Mutation-Specific Calcium Dysregulation in Troponin T Linked Hypertrophic Cardiomyopathy

Abstract

Cardiac troponin T (cTnT) is a protein of the cardiac thin filament (CTF) and assists in conferring calcium regulation to muscle contraction. Genetic mutations in cTnT often cause hypertrophic cardiomyopathy (HCM), a disease affecting 1/200-1/500 people worldwide, and are known to cause differential disease severity and phenotypic variability. This study focused on six HCM-causing, highly penetrant mutations located within the cTnT N-terminus (R94H/C, R92L/W/Q, and I79N) which are each associated with distinct phenotypes and severities in human patients. In the first aim, we determined the effects of HCM-causing mutations in cTnT on the calcium-based regulation of muscle activation across varying in vitro systems of increasing biological complexity using a collection of steady-state and kinetic measurements. Results showed that cTnT mutations can cause similar changes to calcium sensitivity via distinct combinatorial changes to the kinetics of thin filament calcium association and release and were used to generate a binning scheme representing potential drug-targetable groups. In the second aim, we utilized a four-state mathematical model of cardiac troponin C (cTnC) activation to simulate the results obtained towards aim 1. We systematically applied changes to the model in accordance with each bin to successfully simulate the data for each mutation, thereby providing a bin-specific biophysical mechanism observed effects on calcium handling at the cTn core. Results from these simulations suggested the effects of mutations within the cTnT N-terminus may affect cardiac troponin I (cTnI) switch peptide and cTnC interactions. In the last aim, we tested the generalizability of a proposed mechanistic link between accelerated thin filament calcium release and abnormal CaMKIIδ activity via the generation and characterization of two novel, independent mouse models of mixed cardiomyopathy. Despite demonstrating progressive systolic dysfunction and mutation-specific phenotypes, results suggested that CaMKIIδ activity was normal throughout disease progression in both models. Further studies are necessary to clarify the role of CaMKIIδ signaling as a nodal point in cTnT-linked HCM.