The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity

VPAC1 is class B G protein-coupled receptors (GPCR) shared by pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP). The first cysteine (Cys37) in the N-terminal extracellular domain of mature VPAC1 is a free Cys not involved in the formation of conserved...

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Autores principales: Yu, Rongjie, Liu, Hongyu, Peng, Xinhe, Cui, Yue, Song, Suqin, Wang, Like, Zhang, Huahua, Hong, An, Zhou, Tianhong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Impact Journals LLC 2017
Materias:
Research Paper
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522101/
https://www.ncbi.nlm.nih.gov/pubmed/28473666
http://dx.doi.org/10.18632/oncotarget.17449
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author Yu, Rongjie
Liu, Hongyu
Peng, Xinhe
Cui, Yue
Song, Suqin
Wang, Like
Zhang, Huahua
Hong, An
Zhou, Tianhong
author_facet Yu, Rongjie
Liu, Hongyu
Peng, Xinhe
Cui, Yue
Song, Suqin
Wang, Like
Zhang, Huahua
Hong, An
Zhou, Tianhong
author_sort Yu, Rongjie
collection PubMed
description VPAC1 is class B G protein-coupled receptors (GPCR) shared by pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP). The first cysteine (Cys37) in the N-terminal extracellular domain of mature VPAC1 is a free Cys not involved in the formation of conserved intramolecular disulfide bonds. In order to investigate the biological role of this Cys37 in VPAC1, the wild-type VPAC1 and Cys37/Ala mutant (VPAC1-C37/A) were expressed stably as fusion proteins with enhanced yellow fluorescent protein (EYFP) respectively in Chinese hamster ovary (CHO) cells. Both VPAC1-EYFP and VPAC1-C37/A-EYFP trafficked to the plasma membrane normally, and CHO cells expressing VPAC1-EYFP displayed higher anti-apoptotic activity against camptothecin (CPT) induced apoptosis than the cells expressing VPAC1-C37/A-EYFP, while VPAC1-C37/A-CHO cells showed higher proliferative activity than VPAC1-CHO cells. Confocal microscopic analysis, western blotting and fluorescence quantification assay showed VPAC1-EYFP displayed significant nuclear translocation while VPAC1-C37/A-EYFP did not transfer into nucleus under the stimulation of VIP (0.1 nM). Acyl-biotin exchange assay and click chemistry-based palmitoylation assay confirmed for the first time the palmitoylation of Cys37, which has been predicted by bioinformatics analysis. And the palmitoylation inhibitor 2-bromopalmitate significantly inhibited the nuclear translocation of VPAC1-EYFP and its anti-apoptotic activity synchronously. These results indicated the palmitoylation of the Cys37 in the N-terminal extracellular domain of VPAC1 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity. These findings reveal for the first time the lipidation-mediating nuclear translocation of VPAC1 produces a novel anti-apoptotic signal pathway, which may help to promote new drug development strategy targeting VPAC1.
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spelling pubmed-55221012017-08-08 The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity Yu, Rongjie Liu, Hongyu Peng, Xinhe Cui, Yue Song, Suqin Wang, Like Zhang, Huahua Hong, An Zhou, Tianhong Oncotarget Research Paper VPAC1 is class B G protein-coupled receptors (GPCR) shared by pituitary adenylate cyclase activating polypeptide (PACAP) and vasoactive intestinal peptide (VIP). The first cysteine (Cys37) in the N-terminal extracellular domain of mature VPAC1 is a free Cys not involved in the formation of conserved intramolecular disulfide bonds. In order to investigate the biological role of this Cys37 in VPAC1, the wild-type VPAC1 and Cys37/Ala mutant (VPAC1-C37/A) were expressed stably as fusion proteins with enhanced yellow fluorescent protein (EYFP) respectively in Chinese hamster ovary (CHO) cells. Both VPAC1-EYFP and VPAC1-C37/A-EYFP trafficked to the plasma membrane normally, and CHO cells expressing VPAC1-EYFP displayed higher anti-apoptotic activity against camptothecin (CPT) induced apoptosis than the cells expressing VPAC1-C37/A-EYFP, while VPAC1-C37/A-CHO cells showed higher proliferative activity than VPAC1-CHO cells. Confocal microscopic analysis, western blotting and fluorescence quantification assay showed VPAC1-EYFP displayed significant nuclear translocation while VPAC1-C37/A-EYFP did not transfer into nucleus under the stimulation of VIP (0.1 nM). Acyl-biotin exchange assay and click chemistry-based palmitoylation assay confirmed for the first time the palmitoylation of Cys37, which has been predicted by bioinformatics analysis. And the palmitoylation inhibitor 2-bromopalmitate significantly inhibited the nuclear translocation of VPAC1-EYFP and its anti-apoptotic activity synchronously. These results indicated the palmitoylation of the Cys37 in the N-terminal extracellular domain of VPAC1 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity. These findings reveal for the first time the lipidation-mediating nuclear translocation of VPAC1 produces a novel anti-apoptotic signal pathway, which may help to promote new drug development strategy targeting VPAC1. Impact Journals LLC 2017-04-27 /pmc/articles/PMC5522101/ /pubmed/28473666 http://dx.doi.org/10.18632/oncotarget.17449 Text en Copyright: © 2017 Yu et al. http://creativecommons.org/licenses/by/3.0/ This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0/) (CC-BY), which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Research Paper
Yu, Rongjie
Liu, Hongyu
Peng, Xinhe
Cui, Yue
Song, Suqin
Wang, Like
Zhang, Huahua
Hong, An
Zhou, Tianhong
The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title_full The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title_fullStr The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title_full_unstemmed The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title_short The palmitoylation of the N-terminal extracellular Cys37 mediates the nuclear translocation of VPAC1 contributing to its anti-apoptotic activity
title_sort palmitoylation of the n-terminal extracellular cys37 mediates the nuclear translocation of vpac1 contributing to its anti-apoptotic activity
topic Research Paper
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5522101/
https://www.ncbi.nlm.nih.gov/pubmed/28473666
http://dx.doi.org/10.18632/oncotarget.17449
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