Proteomics of a fuzzy organelle: Interphase chromatin

Abstract

Chromatin proteins mediate replication, regulate expression, and ensure integrity of the genome. So far, a comprehensive inventory of interphase chromatin has not been determined. This is largely due to its heterogeneous and dynamic composition, which makes conclusive biochemical purification difficult, if not impossible. As a fuzzy organelle, it defies classical organellar proteomics and cannot be described by a single and ultimate list of protein components. Instead, we propose a new approach that provides a quantitative assessment of a protein's probability to function in chromatin. We integrate chromatin composition over a range of different biochemical and biological conditions. This resulted in interphase chromatin probabilities for 7635 human proteins, including 1840 previously uncharacterized proteins. We demonstrate the power of our large-scale data-driven annotation during the analysis of cyclin-dependent kinase (CDK) regulation in chromatin. Quantitative protein ontologies may provide a general alternative to list-based investigations of organelles and complement Gene Ontology. Synopsis A probabilistic definition of interphase chromatin captures its dynamic and regulated composition with a new proteomics-based quantitative protein ontology approach. This resource is subsequently applied for the identification of novel factors whose interphase chromatin association depends on cyclin-dependent kinase (CDK). Static protein lists in organellar proteomics are replaced by degrees of chromatin involvement. Chromatin components are defined by biological co-behavior instead of biochemical co-purification. Interphase chromatin probabilities (ICPs) for 7635 proteins focus datasets on likely chromatin factors. Chromatin enrichment for proteomics (ChEP) defines a simple but efficient procedure to enrich chromatin for proteomic analysis. Proteomic analysis of CDK-dependent chromatin composition reveals novel cell cycle-regulated chromatin factors. A combination of biochemical and biological classifiers results in assignment of a protein 'interphase chromatin probability' (ICP), making an organelle whose dynamic composition defies rigorous black-and-white delineation amenable to quantitative proteomics. © 2014 The Authors.