Abstract 1446: Role of the Conserved Acidic N-terminus of Cardiac Troponin I in Regulating Myocardial Function
Background: Cardiac Troponin I (cTnI) phosphorylation plays a major role in regulating myocardial contractility and relaxation during β-adrenergic stimulation. The role of the conserved acidic N-terminus of cTnI in the modulation of cardiac contractility is not clear although NMR and modeling studies have shown that this region can extend from its position on the N-lobe of cardiac troponin C (cTnC) and interact with the basic inhibitory region of cTnI.
Hypothesis: The acidic N-terminus of cTnI is necessary to regulate myocardial function.
Methods: We generated cardiac-specific transgenic mice in which the first 10 amino acids of cTnI (cTnIΔ2–11) were deleted. To determine the effect(s) of this deletion, in vitro phosphorylation, Ca2+ dependent Mg2+-ATPase activity and maximal force were measured with and without protein kinase A. To map the acidic N-terminus of cTnI on the N-lobe of cTnC, we compared 1H-15N chemical shifts from 1H-15N HSQC spectra collected from [15N,2H]cTnC complexed with either cTnI or cTnIΔ2–11.
Results: Replacement of cTnI with cTnIΔ2–11 was >95% but was benign in that no hypertrophic response or pathology resulted. The cTnIΔ2–11 hearts displayed significantly decreased contraction and relaxation under basal and β-adrenergic stress compared to non-transgenic hearts, with a reduction in maximal Mg2+-ATPase activity and maximal Ca2+ dependent force. However, Ca2+ sensitivity of force development and phosphorylation at Ser23/24 was not affected. Electron microscopic studies revealed an altered sarcomeric structure: the length of I-bands in cTnIΔ2–11 sarcomeres were decreased by 50% (0.15 ± 0.03 μm) as compared to non-transgenic sarcomeres (0.31 ± 0.01 μm), with no changes in A-band length. Chemical shift difference mapping suggested that, in the absence of phosphorylation, residues 2–11 of cTnI do not strongly interact with cTnC and likely do not directly affect Ca2+ sensitivity. In conclusion, the data show that the acidic N-terminus of the cTnI plays an important role in regulating myocardial contractility and sarcomeric structure, and is necessary in mediating the heart’s response to β-adrenergic stimulation.