Epac1-dependent phospholamban phosphorylation mediates the cardiac response to stresses
Satoshi Okumura, … , Hikaru Tanaka, Yoshihiro Ishikawa
Satoshi Okumura, … , Hikaru Tanaka, Yoshihiro Ishikawa
Published April 24, 2014
Citation Information: J Clin Invest. 2014;124(6):2785-2801. https://doi.org/10.1172/JCI64784.
View: Text | PDF
Research Article Cardiology

Epac1-dependent phospholamban phosphorylation mediates the cardiac response to stresses

Abstract

PKA phosphorylates multiple molecules involved in calcium (Ca2+) handling in cardiac myocytes and is considered to be the predominant regulator of β-adrenergic receptor–mediated enhancement of cardiac contractility; however, recent identification of exchange protein activated by cAMP (EPAC), which is independently activated by cAMP, has challenged this paradigm. Mice lacking Epac1 (Epac1 KO) exhibited decreased cardiac contractility with reduced phospholamban (PLN) phosphorylation at serine-16, the major PKA-mediated phosphorylation site. In Epac1 KO mice, intracellular Ca2+ storage and the magnitude of Ca2+ movement were decreased; however, PKA expression remained unchanged, and activation of PKA with isoproterenol improved cardiac contractility. In contrast, direct activation of EPAC in cardiomyocytes led to increased PLN phosphorylation at serine-16, which was dependent on PLC and PKCε. Importantly, Epac1 deletion protected the heart from various stresses, while Epac2 deletion was not protective. Compared with WT mice, aortic banding induced a similar degree of cardiac hypertrophy in Epac1 KO; however, lack of Epac1 prevented subsequent cardiac dysfunction as a result of decreased cardiac myocyte apoptosis and fibrosis. Similarly, Epac1 KO animals showed resistance to isoproterenol- and aging-induced cardiomyopathy and attenuation of arrhythmogenic activity. These data support Epac1 as an important regulator of PKA-independent PLN phosphorylation and indicate that Epac1 regulates cardiac responsiveness to various stresses.

Authors

Satoshi Okumura, Takayuki Fujita, Wenqian Cai, Meihua Jin, Iyuki Namekata, Yasumasa Mototani, Huiling Jin, Yoshiki Ohnuki, Yayoi Tsuneoka, Reiko Kurotani, Kenji Suita, Yuko Kawakami, Shogo Hamaguchi, Takaya Abe, Hiroshi Kiyonari, Takashi Tsunematsu, Yunzhe Bai, Sayaka Suzuki, Yuko Hidaka, Masanari Umemura, Yasuhiro Ichikawa, Utako Yokoyama, Motohiko Sato, Fumio Ishikawa, Hiroko Izumi-Nakaseko, Satomi Adachi-Akahane, Hikaru Tanaka, Yoshihiro Ishikawa

×

Figure 2

Effects of EPAC activation on PLN phosphorylation in neonatal rat cardiac myocytes.

Options: View larger image (or click on image) Download as PowerPoint
Effects of EPAC activation on PLN phosphorylation in neonatal rat cardia...
(A and B) Treatment of neonatal rat cardiac myocytes with 8-CPT-AM (10 μM). PLN phosphorylation on serine-16 was significantly increased at 15 minutes and remained significantly (*P < 0.05 or **P < 0.01 versus 0 minutes, n = 4) greater at 120 minutes than at 0 minutes (A). PLN phosphorylation on threonine-17 was also significantly increased at 15 minutes after treatment. However, increase fell below significance at 60 minutes versus 0 minutes and remained unchanged at 120 minutes (P = NS, versus 0 minutes, n = 4) (B). Ratio of phosphorylated/total protein expression of PLN at baseline (0 min: Ctrl) was taken as 1-fold. (C) EPAC-mediated PLN phosphorylation on serine-16 was examined in neonatal rat cardiac myocytes transfected with PKCε siRNA or control siRNA (*P < 0.01, n = 5–7). Ratio of phosphorylated/total protein expression of PLN in cells transfected with control siRNA at baseline was taken as 100%. (D) PLN phosphorylation on serine-16 was examined in cells treated with Bnz-cAMP (50 μM) and/or 8-CPT-AM (10 μM). (##P < 0.001 versus Bnz-cAMP [50 μM] alone; ††P < 0.001 versus 8-CPT-AM [10 μM] alone). A similar tendency was observed when 10 μM Bnz-cAMP and 5 μM 8-CPT-AM were used together (#P < 0.001 versus Bnz-cAMP [10 μM] alone; †P < 0.001 versus 8-CPT-AM [5 μM] alone, n = 4–8; *P < 0.01 versus baseline, n = 4–8). Ratio of phosphorylated/total protein expression of PLN at baseline was taken as 100%.