Purification and biochemical characterization of interchromatin granule clusters

• Published in: The EMBO journal Vol. 18; no. 15; pp. 4308 - 4320
• Main Authors: Mintz, Paul J, Patterson, Scott D, Neuwald, Andrew F, Spahr, Chris S, Spector, David L
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 02-08-1999; Blackwell Publishing Ltd
• Subjects: Amino Acid Sequence; Animals; Cell Nucleus - metabolism; Cell Nucleus - ultrastructure; Chromatin - chemistry; Chromatin - isolation & purification; Chromatin - metabolism; Electrophoresis, Gel, Two-Dimensional; Female; interchromatin granule clusters; Liver - metabolism; Liver - ultrastructure; Mass Spectrometry; Mice; Microscopy, Electron; Molecular Sequence Data; nuclear structure; RNA Precursors - genetics; RNA processing; RNA Splicing; RNA, Messenger - genetics; Sequence Homology, Amino Acid; speckles; xeroderma pigmentosum
• ISSN: 0261-4189; 1460-2075; 1460-2075
• DOI: 10.1093/emboj/18.15.4308

Components of the pre‐mRNA splicing machinery are localized in interchromatin granule clusters (IGCs) and perichromatin fibrils (PFs). Here we report the biochemical purification of IGCs. Approximately 75 enriched proteins were present in the IGC fraction. Protein identification employing a novel mass spectrometry strategy and peptide microsequencing identified 33 known proteins, many of which have been linked to pre‐mRNA splicing, as well as numerous uncharacterized proteins. Thus far, three new protein constituents of the IGCs have been identified. One of these, a 137 kDa protein, has a striking sequence similarity over its entire length to UV‐damaged DNA‐binding protein, a protein associated with the hereditary disease xeroderma pigmentosum group E, and to the 160 kDa subunit of cleavage polyadenylation specificity factor. Overall, these results provide a key framework that will enable the biological functions associated with the IGCs to be elucidated.