxygenase-1 expression as a defence mechanism, which subsequently diminishes aquaporin 1 expression in RPTCs. Conclusions: Angiotensinogen-mediated downregulation of aquaporin 1 and Nrf2 signalling may play an important role in intrarenal renin–angiotensin system-induced hypertension and kidney injury.