FUNCTIONAL EFFECTS OF THE H1-HELIX AND THE FLEXIBLE-LINKER OF CARDIAC TROPONIN T IN HEALTH AND DISEASE
MICHAEL, JOHN JESHURUN
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Mutations in cardiac contractile proteins are associated with heart diseases, but it is not understood how such alterations to proteins cause contractile dysfunction. Force generation and contraction of the heart is mediated by coordinated interaction between the thick and thin filament proteins. The thick filament proteins mainly consist of myosin, whereas, the thin filament proteins include actin, tropomyosin (Tm), and the troponin (Tn) complex. Troponin T (TnT), troponin C (TnC), and troponin I (TnI) constitute the Tn complex. The strategic position of TnT allows it to regulate Ca2+-, muscle length (ML)-, and myosin crossbridge (XB)-mediated contractile activation. The N-terminal region of TnT (~residues 1-180) partially interacts with Tm and the C-terminal region (~residues 181-290) interacts with TnC and TnI; both regions are linked by two pivotally positioned motifs: the H1-helix (~205 -221 residues) and the flexible linker (~185-202). There is a lack of knowledge how disease-related or isoform-specific structural changes to these motifs affects cardiac contractile function. Therefore, the overall objective of this work is to determine the role of the H1-helix and the flexible-linker of TnT in regulating Ca2+-, XB-, and ML-mediated activation of the cardiac myofilament in health and cardiac disease. The central hypothesis is that either disease-related and isoform-specific structural changes to the H1-helix and the flexible-linker of TnT impact cardiac contractile function. We tested our hypothesis by measuring the contractile function of cardiac fibers containing recombinant TnT variants, carrying either disease-related or isoform-specific structural changes in these motifs. Specific Aim 1 was designed to determine how disease-related structural alterations to these motifs in TnT affect cardiac contractile function. Specific Aim 2 was designed to determine how altering the cardiac-specific structure of these motifs in TnT affects cardiac contractile function. Our findings substantiated our hypothesis because disease-related and isoform-specific structural changes to the H1-helix and the flexible-linker of TnT indeed impaired cardiac contractile function. This work is anticipated to contribute mechanistic insights into functioning of the cardiac muscle at the myofilament level which is fundamental for developing therapies to treat cardiac contractile dysfunction.