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Characterization of five novel vasopressin V2 receptor mutants causing nephrogenic diabetes insipidus reveals a role of tolvaptan for M272R-V2R mutation.

ABSTRACT: Nephrogenic diabetes insipidus (NDI) is a rare tubulopathy characterized by urinary concentration defect due to renal resistance to vasopressin. Loss-of-function mutations of vasopressin V2 receptor (V2R) gene (AVPR2) is the most common cause of the disease. We have identified five novel mutations L86P, R113Q, C192S, M272R, and W323_I324insR from NDI-affected patients. Functional characterization of these mutants revealed that R113Q and C192S were normally localized at the basolateral membrane of polarized Madin-Darby Canine Kidney (MDCK) cells and presented proper glycosylation maturation. On the other side, L86P, M272R, and W323_I324insR mutants were retained in endoplasmic reticulum and exhibited immature glycosylation and considerably reduced stability. All five mutants were resistant to administration of vasopressin analogues as evaluated by defective response in cAMP release. In order to rescue the function of the mutated V2R, we tested VX-809, sildenafil citrate, ibuprofen and tolvaptan in MDCK cells. Among these, tolvaptan was effective in rescuing the function of M272R mutation, by both allowing proper glycosylation maturation, membrane sorting and response to dDAVP. These results show an important proof of concept for the use of tolvaptan in patients affected by M272R mutation of V2R causing NDI.

SUBMITTER: Prosperi F

PROVIDER: S-EPMC7532466 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Characterization of five novel vasopressin V2 receptor mutants causing nephrogenic diabetes insipidus reveals a role of tolvaptan for M272R-V2R mutation.

Prosperi Federica F Suzumoto Yoko Y Marzuillo Pierluigi P Costanzo Vincenzo V Jelen Sabina S Iervolino Anna A Guarino Stefano S La Manna Angela A Miraglia Del Giudice Emanuele E Perna Alessandra F AF Zacchia Miriam M Cordat Emmanuelle E Capasso Giovambattista G Trepiccione Francesco F

Scientific reports 20201002 1

Nephrogenic diabetes insipidus (NDI) is a rare tubulopathy characterized by urinary concentration defect due to renal resistance to vasopressin. Loss-of-function mutations of vasopressin V2 receptor (V2R) gene (AVPR2) is the most common cause of the disease. We have identified five novel mutations L86P, R113Q, C192S, M272R, and W323_I324insR from NDI-affected patients. Functional characterization of these mutants revealed that R113Q and C192S were normally localized at the basolateral membrane o ...[more]

PMID: 33009446

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Project description:Tolvaptan, a vasopressin antagonist selective for the V2-subtype vasopressin receptor (V2R), is widely used in the treatment of hyponatremia and autosomal-dominant polycystic kidney disease (ADPKD). Its effects on signaling in collecting duct cells have not been fully characterized. Here, we perform RNA-seq in a collecting duct cell line (mpkCCD). The data show that tolvaptan inhibits the expression of mRNAs that were previously shown to be increased in response to vasopressin including aquaporin-2, but also reveals mRNA changes that were not readily predictable and suggest off-target actions of tolvaptan. One such action is activation of the MAPK kinase (ERK1/ERK2) pathway. Prior studies have shown that ERK1/ERK2 activation is essential in the regulation of a variety of cellular and physiological processes and can be associated with cell proliferation. In immunoblotting experiments, we demonstrated that ERK1/ERK2 phosphorylation in mpkCCD cells was significantly reduced by vasopressin, in contrast to the increases seen in non-collecting-duct cells overexpressing V2R in prior studies. We also found that tolvaptan has a strong effect to increase ERK1/ERK2 phosphorylation in the presence of vasopressin and that tolvaptan's effect to increase ERK1/ERK2 phosphorylation is absent in mpkCCD cells in which both protein kinase A (PKA)-catalytic subunits have been deleted. Thus, it appears that the tolvaptan effect to increase ERK activation is PKA-dependent and is not due to an off-target effect of tolvaptan. We conclude that in cells expressing V2R at endogenous levels: 1) vasopressin decreases ERK1/ERK2 activation; 2) in the presence of vasopressin, tolvaptan increases ERK1/ERK2 activation; and 3) these effects are PKA-dependent.NEW & NOTEWORTHY Vasopressin is a key hormone that regulates the function of the collecting duct of the kidney. ERK1 and ERK2 are enzymes that play key roles in physiological regulation in all cells. The authors used collecting duct cell cultures to investigate the effects of vasopressin and the vasopressin receptor antagonist tolvaptan on ERK1 and ERK2 phosphorylation and activation.

Dataset Information

Characterization of five novel vasopressin V2 receptor mutants causing nephrogenic diabetes insipidus reveals a role of tolvaptan for M272R-V2R mutation.

Publications

Characterization of five novel vasopressin V2 receptor mutants causing nephrogenic diabetes insipidus reveals a role of tolvaptan for M272R-V2R mutation.

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