# Project 3: Molecular Mechanisms of Myofilament Dysfunction in Heart Failure

> **NIH NIH P01** · UNIVERSITY OF ILLINOIS AT CHICAGO · 2020 · $410,839

## Abstract

Despite aggressive treatment, heart failure (HF) remains a major clinical problem with high mortality. HF is
characterized by depressed cardiac pump function. Recent studies have shown that depressed contractile
protein function underlies, in part, depressed pump function in HF; however, the molecular mechanisms are
largely unknown. Studies proposed in project 3 emphasize “Control of Sarcomere Dynamics”, with a strong
focus on cardiac myosin light chain 2 (cMLC2) phosphorylation. Strong synergistic interactions of project 3
continue with project 1 (biased ligand signaling targeting sarcomeres) and project 2 (sarcomere growth and
remodeling). In addition project 3 relies critically on the support from Core B (Human Tissue; aims 1 and 2) and
Core C (Proteomics and Analytical Biochemistry; aim 3). The significance of cMLC2 phosphorylation was
substantially enhanced by the recent identification of a cardiac specific isoform of myosin light chain kinase
(cMLCK), which is upregulated in HF. However, both the extent and regional variation of cMLC2
phosphorylation in health and disease, as well as the functional impact of this post-translational modification on
cardiac contractile biology has not been studied in detail. Accordingly, the overall goal of project 3 is to define
the role of cMLC2 on human myofilament function in health and disease. We hypothesize that human cMLC2
is a critical regulator of myocyte contraction that is dysregulated in HF, and that increasing MLC2
phosphorylation will reverse myofilament dysfunction and improve cardiac pump function. To test this
hypothesis, we have developed three independent specific aims. In aim 1 we will determine the role of cMLC2
in skinned myocardium from healthy and failing human hearts. Biophysical experiments will focus on skinned
human isolated myocardium where we will alter contractile protein phosphorylation status by treatment with
kinases or phosphatase, or recombinant protein exchange. In aim 2 we will determine the role of cMLC2 in
healthy and diseased intact cells under simulated length-force relationships that approximate in-situ pressure-
volume loops. Finally, in aim 3 we will determine the extent of cMLC2 biochemical alterations by using state of
the art proteomic analysis. Specific attention will be on MLC2 phosphorylation on S15 as well as the extent of
N14 deamidation, and the regional variation of this parameter. We expect to identify decreased cMLC2
phosphorylation as a significant mechanism underlying sarcomeric dysfunction in heart failure. Our results may
provide a path to a novel therapeutic target in the treatment of heart failure, where restoration of cardiac
myosin light chain 2 phosphorylation levels is expected to improve cardiac pump performance.

## Key facts

- **NIH application ID:** 9941113
- **Project number:** 5P01HL062426-20
- **Recipient organization:** UNIVERSITY OF ILLINOIS AT CHICAGO
- **Principal Investigator:** Pieter P. de TOMBE
- **Activity code:** P01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $410,839
- **Award type:** 5
- **Project period:** — → 2023-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9941113

## Citation

> US National Institutes of Health, RePORTER application 9941113, Project 3: Molecular Mechanisms of Myofilament Dysfunction in Heart Failure (5P01HL062426-20). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/9941113. Licensed CC0.

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