Pedicini, L, Wiktor, SD, Simmons, KJ orcid.org/0000-0003-4846-9097 et al. (2 more authors) (2021) Affinity-based proteomics reveals novel binding partners for Rab46 in endothelial cells. Scientific Reports, 11. 4054. ISSN 2045-2322
Abstract
Rab46 is a novel Ca2+-sensing Rab GTPase shown to have important functions in endothelial and immune cells. The presence of functional Ca2+-binding, coiled-coil and Rab domains suggest that Rab46 will be important for coupling rapid responses to signalling in many cell types. The molecular mechanisms underlying Rab46 function are currently unknown. Here we provide the first resource for studying Rab46 interacting proteins. Using liquid chromatography tandem mass spectrometry (LC–MS/MS) to identify affinity purified proteins that bind to constitutively active GFP-Rab46 or inactive GFP-Rab46 expressed in endothelial cells, we have revealed 922 peptides that interact with either the GTP-bound Rab46 or GDP-bound Rab46. To identify proteins that could be potential Rab46 effectors we performed further comparative analyses between nucleotide-locked Rab46 proteins and identified 29 candidate effector proteins. Importantly, through biochemical and imaging approaches we have validated two potential effector proteins; dynein and the Na2+/ K+ ATPase subunit alpha 1 (ATP1α1). Hence, our use of affinity purification and LC–MS/MS to identify Rab46 neighbouring proteins provides a valuable resource for detecting Rab46 effector proteins and analysing Rab46 functions.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © The Author(s) 2021. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Medicine and Health (Leeds) > School of Medicine (Leeds) > Leeds Institute of Cardiovascular and Metabolic Medicine (LICAMM) > Discovery & Translational Science Dept (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 13 Dec 2021 14:02 |
Last Modified: | 13 Dec 2021 14:02 |
Status: | Published |
Publisher: | Nature Research |
Identification Number: | 10.1038/s41598-021-83560-y |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:180326 |