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Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome

Specific nuclear sub-compartments that are regions of fundamental processes such as gene expression or DNA repair, contain phosphoinositides (PIPs). PIPs thus potentially represent signals for the localization of specific proteins into different nuclear functional domains. We performed limited prote...

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Autores principales: Sztacho, Martin, Šalovská, Barbora, Červenka, Jakub, Balaban, Can, Hoboth, Peter, Hozák, Pavel
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7824793/
https://www.ncbi.nlm.nih.gov/pubmed/33406800
http://dx.doi.org/10.3390/cells10010068
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author Sztacho, Martin
Šalovská, Barbora
Červenka, Jakub
Balaban, Can
Hoboth, Peter
Hozák, Pavel
author_facet Sztacho, Martin
Šalovská, Barbora
Červenka, Jakub
Balaban, Can
Hoboth, Peter
Hozák, Pavel
author_sort Sztacho, Martin
collection PubMed
description Specific nuclear sub-compartments that are regions of fundamental processes such as gene expression or DNA repair, contain phosphoinositides (PIPs). PIPs thus potentially represent signals for the localization of specific proteins into different nuclear functional domains. We performed limited proteolysis followed by label-free quantitative mass spectrometry and identified nuclear protein effectors of the most abundant PIP—phosphatidylinositol 4,5-bisphosphate (PIP2). We identified 515 proteins with PIP2-binding capacity of which 191 ‘exposed’ proteins represent a direct PIP2 interactors and 324 ‘hidden’ proteins, where PIP2 binding was increased upon trypsin treatment. Gene ontology analysis revealed that ‘exposed’ proteins are involved in the gene expression as regulators of Pol II, mRNA splicing, and cell cycle. They localize mainly to non-membrane bound organelles—nuclear speckles and nucleolus and are connected to the actin nucleoskeleton. ‘Hidden’ proteins are linked to the gene expression, RNA splicing and transport, cell cycle regulation, and response to heat or viral infection. These proteins localize to the nuclear envelope, nuclear pore complex, or chromatin. Bioinformatic analysis of peptides bound in both groups revealed that PIP2-binding motifs are in general hydrophilic. Our data provide an insight into the molecular mechanism of nuclear PIP2 protein interaction and advance the methodology applicable for further studies of PIPs or other protein ligands.
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spelling pubmed-78247932021-01-24 Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome Sztacho, Martin Šalovská, Barbora Červenka, Jakub Balaban, Can Hoboth, Peter Hozák, Pavel Cells Article Specific nuclear sub-compartments that are regions of fundamental processes such as gene expression or DNA repair, contain phosphoinositides (PIPs). PIPs thus potentially represent signals for the localization of specific proteins into different nuclear functional domains. We performed limited proteolysis followed by label-free quantitative mass spectrometry and identified nuclear protein effectors of the most abundant PIP—phosphatidylinositol 4,5-bisphosphate (PIP2). We identified 515 proteins with PIP2-binding capacity of which 191 ‘exposed’ proteins represent a direct PIP2 interactors and 324 ‘hidden’ proteins, where PIP2 binding was increased upon trypsin treatment. Gene ontology analysis revealed that ‘exposed’ proteins are involved in the gene expression as regulators of Pol II, mRNA splicing, and cell cycle. They localize mainly to non-membrane bound organelles—nuclear speckles and nucleolus and are connected to the actin nucleoskeleton. ‘Hidden’ proteins are linked to the gene expression, RNA splicing and transport, cell cycle regulation, and response to heat or viral infection. These proteins localize to the nuclear envelope, nuclear pore complex, or chromatin. Bioinformatic analysis of peptides bound in both groups revealed that PIP2-binding motifs are in general hydrophilic. Our data provide an insight into the molecular mechanism of nuclear PIP2 protein interaction and advance the methodology applicable for further studies of PIPs or other protein ligands. MDPI 2021-01-04 /pmc/articles/PMC7824793/ /pubmed/33406800 http://dx.doi.org/10.3390/cells10010068 Text en © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Sztacho, Martin
Šalovská, Barbora
Červenka, Jakub
Balaban, Can
Hoboth, Peter
Hozák, Pavel
Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title_full Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title_fullStr Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title_full_unstemmed Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title_short Limited Proteolysis-Coupled Mass Spectrometry Identifies Phosphatidylinositol 4,5-Bisphosphate Effectors in Human Nuclear Proteome
title_sort limited proteolysis-coupled mass spectrometry identifies phosphatidylinositol 4,5-bisphosphate effectors in human nuclear proteome
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7824793/
https://www.ncbi.nlm.nih.gov/pubmed/33406800
http://dx.doi.org/10.3390/cells10010068
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