@article{132351,
  keywords = {Humans, proteomics, Transcription, Genetic, RNA, Messenger, Real-Time Polymerase Chain Reaction, Blotting, Western, Green Fluorescent Proteins, Chromatography, Liquid, Protein Interaction Maps, Cell Nucleus, Cells, Cultured, RNA, Small Interfering, Sirtuins, RNA Polymerase I, DNA, Ribosomal, Chromatin Assembly and Disassembly, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization},
  author = {Yuan-Chin Tsai and Todd Greco and Apaporn Boonmee and Yana Miteva and Ileana Cristea},
  title = {Functional proteomics establishes the interaction of SIRT7 with chromatin remodeling complexes and expands its role in regulation of RNA polymerase I transcription},
  abstract = {<p>Among mammalian sirtuins, SIRT7 is the only enzyme residing in nucleoli where ribosomal DNA is transcribed. Recent reports established that SIRT7 associates with RNA Pol I machinery and is required for rDNA transcription. Although defined by its homology to the yeast histone deacetylase Sir2, current knowledge suggests that SIRT7 itself has little to no deacetylase activity. Because only two SIRT7 interactions have been thus far described: RNA Pol I and upstream binding factor, identification of proteins and complexes associating with SIRT7 is critical to understanding its functions. Here, we present the first characterization of SIRT7 interaction networks. We have systematically investigated protein interactions of three EGFP-tagged SIRT7 constructs: wild type, a point mutation affecting rDNA transcription, and a deletion mutant lacking the predicted coiled-coil domain. A combinatorial proteomics and bioinformatics approach was used to integrate gene ontology classifications, functional protein networks, and normalized abundances of proteins co-isolated with SIRT7. The resulting refined proteomic data set confirmed SIRT7 interactions with RNA Pol I and upstream binding factor and highlighted association with factors involved in RNA Pol I- and II-dependent transcriptional processes and several nucleolus-localized chromatin remodeling complexes. Particularly enriched were members of the B-WICH complex, such as Mybbp1a, WSTF, and SNF2h. Prominent interactions were validated by a selected reaction monitoring-like approach using metabolic labeling with stable isotopes, confocal microscopy, reciprocal immunoaffinity precipitation, and co-isolation with endogenous SIRT7. To extend the current knowledge of mechanisms involved in SIRT7-dependent regulation of rDNA transcription, we showed that small interfering RNA-mediated SIRT7 knockdown leads to reduced levels of RNA Pol I protein, but not messenger RNA, which was confirmed in diverse cell types. The down-regulation of RNA Pol I protein levels placed in the context of SIRT7 interaction networks led us to propose that SIRT7 plays a crucial role in connecting the function of chromatin remodeling complexes to RNA Pol I machinery during transcription.</p>
},
  year = {2012},
  journal = {Mol Cell Proteomics},
  volume = {11},
  pages = {M111.015156},
  month = {02/2012},
  issn = {1535-9484},
  doi = {10.1074/mcp.M111.015156},
  language = {eng},
}