Protocol for nuclear export signal characterization of cGAS in mammalian cells

Protocol for nuclear export signal characterization of cGAS in mammalian cells

Summary: The cyclic GMP-AMP synthase (cGAS) is the principal DNA sensor, which binds DNA and triggers the type I interferon production. We used ISD45 or inactivated Vaccinia Virus (VACV) to stimulate cGAS and monitored cellular localization by immunofluorescence microscopy, Operetta high-content scr...

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Main Authors: Yu Huang, Myles McLean, Chen Liang, Fei Guo
Format: Article
Language:English
Published: Elsevier 2021-09-01
Series:STAR Protocols
Subjects:
Cell Biology
Immunology
Molecular Biology
Online Access:http://www.sciencedirect.com/science/article/pii/S2666166721003567
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spelling doaj-2b26fd1244f4474bbf2c3caebc1f071d2021-09-19T05:00:10ZengElsevierSTAR Protocols2666-16672021-09-0123100649Protocol for nuclear export signal characterization of cGAS in mammalian cellsYu Huang0Myles McLean1Chen Liang2Fei Guo3NHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, and Center for AIDS Research, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, P. R. China; Corresponding authorMcGill University AIDS Centre, Lady Davis Institute, Jewish General Hospital, Montreal H3T 1E2, CanadaMcGill University AIDS Centre, Lady Davis Institute, Jewish General Hospital, Montreal H3T 1E2, CanadaNHC Key Laboratory of Systems Biology of Pathogens, Institute of Pathogen Biology, and Center for AIDS Research, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100730, P. R. China; Corresponding authorSummary: The cyclic GMP-AMP synthase (cGAS) is the principal DNA sensor, which binds DNA and triggers the type I interferon production. We used ISD45 or inactivated Vaccinia Virus (VACV) to stimulate cGAS and monitored cellular localization by immunofluorescence microscopy, Operetta high-content screening, and cytoplasmic/nuclear fractionation. LocNES server was used to predict cGAS nuclear export signal (NES) sequence and characterized the function by mutagenesis. This protocol provides a prototype of cGAS subcellular distribution or the identification of NES in other proteins.For complete details on the use and execution of this protocol, please refer to Sun et al. Sun et al. (2021)http://www.sciencedirect.com/science/article/pii/S2666166721003567Cell BiologyImmunologyMolecular Biology
collection DOAJ
language English
format Article
sources DOAJ
author Yu Huang
Myles McLean
Chen Liang
Fei Guo
spellingShingle Yu Huang
Myles McLean
Chen Liang
Fei Guo
Protocol for nuclear export signal characterization of cGAS in mammalian cells
STAR Protocols
Cell Biology
Immunology
Molecular Biology
author_facet Yu Huang
Myles McLean
Chen Liang
Fei Guo
author_sort Yu Huang
title Protocol for nuclear export signal characterization of cGAS in mammalian cells
title_short Protocol for nuclear export signal characterization of cGAS in mammalian cells
title_full Protocol for nuclear export signal characterization of cGAS in mammalian cells
title_fullStr Protocol for nuclear export signal characterization of cGAS in mammalian cells
title_full_unstemmed Protocol for nuclear export signal characterization of cGAS in mammalian cells
title_sort protocol for nuclear export signal characterization of cgas in mammalian cells
publisher Elsevier
series STAR Protocols
issn 2666-1667
publishDate 2021-09-01
description Summary: The cyclic GMP-AMP synthase (cGAS) is the principal DNA sensor, which binds DNA and triggers the type I interferon production. We used ISD45 or inactivated Vaccinia Virus (VACV) to stimulate cGAS and monitored cellular localization by immunofluorescence microscopy, Operetta high-content screening, and cytoplasmic/nuclear fractionation. LocNES server was used to predict cGAS nuclear export signal (NES) sequence and characterized the function by mutagenesis. This protocol provides a prototype of cGAS subcellular distribution or the identification of NES in other proteins.For complete details on the use and execution of this protocol, please refer to Sun et al. Sun et al. (2021)
topic Cell Biology
Immunology
Molecular Biology
url http://www.sciencedirect.com/science/article/pii/S2666166721003567
work_keys_str_mv AT yuhuang protocolfornuclearexportsignalcharacterizationofcgasinmammaliancells
AT mylesmclean protocolfornuclearexportsignalcharacterizationofcgasinmammaliancells
AT chenliang protocolfornuclearexportsignalcharacterizationofcgasinmammaliancells
AT feiguo protocolfornuclearexportsignalcharacterizationofcgasinmammaliancells
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