title: Proteomic interrogation of human chromatin.
abstract: Chromatin proteins provide a scaffold for DNA packaging and a basis for epigenetic regulation and genomic maintenance. Despite understanding its functional roles, mapping the chromatin proteome (i.e. the "Chromatome") is still a continuing process. Here, we assess the biological specificity and proteomic extent of three distinct chromatin preparations by identifying proteins in selected chromatin-enriched fractions using mass spectrometry-based proteomics. These experiments allowed us to produce a chromatin catalog, including several proteins ranging from highly abundant histone proteins to less abundant members of different chromatin machinery complexes. Using a Normalized Spectral Abundance Factor approach, we quantified relative abundances of the proteins across the chromatin enriched fractions giving a glimpse into their chromosomal abundance. The large-scale data sets also allowed for the discovery of a variety of novel post-translational modifications on the identified chromatin proteins. With these comparisons, we find one of the probed methods to be qualitatively superior in specificity for chromatin proteins, but inferior in proteomic extent, evidencing a compromise that must be made between biological specificity and broadness of characterization. Additionally, we attempt to identify proteins in eu- and heterochromatin, verifying the enrichments by characterizing the post-translational modifications detected on histone proteins from these chromatin regions. In summary, our results provide insights into the value of different methods to extract chromatin-associated proteins and provide starting points to study the factors that may be involved in directing gene expression and other chromatin-related processes.
authors.affiliation: Department of Chemistry, Princeton University, Princeton, New Jersey, United States of America.
authors.forename: Mariana P
authors.initials: MP
authors.lastname: Torrente
authors.forename: Barry M
authors.initials: BM
authors.lastname: Zee
authors.forename: Nicolas L
authors.initials: NL
authors.lastname: Young
authors.forename: Richard C
authors.initials: RC
authors.lastname: Baliban
authors.forename: Gary
authors.initials: G
authors.lastname: LeRoy
authors.forename: Christodoulos A
authors.initials: CA
authors.lastname: Floudas
authors.forename: Sandra B
authors.initials: SB
authors.lastname: Hake
authors.forename: Benjamin A
authors.initials: BA
authors.lastname: Garcia
mesh_terms: D002843:Chromatin / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D020543:Proteome / Q000032:analysis*; D040901:Proteomics; D053719:Tandem Mass Spectrometry
chemical_list: D002843:Chromatin; D020543:Proteome