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N‐terminal phosphorylation of cardiac troponin‐I reduces length‐dependent calcium sensitivity of contraction in cardiac muscle
Author(s) -
Rao Vijay S.,
Korte F. Steven,
Razumova Maria V.,
Feest Erik R.,
Hsu Hsiaoman,
Irving Thomas C.,
Regnier Michael,
Martyn Donald A.
Publication year - 2013
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1113/jphysiol.2012.241604
Subject(s) - cardiac muscle , contraction (grammar) , calcium , troponin , cardiology , phosphorylation , medicine , muscle contraction , troponin i , myofilament , chemistry , myocyte , biochemistry , myocardial infarction
Key points• β‐Adrenergic stimulation is an important control mechanism, matching cardiac output to venous return during increased metabolic demand. • β‐Adrenergic signalling leads to protein kinase A (PKA) phosphorylation of myofilament proteins cardiac troponin I (cTnI), cardiac myosin binding protein‐C (cMyBP‐C) and titin, but their specific effects on the sarcomeric length (SL) dependence of contraction – which underlies the Frank–Starling Law of the Heart – is debated. • Recombinant cTnI phosphomimetics were exchanged into cardiac muscle to isolate the effects of cTnI from those of cMyBP‐C/titin phosphorylation on SL‐dependent force–Ca 2+ relations and sarcomeric structure. • Results suggest cTnI or cMyBP‐C/titin phosphorylation, separately or together, eliminate the SL dependence of Ca 2+ sensitivity of force, but not maximal force. The reduction occurs particularly at long SL, suggesting effects on thin filament access and crossbridge recruitment. • The net effect of PKA phosphorylation is to blunt SL dependence of force at submaximal [Ca 2+ ] to maintain elevated systolic function.Abstract Protein kinase A (PKA) phosphorylation of myofibrillar proteins constitutes an important pathway for β‐adrenergic modulation of cardiac contractility. In myofilaments PKA targets troponin I (cTnI), myosin binding protein‐C (cMyBP‐C) and titin. We studied how this affects the sarcomere length (SL) dependence of force– p Ca relations in demembranated cardiac muscle. To distinguish cTnI from cMyBP‐C/titin phosphorylation effects on the force– p Ca relationship, endogenous troponin (Tn) was exchanged in rat ventricular trabeculae with either wild‐type (WT) Tn, non‐phosphorylatable cTnI (S23/24A) Tn or phosphomimetic cTnI (S23/24D) Tn. PKA cannot phosphorylate either cTnI S23/24 variant, leaving cMyBP‐C/titin as PKA targets. Force was measured at 2.3 and 2.0 μm SL. Decreasing SL reduced maximal force ( F max ) and Ca 2+ sensitivity of force ( p Ca 50 ) similarly with WT and S23/24A trabeculae. PKA treatment of WT and S23/24A trabeculae reduced p Ca 50 at 2.3 but not at 2.0 μm SL, thus eliminating the SL dependence of p Ca 50 . In contrast, S23/24D trabeculae reduced p Ca 50 at both SL values, primarily at 2.3 μm, also eliminating SL dependence of p Ca 50 . Subsequent PKA treatment moderately reduced p Ca 50 at both SLs. At each SL, F max was unaffected by either Tn exchange and/or PKA treatment. Low‐angle X‐ray diffraction was performed to determine whether p Ca 50 shifts were associated with changes in myofilament spacing ( d 1,0 ) or thick–thin filament interaction. PKA increased d 1,0 slightly under all conditions. The ratios of the integrated intensities of the equatorial X‐ray reflections ( I 1,1 / I 1,0 ) indicate that PKA treatment increased crossbridge proximity to thin filaments under all conditions. The results suggest that phosphorylation by PKA of either cTnI or cMyBP‐C/titin independently reduces the p Ca 50 preferentially at long SL, possibly through reduced availability of thin filament binding sites (cTnI) or altered crossbridge recruitment (cMyBP‐C/titin). Preferential reduction of p Ca 50 at long SL may not reduce cardiac output during periods of high metabolic demand because of increased intracellular Ca 2+ during β‐adrenergic stimulation.