PHD2/3-dependent hydroxylation tunes cardiac response to β-adrenergic stress via phospholamban
Liang Xie, … , Gerhard Meissner, Cam Patterson
Liang Xie, … , Gerhard Meissner, Cam Patterson
Published July 1, 2015; First published June 15, 2015
Citation Information: J Clin Invest. 2015;125(7):2759-2771. https://doi.org/10.1172/JCI80369.
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Research Article Cardiology

PHD2/3-dependent hydroxylation tunes cardiac response to β-adrenergic stress via phospholamban

Abstract

Ischemic heart disease is the leading cause of heart failure. Both clinical trials and experimental animal studies demonstrate that chronic hypoxia can induce contractile dysfunction even before substantial ventricular damage, implicating a direct role of oxygen in the regulation of cardiac contractile function. Prolyl hydroxylase domain (PHD) proteins are well recognized as oxygen sensors and mediate a wide variety of cellular events by hydroxylating a growing list of protein substrates. Both PHD2 and PHD3 are highly expressed in the heart, yet their functional roles in modulating contractile function remain incompletely understood. Here, we report that combined deletion of Phd2 and Phd3 dramatically decreased expression of phospholamban (PLN), resulted in sustained activation of calcium/calmodulin-activated kinase II (CaMKII), and sensitized mice to chronic β-adrenergic stress–induced myocardial injury. We have provided evidence that thyroid hormone receptor-α (TR-α), a transcriptional regulator of PLN, interacts with PHD2 and PHD3 and is hydroxylated at 2 proline residues. Inhibition of PHDs increased the interaction between TR-α and nuclear receptor corepressor 2 (NCOR2) and suppressed Pln transcription. Together, these observations provide mechanistic insight into how oxygen directly modulates cardiac contractility and suggest that cardiac function could be modulated therapeutically by tuning PHD enzymatic activity.

Authors

Liang Xie, Xinchun Pi, W.H. Davin Townley-Tilson, Na Li, Xander H.T. Wehrens, Mark L. Entman, George E. Taffet, Ashutosh Mishra, Junmin Peng, Jonathan C. Schisler, Gerhard Meissner, Cam Patterson

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Figure 1

Depletion of PHD2/3 results in decreased PLN protein levels and increased SR Ca2+ uptake.

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Depletion of PHD2/3 results in decreased PLN protein levels and increase...
Phd2/3fl/flCre+/– mice were i.p. injected with tamoxifen once daily for 5 consecutive days. (A) Hearts were harvested on the indicated day after the first injection of tamoxifen, and real-time PCR was performed to determine the relative mRNA levels of Phd2 and Phd3. Phd2 and Phd3 mRNA levels were significantly decreased after 2 days of tamoxifen injection. n = 4, *P < 0.01, compared with day 0, 1-way ANOVA. (B and C) Seven days after the first injection of tamoxifen, mouse hearts were harvested, and Western blot analysis was performed. Densitometric analysis of PLN protein levels is shown in C. PLN, but not SERCA2a or RyR2, protein levels were significantly decreased. n = 6, *P < 0.01, 2-tailed Student’s t test. (D) Neonatal ventricular myocytes were isolated from Phd2/3fl/fl Cre+/– and Phd2/3fl/fl Cre–/– mice. After treatment with 4-OHT for 5 days, myocytes were treated with ISO for 0, 1, 2, or 4 hours. Western blot analysis was then performed. (E) After infection with adenovirus expressing normoxia-stable HIF-1α for 0, 1, 2, 4, and 7 days, neonatal rat ventricular myocytes were harvested, and Western blot analysis was performed. p-PLN, phosphorylated PLN. (F) SR Ca2+ uptake rates were measured using heart homogenates from Phd2/3fl/fl Cre+/– and Phd2/3fl/fl Cre–/– mice after treatment with tamoxifen for 7 days. The Ca2+ uptake rate was increased in mice lacking PHD2 and PHD3. n = 4, *P < 0.05, 2-tailed Student’s t test.