Abstract:
UNLABELLED:Mapping protein-protein interactions for chromatin-associated proteins remains challenging. Here we explore the use of BioID, a proximity biotinylation approach in which a mutated biotin ligase (BirA*) is fused to a bait of interest, allowing for the local activation of biotin and subsequent biotinylation of proteins in the bait vicinity. BioID allowed for successful interactome mapping of core histones and members of the mediator complex. We explored the background signal produced by the BioID approach and found that using distinct types of controls increased the stringency of our statistical analysis with SAINTexpress. A direct comparison of BioID with our AP-MS protocol optimized for chromatin-associated protein complexes revealed that the approaches identified few shared interaction partners and enriched for distinct biological processes; yet, both approaches permitted the recovery of biologically meaningful interactions. While no clear bias could be observed for either technique toward protein complexes of particular functions, BioID allowed for the purification of proteins of lower cellular abundance. Finally, we were able to identify a strong association of MED4 with the centrosome by BioID and validated this finding by immunofluorescence. In summary, BioID complements AP-MS for the study of chromatin-associated protein complexes. BIOLOGICAL SIGNIFICANCE:This manuscript describes the application of BioID, a proximity biotinylation approach, to chromatin-associated proteins, namely core histones and members of the mediator complex. We observed that BioID was successful at identifying known interaction partners for the baits tested, but also allowed novel putative interaction partners to be identified. By performing a detailed comparison of BioID versus a standard method for interactome mapping (affinity purification coupled to mass spectrometry, AP-MS), we show that the approaches were complementary, allowing for purification of different interaction partners. These interaction partners were different in the biological processes they are associated with, but also in their abundance. BioID represents a significant technical development in the field of chromatin research by expanding the search space for interactome mapping beyond what is possible with AP-MS. This article is part of a Special Issue entitled: Protein dynamics in health and disease. Guest Editors: Pierre Thibault and Anne-Claude Gingras.
journal_name
J Proteomicsjournal_title
Journal of proteomicsauthors
Lambert JP,Tucholska M,Go C,Knight JD,Gingras ACdoi
10.1016/j.jprot.2014.09.011subject
Has Abstractpub_date
2015-04-06 00:00:00pages
81-94eissn
1874-3919issn
1876-7737pii
S1874-3919(14)00435-7journal_volume
118pub_type
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journal_title:Journal of proteomics
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abstract:UNLABELLED:Systems biology studies require the capability to quantify with high precision proteins spanning a broad range of abundances across multiple samples. However, the broad range of protein expression in cells often precludes the detection of low-abundance proteins. Different sample processing techniques can be ...
journal_title:Journal of proteomics
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journal_title:Journal of proteomics
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更新日期:2014-05-30 00:00:00
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journal_title:Journal of proteomics
pub_type: 杂志文章
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pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章,评审
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
doi:10.1016/j.jprot.2013.06.031
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journal_title:Journal of proteomics
pub_type: 杂志文章
doi:10.1016/j.jprot.2013.07.026
更新日期:2013-12-16 00:00:00
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journal_title:Journal of proteomics
pub_type: 杂志文章,评审
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
doi:10.1016/j.jprot.2018.05.001
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
doi:10.1016/j.jprot.2013.12.012
更新日期:2014-03-17 00:00:00
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章
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journal_title:Journal of proteomics
pub_type: 杂志文章,评审
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journal_title:Journal of proteomics
pub_type: 杂志文章
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更新日期:2012-06-18 00:00:00
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journal_title:Journal of proteomics
pub_type: 杂志文章
doi:10.1016/j.jprot.2012.06.024
更新日期:2012-09-18 00:00:00
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journal_title:Journal of proteomics
pub_type: 杂志文章
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abstract:UNLABELLED:The composition and toxicological profile of the venom of the rattlesnake Crotalus simus in Mexico was analyzed at the subspecies and individual levels. Venoms of the subspecies C. s. simus, C. s. culminatus and C. s. tzabcan greatly differ in the expression of the heterodimeric neurotoxin complex 'crotoxin'...
journal_title:Journal of proteomics
pub_type: 杂志文章
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abstract:UNLABELLED:Brevibacterium linens DSM 20158 is an industrially important actinobacterium which is well-known for the production of amino acids and enzymes. However, as this strain has an unsequenced genome, there is no detailed information regarding its proteome although another strain of this microbe, BL2, has a shotgu...
journal_title:Journal of proteomics
pub_type: 杂志文章
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abstract:UNLABELLED:Quercus suber L. is a forest tree with remarkable ecological, social and economic value in the southern Europe ecosystems. To circumvent the difficulties of breeding such long-lived species like Q. suber in a conventional fashion, clonal propagation of Q. suber elite trees can be carried out, although this p...
journal_title:Journal of proteomics
pub_type: 杂志文章
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