The NHLBI Johns Hopkins University Proteomics Innovation Center in Heart Failure

Overview

Rationale:

The Johns Hopkins University (JHU) proposal applies state-of-the-art proteomic methods and develops new approaches and techniques to investigate the biological and clinical aspects of heart failure (HF).  HF is a clinical syndrome in which cardiovascular function is insufficient to support the metabolic needs of the body.  On a population basis, its burden is substantial and increasing with 5-6 million people affected in the United States.  Several factors contribute to the etiology and evolution of the clinical presentation including ischemia, hypertension, diabetes and genetic predisposition. Regardless of therapeutic efforts, mortality and morbidity remains high.  Many gaps in understanding HF remain and it is the goal of this center to identify how HF impacts signaling cascades and several subproteomes (mitochondrial, contractile, cell surface and secretory) emphasizing post-translational modifications (PTMs) in order to discover novel ways by which protein modifications either contribute to disease or could be targeted to improve disease outcome.

Philosophy and goals:

 The central philosophy of our center is that innovative technologies in proteomics will be essential to pursue compelling biological and clinical questions. Building on our past successes in developing innovative technologies, we will broaden our developmental scope and focus on JHU’s clinical and research strength in HF. A high level overview is presented below.

Technical challenges to be addressed:

1. Exploitation and continued development of proteomic methods for the global investigation of a wide range of biologically important PTMs with the explicit goals of identifying and quantifying each modified amino acid residue. By achieving this we can explore the dynamic nature of the proteome and the crosstalk between signaling cascades and the end effector molecules.  Specific emphasis is placed on:
   
   
   1. Phosphorylation and its dynamic interaction with O-GlcNAcylation.
   2. Cys and Lys residue modifications including S-nitrosylation, disulfide bonds and acetylation.
   3. The cell surface and secretory subproteome and N-linked glycosylation.

2. Development of workflows for improved separation and enrichment of proteins and peptides, as well as, mass spectrometry methods to extend detection and quantification to the full dynamic range observed in tissue and body fluids.
3. Innovation in bioinformatics focused on automated solutions for protein name redundancy, comparisons/querying, PTM iteration for both improved protein identification and maximized protein characterization (isoforms, PTMs).  Including the development of accessible databases for human cardiac proteins and their PTMs and CardioMRM, a peptide database specific for the cardiac proteome.  The bioinformatics core is designed to improve integration between projects and to help to emphasize targets for further study.
4. Integration of sophisticated cell modeling with quantitative proteomic data and functional endpoints to provide mechanistic insights into the interplay between subproteomes of the human cardiac myocyte.

Technical application to biological and clinical challenges. Using defined experimental animal models and myocardial and blood samples from patients with HF we will:

1. Reveal the full dynamic range and interconnection of the intracellular signaling proteome.
2. Assess established and novel PTMs of proteins of the contractile system.
3. Assess the plasma membrane proteome and its subdomains as relevant to HF.
4. Pursue plasma biomarker identification for the risk stratification of HF patients by i) exploiting our newly developed PTM-enrichment methods for de novo discovery and ii) targeting and quantifying these and other specific disease-associated PTMs and unique protein isoforms of known or candidate markers. 

Summary: 

The exceptional group of scientists comprising the JHU NHLBI Proteomic Center will continue developing and disseminating new tools and proteomic technologies focused on central questions relevant to HF.  Our strengths lie not only in our excellence in proteomics (analytical approaches, bioinformatics, systems modeling and range of functional assays), but also in the breadth of animal models and unique human samples brought to the group by world class cardiologists, epidemiologists and physiologists.  The highly collaborative and integrated nature of this group of investigators allows us to keep focused on clinically-relevant problems and provide rapid paths to translational medicine.  Given our concerted approach, our center will push the frontiers forward providing accelerated intellectual payoffs and innovation in technological and data analysis resources for the broader scientific community.  The application of state-of-the-art and truly innovative methods, coupled equally with innovative biology, is the cornerstone of the JHU proposal.

Technology Innovation:

Phosphorylation
O-GlcNAc and Crosstalk with Phosphorylation
Cys and Lys Modifications in Heart
Cell Surface and the Secretory Pathway

Biological/Clinical Applications:

G-coupled Signaling in Heart Failure
Myofilament in Diabetic Cardiomyopathy
HIF-1α Regulation in Heart Failure
Mitochondria Acetylation
Biomarker Development for Heart Failure Risk Stratification

This project is funded by the National Heart, Lung, and Blood Institute