Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure.
Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure.
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DOI：
10.1161/circresaha.116.308964
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发表时间：
2016-09-30
影响因子：
20.1
通讯作者：
Xiang YK
中科院分区：
文献类型：
--
作者：
Barbagallo F;Xu B;Reddy GR;West T;Wang Q;Fu Q;Li M;Shi Q;Ginsburg KS;Ferrier W;Isidori AM;Naro F;Patel HH;Bossuyt J;Bers D;Xiang YK
在心力衰竭中，肌丝蛋白表现出异常磷酸化，从而导致收缩功能障碍。肌丝上蛋白质磷酸化失调的机制尚不清楚。本研究旨在了解心力衰竭中肌丝蛋白磷酸化改变的机制。我们生成了一种新型基因编码蛋白激酶 A (PKA) 生物传感器，锚定在兔心肌细胞的肌丝上，以检查肌丝对肾上腺素刺激的反应中的 PKA 活性。我们发现，在肥大衰竭的兔肌细胞中，PKA 活性从肌膜转移到肌丝。特别是，肌丝上 PKA 活性的增加是由于选择性定向至肌丝的 β2 肾上腺素受体 (β2AR) 信号增强以及与肌原纤维相关的磷酸二酯酶活性降低所致。从机制上讲，肌丝上 PKA 活性的增强与肥大衰竭兔肌细胞中 Caveolin-3 的下调有关。在衰竭的肌细胞中重新引入caveolin-3能够通过阻止PKA信号进入肌丝来使β2AR信号的分布正常化，并恢复对肾上腺素刺激的收缩反应。在肥大的兔肌细胞中，选择性增强针对肌丝的 β2AR 信号传导有助于提高肌丝蛋白的 PKA 活性和 PKA 磷酸化。重新引入 Caveolin-3 能够限制 β2AR 信号传导并恢复肌细胞响应 β-肾上腺素能刺激的收缩性。
In heart failure, myofilament proteins display abnormal phosphorylation, which contributes to contractile dysfunction. The mechanisms underlying the dysregulation of protein phosphorylation on myofilaments is not clear. This study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure. We generate a novel genetically encoded protein kinase A (PKA) biosensor anchored onto the myofilaments in rabbit cardiac myocytes to examine PKA activity at the myofilaments in responses to adrenergic stimulation. We show that PKA activity is shifted from the sarcolemma to the myofilaments in hypertrophic failing rabbit myocytes. In particular, the increased PKA activity on the myofilaments is due to an enhanced β2 adrenergic receptor (β2AR) signal selectively directed to the myofilaments together with a reduced phosphodiesterase activity associated with the myofibrils. Mechanistically, the enhanced PKA activity on the myofilaments is associated with downregulation of caveolin-3 in the hypertrophic failing rabbit myocytes. Reintroduction of caveolin-3 in the failing myocytes is able to normalize the distribution of β2AR signal by preventing PKA signal access to the myofilaments, and to restore contractile response to adrenergic stimulation. In hypertrophic rabbit myocytes, selectively enhanced β2AR signaling toward the myofilaments contributes to elevated PKA activity and PKA phosphorylation of myofilament proteins. Reintroduction of caveolin-3 is able to confine β2AR signaling and restore myocyte contractility in response to β-adrenergic stimulation.