Functional Modulation and Adaptive Responses to PKC Targets on Cardiac Troponin I.

[摘要] Cardiac pump performance is highly dynamic and the rhythmic contraction and relaxation during a single heart beat is produced by myofilaments composed of highly ordered thin and thick filaments arranged in sarcomeres within each cardiac myocyte.The thin filament, molecular switch protein cardiac troponin I (cTnI) toggles between actin and the Ca2+-binding subunit, troponin C (cTnC) to regulate muscle contraction.Cellular signaling via the protein kinase C (PKC) pathway, modulates contractile function and phosphorylates cTnI on serines (Ser)23/24, Ser43/45, and threonine (Thr)144. While the functional effects of cTnISer23/24 phosphorylation are well characterized, the role of cTnISer43/45 remains controversial. During cardiac dysfunction, cTnISer43/45 phosphorylation increases, but it is unclear whether this phosphorylation improves or reduces function. Gene transfer into intact cardiac myocytes is used in the present study to examine the functional role of cTnISer43/45 and reconcile divergent in vitro and in vivo responses.Phospho-null (Ala) substitution at cTnISer43/45 diminished myocyte shortening amplitude and myofilament Ca2+ sensitivity of tension, indicating Ala substitution is not functionally conservative at this site.However, cTnISer43/45 phospho-mimetic (Asp) substitutions produced the anticipated decrease in basal contractile function and myofilament Ca2+ sensitivity 2 days after gene transfer into myocytes.Moreover, cTnISer43Asp and cTnISer45Asp individually modulated function.Interestingly, the slowed shortening and re-lengthening rates returned toward control values after more extensive sarcomeric cTnI replacement at day 4. Further studies demonstrated adaptations within myocytes, including accelerated Ca2+ re-uptake into the sarcoplasmic reticulum and increased sarcomeric protein phosphorylation.Overall, these adaptations are proposed to communicate and fine-tune the modulatory effects of cTnISer43/45 phosphorylation and/or phospho-mimetic substitution.This communication was further examined with Asp substitutions at cTnI Ser23/24 and Ser43/45 (e.g. cTnIAspQuad).Studies with cTnIAspQuad showed Ser23/24 and Ser43/45 modulate function through separate mechanisms, Ser23/24 enhances the functional response to Ser43/45,and cTnIAspQuad induces a different adaptive signaling response than cTnISer43/45Asp. Overall, the present experiments identify cTnISer43/45 phosphorylation as a modulatory brake in the heart, which is fine-tuned by adaptive signaling. This modulatory brake is predicted to play an important role during sustained stress and may contribute to heart failure with the loss of adaptive signaling in cardiac myocytes.

[发布机构] University of Michigan

[效力级别] Protein Kinase C

[关键词] Heart;Protein Kinase C;Cardiac Troponin I;Molecular;Cellular and Developmental Biology;Science;Cellular & Molecular Biology