Advanced targeted mass spectrometry technologies for discovery of protein isoforms that are drivers of Coronary Artery disease (CAD)

• Grant number: 22/16606-4
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• Start date: June 01, 2023
• End date: May 31, 2024
• Field of knowledge: Biological Sciences - Physiology - Physiology of Organs and Systems
• Principal Investigator: Ana Maria de Lauro Castrucci
• Grantee: José Thalles Jocelino Gomes de Lacerda
• Supervisor: Gloria M. Sheynkman
• Host Institution: Instituto de Biociências (IB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Institution abroad: University of Virginia (UVa), United States
• Associated to the scholarship: 20/04524-8 - Integrating proteogenomics approach and system biology to investigate the role of melanopsin and TRP channels in the circadian rhythmicity, BP.PD

Abstract

The genetic processes that give rise to phenotypic plasticity in complex organisms are primarily orchestrated by alternative splicing (AS), a mechanism by which genetic variants (e.g., single-nucleotide polymorphisms - SNPs located within non-coding genomic regions), in cis, directly influence the alternatively spliced transcripts produced from a gene. Nearly every human gene is alternatively spliced and thus gives rise to diverse phenotypes, particularly across human ethinic population. The product of alternative splicing, protein isoforms (or proteoforms), are central elements in nearly all biological systems and play a central role in modulating complex traits. Even so, functionally distinct proteoforms resulting from AS are poorly understood in terms of their disease-related functions, information that is desperately needed for rationally designed therapies and candidate markers of disease. Coronary artery disease (CAD) is one of the leading global causes of mortality, where there are large heritability estimates for CAD indicating strong associated genetic variants. Notably, vascular smooth endothelial cells (VSMCs) play critical roles in CAD susceptibility and its link to atherosclerosis. In this project, we will seek to establish a mass spectrometry (MS)-based multilevel analytical workflow to thoroughly characterize the CAD-relevant protein isoforms in VSMCs. This effort will discover distinct isoforms associated with the CAD phenotype and uncover deeper knowledge about the splice-related molecular etiology of CAD. We will conduct the study through the collection of VSMCs gathered from 151 multiethnic heart transplant donors, which will be cultured under quiescent (healthy) or proliferative (atherogenic) conditions. First, we will characterize the molecular landscape of CAD through a long-read proteogenomics approach and perform real-time monitoring of peptides derived from spliced protein isoforms. Then, we will improve the identification and quantification of spliced protein isoforms related to CAD through multiplexed targeted proteomics. This project will provide an in-depth look into the impact of splicing on protein isoform diversity for CAD-related phenotypes, highlighting the clinical relevance of spliced protein isoforms for biomedical discoveries, including potential candidate biomarkers and drug targets for preventive and personalized medicine. (AU)