Quantitative phosphoproteomics of vasopressin-sensitive renal cells: Regulation of aquaporin-2 phosphorylation at two sites

Hoffert, Jason D.; Pisitkun, Trairak; Wang, Guanghui; Shen, Rong-Fong; Knepper, Mark A.

doi:10.1073/pnas.0600895103

Cited by 325 publications

(350 citation statements)

References 40 publications

(31 reference statements)

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“…Previous studies have shown that TG2 is phosphorylated by PKA at Ser 215 and Ser 216 [52] and at an unknown site(s) by PTEN-induced putative kinase 1 (PINK1; [53]). Phosphoproteomic based studies have identified numerous phosphorylation sites on Ser, Thr and Tyr residues in human and rat TG2 [54][55][56][57][58][59][60]. It would therefore be of value to identify the specific site(s) of TG2-associated serine and threonine phosphorylation triggered by the A1 adenosine receptor.…”

Section: A1 Adenosine Receptor-induced Phosphorylation Of Tg2mentioning

confidence: 99%

A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival

Vyas

Hargreaves

Bonner

et al. 2016

Biochemical Pharmacology

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The regulation of tissue transglutaminase (TG2) activity by the GPCR family is poorly understood. In this study, we investigated the modulation of TG2 activity by the A1 adenosine receptor in cardiomyocyte-like H9c2 cells.H9c2 cells were lysed following stimulation with the A1 adenosine receptor agonist N 6 -cyclopentyladenosine (CPA).Transglutaminase activity was determined using an amine incorporating and a protein cross linking assay. TG2 phosphorylation was assessed via immunoprecipitation and 3

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Section: A1 Adenosine Receptor-induced Phosphorylation Of Tg2mentioning

confidence: 99%

A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival

Vyas

Hargreaves

Bonner

et al. 2016

Biochemical Pharmacology

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“…Numerous studies, in both cell culture and animal models, suggest that PKA phosphorylation of serine 256 (S256) in the COOH tail of AQP2 plays a critical role in its apical trafficking (2)(3)(4)(5). In addition to S256, we have discovered recently that AQP2 is further phosphorylated on residues S261, S264, and S269 in the COOH tail in response to AVP stimulation (6,7).…”

mentioning

confidence: 94%

Acute regulation of aquaporin-2 phosphorylation at Ser-264 by vasopressin

Fenton

Moeller²,

Hoffert³

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

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131

106

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By phosphoproteome analysis, we identified a phosphorylation site, serine 264 (pS264), in the COOH terminus of the vasopressinregulated water channel, aquaporin-2 (AQP2). In this study, we examined the regulation of AQP2 phosphorylated at serine 264 (pS264 -AQP2) by vasopressin, using a phospho-specific antibody (anti-pS264). Immunohistochemical analysis showed pS264 -AQP2 labeling of inner medullary collecting duct (IMCD) from control mice, whereas AQP2 knockout mice showed a complete absence of labeling. In rat and mouse, pS264 -AQP2 was present throughout the collecting duct system, from the connecting tubule to the terminal IMCD. Immunogold electron microscopy, combined with double-labeling confocal immunofluorescence microscopy with organelle-specific markers, determined that the majority of pS264 resides in compartments associated with the plasma membrane and early endocytic pathways. In Brattleboro rats treated with [deamino-Cys-1, D-Arg-8]vasopressin (dDAVP), the abundance of pS264 -AQP2 increased 4-fold over controls. Additionally, dDAVP treatment resulted in a time-dependent change in the distribution of pS264 from predominantly intracellular vesicles, to both the basolateral and apical plasma membranes. Sixty minutes after dDAVP exposure, a proportion of pS264 -AQP2 was observed in clathrin-coated vesicles, early endosomal compartments, and recycling compartments, but not lysosomes. Overall, our results are consistent with a dynamic effect of AVP on the phosphorylation and subcellular distribution of AQP2.concentrating mechanism ͉ immunohistochemistry ͉ kidney ͉ trafficking T he maintenance of body water homeostasis depends on the fine control of renal water excretion, a process that is regulated by the antidiuretic hormone arginine vasopressin (AVP). An essential step in the action of AVP is the trafficking of intracellular vesicles containing the AVP-sensitive water channel aquaporin-2 (AQP2) to the plasma membrane of kidney collecting duct (CD) principal cells, a process that ultimately increases osmotically driven water reabsorption from the CD lumen and the production of concentrated urine (1). After small increases in plasma osmolality, AVP is released by the posterior pituitary gland and binds to the type II AVP receptor (V2R) in the CD, leading to activation of adenylyl cyclase, increased intracellular cAMP levels, increased intracellular calcium, and activation of PKA. Numerous studies, in both cell culture and animal models, suggest that PKA phosphorylation of serine 256 (S256) in the COOH tail of AQP2 plays a critical role in its apical trafficking (2-5). In addition to S256, we have discovered recently that AQP2 is further phosphorylated on residues S261, S264, and S269 in the COOH tail in response to AVP stimulation (6, 7).In this study, we examined the regulation of AQP2 phosphorylated at serine 264 (pS264-AQP2) by AVP in a number of in vivo models by using a phospho-specific antibody. Our findings demonstrated that pS264-AQP2 abundance is regulated acutely by AVP and that pS264-AQP2 app...

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“…All experiments were conducted in accord with animal protocol H-0110R1, which was approved by the Animal Care and Use Committee of the National Heart, Lung and Blood Institute. peptide enrichment (29). (Note that, other than IMAC, we carried out no upstream fractionation to maximize the precision of the quantification, but at the expense of a lower number of phosphopeptide identifications that would otherwise be the case.…”

Section: Methodsmentioning

confidence: 99%

Quantitative phosphoproteomic analysis reveals cAMP/vasopressin-dependent signaling pathways in native renal thick ascending limb cells

Gunaratne

Braucht

Rinschen

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

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103

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Quantitative mass spectrometry was used to identify hormonedependent signaling pathways in renal medullary thick ascending limb (mTAL) cells via phosphoproteomic analysis. Active transport of NaCl across the mTAL epithelium is accelerated by hormones that increase cAMP levels (vasopressin, glucagon, parathyroid hormone, and calcitonin). mTAL suspensions from rat kidneys were exposed (15 min) to a mixture of these four hormones. Tryptic phosphopeptides (immobilized metal affinity chromatography-enriched) were identified and quantified by mass spectrometry (LTQ-Orbitrap) using label-free methodology. We quantified a total of 654 phosphopeptides, of which 414 were quantified in three experimental pairs (hormone vs. vehicle). Of these phosphopeptides, 82% were statistically unchanged in abundance in response to the hormone mixture. In contrast, 48 phosphopeptides were significantly increased, whereas 28 were significantly decreased. The population of up-regulated phosphopeptides was highly enriched in basophilic kinase substrate motifs (AGC or calmodulin-sensitive kinase families), whereas the down-regulated sites were dominated by "proline-directed" motifs (cyclin-dependent or MAP kinase families). Bioinformatic classification uncovered overrepresentation of transmembrane transporters, protein phosphatase regulators, and cytoskeletal binding proteins among the regulated proteins. Immunoblotting with phospho-specific antibodies confirmed cAMP/vasopressin-dependent phosphorylation at Thr96, Ser126, and Ser874 of the Na + :K + :2Cl − cotransporter NKCC2, at Ser552 of the Na + :H + exchanger NHE3, and at Ser552 of β-catenin. Vasopressin also increased phosphorylation of NKCC2 at both Ser126 (more than fivefold) and Ser874 (more than threefold) in rats in vivo. Both sites were phosphorylated by purified protein kinase A during in vitro assays. These results support the view that, although protein kinase A plays a central role in mTAL signaling, additional kinases, including those that target proline-directed motifs, may be involved.protein phosphatase | glucose transporters | mass spectrometry | ion transporters | protein kinase T he thick ascending limb (TAL) of Henle's loop is a nephron segment that plays a critical role in the control of mammalian water excretion. Active NaCl transport by the medullary TAL (mTAL) drives the countercurrent multiplication process that concentrates the urine (1). Hormones that increase the concentration of the intracellular second messenger, cAMP, have been shown to enhance the rate of NaCl transport in mTAL cells (2). These hormones include parathyroid hormone (PTH), calcitonin, glucagon, and vasopressin (2). Among these, only vasopressin plays a selective role in regulation of water balance. The molecular targets for cAMP-mediated regulation in the mTAL include the apical Na + : K + :2Cl − cotransporter NKCC2 (gene symbol: Slc12a1) and the apical Na + :H + exchanger NHE3 (Slc9a3) (3).The signaling network that accounts for cAMP-dependent regulation of these transporters is largely unk...

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Quantitative phosphoproteomics of vasopressin-sensitive renal cells: Regulation of aquaporin-2 phosphorylation at two sites

Cited by 325 publications

References 40 publications

A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival

A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival

Acute regulation of aquaporin-2 phosphorylation at Ser-264 by vasopressin

Quantitative phosphoproteomic analysis reveals cAMP/vasopressin-dependent signaling pathways in native renal thick ascending limb cells

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