Decoding Nuclear Mechanosignaling and Epigenetic Mediators of Mechanical Memory in Cardiomyocytes

About This Grant

Project summary Mechanotransduction is the process by which cells convert external mechanical signals into biochemical signals that shape their phenotypic adaptations. In cardiomyocytes, short-term extracellular stiffening induces readily reversible phenotypic adaptations, while sustained exposure to extracellular stiffening induces persistent changes in cellular structure and chromatin architecture: a phenomenon referred to as 'mechanical memory' (MM). We recently reported that stabilization of microtubule (MT) architecture is required for both the formation and maintenance of MM in cardiomyocytes. In this proposal, I focus on the time-dependent nuclear responses to extracardiac stiffening, including changes in chromatin architecture, gene expression and DNA damage responses. My working hypothesis is that DNA damage is a central component of persistent responses to extracellular stiffening and disease-relevant mechanical stresses. In Aim 1, I will determine the molecular conduits and epigenetic regulators of MM using normal adult cardiomyocytes and a novel cell-culture system with bidirectionally tunable stiffness. After defining the temporal and magnitude thresholds for inducing MM, I will perform a whole genome ATAC-seq to determine the distinct epigenetic landscapes associated with transience vs. persistence of the stiffness-induced phenotype in cardiomyocytes (K99 phase). Building on this foundation, I will then determine the role of the DNA damage response (DDR) in determining the reversibility of nuclear responses to extracellular stiffening. These studies will define whether DDR elements may be targets for therapeutics to limit or reverse MM in CMs. In Aim 2 studies, I will translate these experiments to the tissue level using living myocardial slices (LMS) from normal rat and human hearts. I will determine whether pathomimetic increases in afterload evoke the same time-dependent MM responses, mechanotransduction cascades, and DNA damage signals observed in isolated cardiomyocyte (K99 phase). I will then examine whether interventions targeting MT dynamics and the DDR mechanism will attenuate afterload induced MM (R00 phase). Finally, in vivo studies will explore whether time-dependent MM dynamics, and mitigating strategies, are relevant to the myocardial dysfunction observed in the viable myocardium following a large myocardial infarction(R00 phase). Through the proposed work, I will significantly expand my expertise and facility with several powerful and versatile skills (RNA-seq, ATAC seq, the LMS model, and rodent experimentation) while broadening my ability to work with large datasets and manage a multifaceted research program. During this process, I will pursue interactions and scientific connections with centers and collaborators within and outside my institution that will contribute to the advancement and completion of this work and prepare me for success as an independent research scientist.