Kunhong Xiao - US grants

Introduction: Changes in protein expression and post-translational modifications (PTMs) are essential mechanisms for biological regulation in normal physiology and numerous diseases, including those of the cardiovascular disease [1-4]. Characterizing these changes can provide valuable information for the elucidation of disease etiology, identification of clinically useful biomarkers, and development of novel therapeutics. Thus, methods for the identification of proteins and characterization of protein PTMs are essential to modern biomedical research. In recent years, mass spectrometry (MS)-based proteomics has become the technology of choice for these purposes. MS-based proteomics takes advantage of the modern mass spectrometer's superior resolution power and accuracy in peptide sequencing [5]. It allows for the rapid, large-scale identification and quantification of proteins and their PTMs in multiprotein complexes, whole cells, tissues and organisms with sub-femto mole level sensitivity (100-1000 times more sensitive than traditional technologies). Recently, MS combined with stable isotope labeling technologies (i.e. quantitative proteomics) has emerged as a powerful tool to quantitatively assess dynamic changes in protein expression, subcellular compartmentalization and PTMs on a proteome-wide scale [6]. Therefore, MS-based proteomics is unequaled as a tool for studying complex biological systems and disease in the post-genomic era. All these various tools are particularly applicable to cardiovascular research and should allow us to carry out the goals of this PPG. The ability to deliver genes efficiently to cultured cardiomyocytes and in vivo to the rodent heart is critical to many of the experiments described in the three PPG projects. Viral vectors offer greater transduction efficiency to cultured cardiomyocytes than nonviral methodology such as plasmid DNA delivery via liposomal reagents, electroporation or nucleofector techniques [7]. Similarly, viral vectors are more efficient than non-viral gene delivery methods in mediating gene delivery in vivo to the heart [8]. The Viral Vector Core will provide adenoviral and adeno-associated viral vector development, manufacturing, purification, and validation services to this PPG.

PROJECT SUMMARY / ABSTRACT: Core C ? Proteomics Core Mass spectrometry (MS)-based proteomics is unequaled as a tool for studying complex biological systems and diseases in the post-genomic era. In the past decade, Drs. Gygi and Xiao have worked together and established close collaborations with the PIs of this PPG (Drs. Lefkowitz, Rockman, Stamler and Koch) to take full advantage of MS-based proteomics as a powerful tool to study ?-adrenergic receptor and angiotensin II type I receptor signaling. These collaborations form the basis for the Proteomics Core C, which will continue to provide cutting-edge, high-throughput MS technologies to the investigators of this PPG. Four major functions of the Proteomics Core are: 1) To serve as a proteomics resource for the three projects of this PPG, offering state-of-the-art proteomics service and consultation. The Core will provide cutting-edge MS-based proteomics technologies for both single protein characterization and large-scale, high-throughput MS-based proteomics analysis to support the cardiovascular GPCR research in this PPG. With these services, the Core will function as a ?discovery engine? in order to intensify the innovation efforts of the overall research program. 2) To support translational research in this PPG by conducting proteomic studies of heart samples from different sources. 3) To provide MS-based structural tools, such as Hydrogen-Deuterium Exchange MS (HDXMS), to study protein conformations and dynamics. 4) To continue to educate investigators involved in this PPG in the field of proteomics research, thereby disseminating this powerful technology. Three major services that the Proteomics Core will provide to this PPG are: 1) Basic MS-based proteomics services for protein identification and protein PTM analysis (such as phosphorylation, S-nitrosylation, ubiquitination, glycosylation, and prolyl hydroxylation, etc.). 2) Advanced large-scale high-throughput MS-based proteomics services (such as qualitative and quantitative interactome and phosphoproteome analysis). 3) MS services for studying protein conformation and conformational dynamics (such as HDXMS)