Proteome Research

The human genome has been sequenced and now sophisticated technologies are used to characterize the proteome, the set of proteins generated by that genome, as a function of developmental stage, environment, tissue-type, and pathology. The goal of our research is to understand, on a systems level, the complex biological processes that are fundamental to physiology and disease. A particular focus of our laboratory is cancer research (ovarian and breast cancer) and vascular/cardiovascular biology.

Our laboratory develops and applies powerful tools of proteomics and bioinformatics to analyze the protein complement in protein complexes, cells, tissues and organisms. Our goal is not only to determine the level of proteins but also their localization, dynamics, and interaction partners. In recent years we have developed sensitive methodologies to isolate specific subcellular fractions from mammalian cell culture or tissues.

Currently our focus is on optimizing technologies that allow for the sensitive and selective isolation and characterization of surface, plasma membrane proteins from cell culture (in vitro) or directly from the surface of microvascular endothelial cells (in vivo). The ultimate goal of these studies is to investigate the proteome dynamics of surface membrane proteins in the vasculature of disease models or in cell culture in response to specific treatments. Additionally, we are interested in dynamics of proteinprotein interactions. To accomplish this goal we are applying tandem affinity purifications (TAP-tagging) in combination with mass spectrometry. Briefly, the protein of interest (bait) is expressed as a fusion protein with a dual affinity tag. This allows for mild and efficient purification of this protein and most of its interacting partners.

Our laboratory is well equipped and currently has fulltime access to two latest generation mass spectrometers, a linear ion-trap (Thermo Scientific LTQ) and the new Thermo Scientific LTQ-Orbitrap XL, a high resolution, high mass accuracy hybrid mass spectrometer. These mass spectrometers are currently the standard for global, high-throughput proteomics. Additionally, we have access to a new triple quadrupole mass spectrometer for target driven proteomics and accurate quantification.

Graduate Students:

• Lusia Sepiashvili (co-supervised)

Selected References:

• Gortzak-Uzan L, Ignatchenko A, Evangelou A, Agochiya M, Brown KA, St.Onge P, Kireeva I, Schmitt-Ulms G, Brown TJ, Rosen B, Shaw P, Jurisica I, & Kislinger T. A Proteome resource of ovarian cancer ascites: integrated proteomic and bioinformatic analyses to identify putative biomarkers. J Proteome Research 2007 (in press)

• Kislinger T, Cox B, Kannan A, Chung C, Hu P, Ignatchenko A, Scott SS, Gramolini AO, Morris Q, Hallett MT, Rossant J, Hughes TR, Frey B, & Emili A. Global Survey of Organ and Organelle Protein Expression in Mouse: Combined Proteomic and Transcriptomic Profiling. Cell 2006, 125: 173-186

• Kislinger T, Gramolini AO, Pan Y, Rahman K, MacLennan DH, & Emili A. Proteome dynamics during C2C12 myoblast differentiation. Mol Cell Proteomics 2005; 7: 887-901

• Kislinger T, Rahman K, Radulovic D, Cox B, Rossant J, & Emili A. PRISM: A generic large-scale proteomics investigation strategy for mammals. Mol Cell Proteomics 2003; 2:96-106