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A quantitative mass spectrometry-based approach to monitor the dynamics of endogenous chromatin-associated protein complexes.

Published version
Peer-reviewed

Type

Article

Change log

Authors

Papachristou, Evangelia K 
Holding, Andrew N 
Harvey, Kate 
Roumeliotis, Theodoros I 

Abstract

Understanding the dynamics of endogenous protein-protein interactions in complex networks is pivotal in deciphering disease mechanisms. To enable the in-depth analysis of protein interactions in chromatin-associated protein complexes, we have previously developed a method termed RIME (Rapid Immunoprecipitation Mass spectrometry of Endogenous proteins). Here, we present a quantitative multiplexed method (qPLEX-RIME), which integrates RIME with isobaric labelling and tribrid mass spectrometry for the study of protein interactome dynamics in a quantitative fashion with increased sensitivity. Using the qPLEX-RIME method, we delineate the temporal changes of the Estrogen Receptor alpha (ERα) interactome in breast cancer cells treated with 4-hydroxytamoxifen. Furthermore, we identify endogenous ERα-associated proteins in human Patient-Derived Xenograft tumours and in primary human breast cancer clinical tissue. Our results demonstrate that the combination of RIME with isobaric labelling offers a powerful tool for the in-depth and quantitative characterisation of protein interactome dynamics, which is applicable to clinical samples.

Description

Keywords

Breast cancer, Immunoprecipitation, Mass spectrometry, Protein–protein interaction networks

Journal Title

Nature Communications

Conference Name

Journal ISSN

2041-1723
2041-1723

Volume Title

9

Publisher

Springer Nature
Sponsorship
Wellcome Trust (108467/Z/15/Z)
Cancer Research UK (C14303/A17197)
Breast Cancer Campaign (2012NovemberPR042)
European Research Council (646876)
Cancer Research UK (19274)
Cancer Research UK (20411)