Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure

Federica Barbagallo, Bing Xu, Gopireddy R. Reddy, Toni West, Qingtong Wang, Qin Fu, Minghui Li, Qian Shi, Kenneth S Ginsburg, William Ferrier, Andrea M. Isidori, Fabio Naro, Hemal H. Patel, Julie B C Bossuyt, Donald M Bers, Yang Kevin Xiang

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

RATIONALE:: 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. OBJECTIVE:: This study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure. METHODS AND RESULTS:: 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. CONCLUSIONS:: 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.

Original languageEnglish (US)
JournalCirculation Research
DOIs
StateAccepted/In press - Aug 31 2016

Fingerprint

Myofibrils
Biosensing Techniques
Cyclic AMP-Dependent Protein Kinases
Heart Failure
Rabbits
Caveolin 3
Muscle Cells
Adrenergic Receptors
Phosphorylation
Adrenergic Agents
Proteins
Sarcolemma
Phosphoric Diester Hydrolases
Cardiac Myocytes
Down-Regulation

ASJC Scopus subject areas

  • Physiology
  • Medicine(all)
  • Cardiology and Cardiovascular Medicine

Cite this

Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure. / Barbagallo, Federica; Xu, Bing; Reddy, Gopireddy R.; West, Toni; Wang, Qingtong; Fu, Qin; Li, Minghui; Shi, Qian; Ginsburg, Kenneth S; Ferrier, William; Isidori, Andrea M.; Naro, Fabio; Patel, Hemal H.; Bossuyt, Julie B C; Bers, Donald M; Xiang, Yang Kevin.

In: Circulation Research, 31.08.2016.

Research output: Contribution to journalArticle

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, JBC, Bers, DM & Xiang, YK 2016, 'Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure', Circulation Research. https://doi.org/10.1161/CIRCRESAHA.116.308964
Barbagallo, Federica ; Xu, Bing ; Reddy, Gopireddy R. ; West, Toni ; Wang, Qingtong ; Fu, Qin ; Li, Minghui ; Shi, Qian ; Ginsburg, Kenneth S ; Ferrier, William ; Isidori, Andrea M. ; Naro, Fabio ; Patel, Hemal H. ; Bossuyt, Julie B C ; Bers, Donald M ; Xiang, Yang Kevin. / Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure. In: Circulation Research. 2016.
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abstract = "RATIONALE:: 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. OBJECTIVE:: This study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure. METHODS AND RESULTS:: 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. CONCLUSIONS:: 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.",
author = "Federica Barbagallo and Bing Xu and Reddy, {Gopireddy R.} and Toni West and Qingtong Wang and Qin Fu and Minghui Li and Qian Shi and Ginsburg, {Kenneth S} and William Ferrier and Isidori, {Andrea M.} and Fabio Naro and Patel, {Hemal H.} and Bossuyt, {Julie B C} and Bers, {Donald M} and Xiang, {Yang Kevin}",
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T1 - Genetically Encoded Biosensors Reveal PKA Hyperphosphorylation on the Myofilaments in Rabbit Heart Failure

AU - Barbagallo, Federica

AU - Xu, Bing

AU - Reddy, Gopireddy R.

AU - West, Toni

AU - Wang, Qingtong

AU - Fu, Qin

AU - Li, Minghui

AU - Shi, Qian

AU - Ginsburg, Kenneth S

AU - Ferrier, William

AU - Isidori, Andrea M.

AU - Naro, Fabio

AU - Patel, Hemal H.

AU - Bossuyt, Julie B C

AU - Bers, Donald M

AU - Xiang, Yang Kevin

PY - 2016/8/31

Y1 - 2016/8/31

N2 - RATIONALE:: 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. OBJECTIVE:: This study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure. METHODS AND RESULTS:: 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. CONCLUSIONS:: 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.

AB - RATIONALE:: 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. OBJECTIVE:: This study aims to understand the mechanisms underlying altered phosphorylation of myofilament proteins in heart failure. METHODS AND RESULTS:: 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. CONCLUSIONS:: 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.

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