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J Proteomics. 2015 Apr 6;118:95-111. doi: 10.1016/j.jprot.2014.09.029. Epub 2014 Oct 18.

BioID identifies novel c-MYC interacting partners in cultured cells and xenograft tumors.

Author information

1
Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
2
The Hospital for Sick Children and Department of Paediatrics, University of Toronto, Toronto, ON Canada.
3
Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada. Electronic address: lpenn@uhnres.utoronto.ca.
4
Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada. Electronic address: brian.raught@uhnres.utoronto.ca.

Abstract

The BioID proximity-based biotin labeling technique was recently developed for the characterization of protein-protein interaction networks [1]. To date, this method has been applied to a number of different polypeptides expressed in cultured cells. Here we report the adaptation of BioID to the identification of protein-protein interactions surrounding the c-MYC oncoprotein in human cells grown both under standard culture conditions and in mice as tumor xenografts. Notably, in vivo BioID yielded >100 high confidence MYC interacting proteins, including >30 known binding partners. Putative novel MYC interactors include components of the STAGA/KAT5 and SWI/SNF chromatin remodeling complexes, DNA repair and replication factors, general transcription and elongation factors, and transcriptional co-regulators such as the DNA helicase protein chromodomain 8 (CHD8). Providing additional confidence in these findings, ENCODE ChIP-seq datasets highlight significant coincident binding throughout the genome for the MYC interactors identified here, and we validate the previously unreported MYC-CHD8 interaction using both a yeast two hybrid analysis and the proximity-based ligation assay. In sum, we demonstrate that BioID can be utilized to identify bona fide interacting partners for a chromatin-associated protein in vivo. This technique will allow for a much improved understanding of protein-protein interactions in a previously inaccessible biological setting.

BIOLOGICAL SIGNIFICANCE:

The c-MYC (MYC) oncogene is a transcription factor that plays important roles in cancer initiation and progression. MYC expression is deregulated in more than 50% of human cancers, but the role of this protein in normal cell biology and tumor progression is still not well understood, in part because identifying MYC-interacting proteins has been technically challenging: MYC-containing chromatin-associated complexes are difficult to isolate using traditional affinity purification methods, and the MYC protein is exceptionally labile, with a half-life of only ~30 min. Developing a new strategy to gain insight into MYC-containing protein complexes would thus mark a key advance in cancer research. The recently described BioID proximity-based labeling technique represents a promising new complementary approach for the characterization of protein-protein interactions (PPIs) in cultured cells. Here we report that BioID can also be used to characterize protein-protein interactions for a chromatin-associated protein in tumor xenografts, and present a comprehensive, high confidence in vivo MYC interactome. This article is part of a Special Issue entitled: Protein dynamics in health and disease. Guest Editors: Pierre Thibault and Anne-Claude Gingras.

KEYWORDS:

BioID; CHD8; Mass spectrometry; Proteomics; c-MYC oncogene

PMID:
25452129
DOI:
10.1016/j.jprot.2014.09.029
[Indexed for MEDLINE]

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