title: BioID identifies novel c-MYC interacting partners in cultured cells and xenograft tumors.
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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Dharmendra
authors.initials: D
authors.lastname: Dingar
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Manpreet
authors.initials: M
authors.lastname: Kalkat
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Pak-Kei
authors.initials: PK
authors.lastname: Chan
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Tharan
authors.initials: T
authors.lastname: Srikumar
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Swneke D
authors.initials: SD
authors.lastname: Bailey
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: William B
authors.initials: WB
authors.lastname: Tu
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Etienne
authors.initials: E
authors.lastname: Coyaud
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Romina
authors.initials: R
authors.lastname: Ponzielli
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Max
authors.initials: M
authors.lastname: Kolyar
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Igor
authors.initials: I
authors.lastname: Jurisica
authors.affiliation: The Hospital for Sick Children and Department of Paediatrics, University of Toronto, Toronto, ON Canada.
authors.forename: Annie
authors.initials: A
authors.lastname: Huang
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Lupien
authors.affiliation: Princess Margaret Cancer Centre, University Health Network, and Department of Medical Biophysics, University of Toronto, Toronto, ON Canada. Electronic address: lpenn@uhnres.utoronto.ca.
authors.forename: Linda Z
authors.initials: LZ
authors.lastname: Penn
authors.affiliation: 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.
authors.forename: Brian
authors.initials: B
authors.lastname: Raught
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D064593:Heterografts; D051548:Histone Acetyltransferases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D000074443:Lysine Acetyltransferase 5; D008297:Male; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D009368:Neoplasm Transplantation; D009374:Neoplasms, Experimental / Q000235:genetics / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: C523331:CHD8 protein, human; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C489427:MYC protein, human; D016271:Proto-Oncogene Proteins c-myc; C445179:SWI-SNF-B chromatin-remodeling complex; D014157:Transcription Factors; D051548:Histone Acetyltransferases; C098489:KAT5 protein, human; D000074443:Lysine Acetyltransferase 5
keywords: BioID; CHD8; Mass spectrometry; Proteomics; c-MYC oncogene