Michael P Washburn, PhD

Associate Professor, Department of Pathology and Laboratory Medicine
Ph.D., Michigan State University (Biochemistry and Environmental Toxicology)
Postdoctoral Fellow, University of Washington (Molecular Biotechnology)


http://www.stowers.org/faculty/washburn-lab

Research Interest:

Quantitative proteomic analysis of transcriptional regulatory complexes and assembly of protein interaction networks

The dynamic changes of a proteome or fractions of a proteome; i.e., organelles and protein complexes, can be analyzed via quantitative proteomic methods.  We largely carry out a chromatography-based proteomic technique known as multidimensional protein identification technology (MudPIT) combined with label free quantitation using normalized spectral counts (normalized spectral abundance factor, NSAF).  In spectral counting, the total number of tandem mass spectra that match peptides to a particular protein is used to measure the abundance of proteins in a complex mixture.  We have demonstrated that the NSAF approach generates datasets that have a high degree of statistical similarity to Affymetrix GeneChipTM datasets.  This provides a foundation for bioinformatics analysis of NSAF based MudPIT datasets using established GeneChipTM tools.  One example of the application of the NSAF approach to quantitative proteomic analysis includes a study on the human Mediator transcriptional complex.  We demonstrated that purifying Mediator through different subunits as tagged baits results in different forms of Mediator. We validated our proteomic and two stage statistical analysis with semi quantitative western blotting and a functional assay for the presence of RNA Polymerase II in Mediator preparations.  In addition, we have developed a novel approach for assembling probabilistic protein interaction networks based on NSAF values.  We applied this to the human Tip49a/Tip49b protein interaction network.  Finally, we determined the function of a poorly characterized but highly conserved protein, Rtr1.  Rtr1 interacts with RNA polymerase II and regulates the phosphorylation state of the C-terminal domain of the largest subunit of RNA polymerase II.  Going forward, we will continue to use quantitative proteomic technologies to develop new insights into the function, regulation, and architecture of transcriptional regulatory complexes.

Selected Publications:

  1. Suganuma T, Mushegian A, Swanson SK, Abmayr SM, Florens L,Washburn MP, Workman JL. The ATAC acetyltransferase complex coordinates MAP kinases to regulate JNK target genes, Cell2010;142:726-36. Abstract
  2. D'Angiolella V, Donato V, Vijayakumar S, Saraf A, Florens L, Washburn MP, Dynlacht B, Pagano M. SCF(Cyclin F) controls centrosome homeostasis and mitotic fidelity through CP110 degradation, Nature2010;466:138-42. Abstract
  3. Zhang Y, Wen Z, Washburn MP, Florens L. Refinements to label free proteome quantitation: how to deal with peptides shared by multiple proteins, Anal Chem. 2010;82: 2272-81. Abstract
  4. Lin C, Smith ER, Takahashi H, Lai KC, Martin-Brown S, Florens L,Washburn MP, Conaway JW, Conaway RC, Shilatifard A. AFF4, a component of the ELL/P-TEFb elongation complex and a shared subunit of MLL chimeras, can link transcription elongation to leukemia, Mol Cell. 2010;37:429-37. Abstract
  5. Fournier ML, Paulson A, Pavelka N, Mosley AL, Zueckert-Gaudenz K, Bradford WD, Glynn E, Li H, Sardiu ME, Fleharty B, Seidel C, Florens LWashburn MP. Delayed Correlation of mRNA and Protein Expression in Rapamycin Treated Cells and a Role for Ggc1 in Cellular Sensitivity to Rapamycin, Mol Cell Proteomics. 2010;9:271-84. Abstract
  6. Sardiu ME, Gilmore JM, Carozza MJ, Li B, Workman JL, Florens L,Washburn MP. Determining protein complex architecture using a probabilistic deletion network derived from quantitative proteomics, PLoS ONE. 2009;4:e7310 Abstract
  7. Zhang Y, Wen Z, Washburn MP, Florens L. Effect of Dynamic Exclusion on Spectral Count Based Quantitative Proteomics. Ana. Chem.2009;81:6317-6326. Abstract
  8. Sardiu, ME, Florens, L, and Washburn, MP. Evaluation of Clustering Algorithms for Protein Complex and Protein Interaction Network Assembly. J Prot Research. 2009;8:2944-2952. Abstract
  9. Mosley AL, Pattenden SG, Carey M, Venkatesh S, Gilmore JM, Florens L, Workman JL, Washburn MP. Rtr1 is a CTD phosphatase that regulates RNA polymerase II during the transition from serine 5 to serine 2 phosphorylation. Mol Cell. 2009;34:168-178. Abstract
  10. Mosley AL, Florens L, Wen Z, Washburn MP. A label free quantitative proteomic analysis of the Saccharomyces cerevisiae nucleus. J Proteomics. 2009;72:110-120. Abstract
  11. Lin CH, Li B, Swanson S, Zhang Y, Florens L, Washburn MP, Abmayr SM, Workman JL. Heterochromatin Protein 1a stimulates histone H3 lysine 36 demethylation by the DrosophilaKDM4A demethylase. Mol Cell. 2008;32:696-706. Abstract
  12. Yao T, Song L, Jin J, Cai Y, Takahashi H, Swanson SK, Washburn MP, Florens L, Conaway RC, Cohen RE, Conaway JW. Distinct modes of regulation of the Uch37 deubiquitinating enzyme in the proteasome and in the Ino80 chromatin-remodeling complex. Mol Cell. 2008;31:909-917.Abstract
  13. Suganuma T, Gutierrez JL, Li B, Florens L, Swanson SK, Washburn MP, Abmayr SM, Workman JL. ATAC is a double histone acetyltransferase complex that stimulates nucleosome sliding. Nat Struct Mol Biol. 2008;15:364-372. Abstract
  14. Pavelka N, Fournier ML, Swanson SK, Pelizzola M, Ricciardi-Castagnoli P, Florens L, Washburn MP. Statistical Similarities between Transcriptomics and Quantitative Shotgun Proteomics Data. Mol Cell Proteomics. 2008;7: 631-644. Abstract
  15. Sardiu ME, Cai Y, Jin J, Swanson SK, Conaway RC, Conaway JW, Florens L, Washburn MP. Probabilistic assembly of human protein interaction networks from label-free quantitative proteomics. Proc Natl Acad Sci U S A. 2008;105:1454-1459. Abstract
  16. Lee JS, Shukla A, Schneider J, Swanson SK, Washburn MP, Florens L, Bhaumik SR, Shilatifard A. Histone crosstalk between H2B monoubiquitination and H3 methylation mediated by COMPASS. Cell.2007;131:1084-1096. Abstract
  17. Cai Y, Jin J, Yao T, Gottschalk AJ, Swanson SK, Wu S, Shi Y, Washburn MP, Florens L, Conaway RC, Conaway JW. YY1 functions with INO80 to activate transcription. Nat Struct Mol Biol. 2007;14:872-874. Abstract
  18. Paoletti AC, Parmely TJ, Tomomori-Sato C, Sato S, Zhu D, Conaway RC, Conaway JW, Florens L, Washburn MP. Quantitative proteomic analysis of distinct mammalian Mediator complexes using normalized spectral abundance factors. Proc Natl Acad Sci U S A. 2006;103:18928-18933.Abstract
  19. Zybailov B, Mosley AL, Sardiu ME, Coleman MK, Florens L, Washburn MP.Statistical analysis of membrane proteome expression changes in Saccharomyces cerevisiae. J Proteome Res. 2006;5:2339-2347. Abstract
  20. Carrozza MJ, Li B, Florens L, Suganuma T, Swanson SK, Lee KK, Shia WJ, Anderson S, Yates J, Washburn MP, Workman JL. Histone H3 Methylation by Set2 Directs Deacetylation of Coding Regions by Rpd3S to Suppress Spurious Intragenic Transcription. Cell. 2005;123:581-592.Abstract
  21. Zybailov B, Coleman MK, Florens L, Washburn MP. Correlation of relative abundance ratios derived from peptide ion chromatograms and spectrum counting for quantitative proteomic analysis using stable isotope labeling.Anal Chem. 2005;77:6218-6224. Abstract
  22. Kusch T, Florens L, Macdonald WH, Swanson SK, Glaser RL, Yates III JR, Abmayr SM, Washburn MP, Workman JL. Acetylation by Tip60 Is Required for Selective Histone Variant Exchange at DNA Lesions. Science. 2004;306:2084-2087. Abstract
  23. Sato S, Tomomori-Sato C, Parmely TJ, Florens L, Zybailov B, Swanson SK, Banks CA, Jin J, Cai Y, Washburn MP, Conaway JW, Conaway RC. A set of consensus Mammalian mediator subunits identified by multidimensional protein identification technology. Mol Cell. 2004;14:685-691. Abstract
  24. Washburn MP, Wolters D, Yates JR III. Large-scale analysis of the yeast proteome via multidimensional protein identification technology. Nat Biotech. 2001;19:242-247. Abstract

 

    Last modified: Mar 14, 2014

    Contact

    Michael P Washburn, PhD
    Associate Professor, Department of Pathology and Laboratory Medicine

    Stowers Institute for Medical Research
    1000 East 50th Street
    Kansas City, MO 64110

    mpw@stowers.org

    ID=x10299