Regulation of Akt/PKB activity by P21-activated kinase in cardiomyocytes (original) (raw)
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Novel insights into mechanisms for Pak1-mediated regulation of cardiac Ca2+ homeostasis
Frontiers in Physiology, 2015
A series of recent studies report novel roles for Pak1, a key member of the highly conserved family of serine-threonine protein kinases regulated by Ras-related small G-proteins, Cdc42/Rac1, in cardiac physiology and cardioprotection. Previous studies had identified Pak1 in the regulation of hypertrophic remodeling that could potentially lead to heart failure. This article provides a review of more recent findings on the roles of Pak1 in cardiac Ca 2+ homeostasis. These findings identified crucial roles for Pak1 in cardiomyocyte Ca 2+ handling and demonstrated that it functions through unique mechanisms involving regulation of the post-transcriptional activity of key Ca 2+-handling proteins, including the expression of Ca 2+-ATPase SERCA2a, along with the speculative possibility of an involvement in the maintenance of transverse (T)-tubular structure. They highlight important regulatory functions of Pak1 in Ca 2+ homeostasis in cardiac cells, and identify novel potential therapeutic strategies directed at manipulation of Pak1 signaling for the management of cardiac disease, particularly heart failure.
Circulation. Arrhythmia and electrophysiology, 2014
Impaired sarcoplasmic reticular Ca(2+) uptake resulting from decreased sarcoplasmic reticulum Ca(2+)-ATPase type 2a (SERCA2a) expression or activity is a characteristic of heart failure with its associated ventricular arrhythmias. Recent attempts at gene therapy of these conditions explored strategies enhancing SERCA2a expression and the activity as novel approaches to heart failure management. We here explore the role of Pak1 in maintaining ventricular Ca(2+) homeostasis and electrophysiological stability under both normal physiological and acute and chronic β-adrenergic stress conditions. Mice with a cardiomyocyte-specific Pak1 deletion (Pak1(cko)), but not controls (Pak1(f/f)), showed high incidences of ventricular arrhythmias and electrophysiological instability during either acute β-adrenergic or chronic β-adrenergic stress leading to hypertrophy, induced by isoproterenol. Isolated Pak1(cko) ventricular myocytes correspondingly showed aberrant cellular Ca(2+) homeostasis. Pak1(...
Akt2 Regulates Cardiac Metabolism and Cardiomyocyte Survival
Journal of Biological Chemistry, 2006
The Akt family of serine-threonine kinases participates in diverse cellular processes, including the promotion of cell survival, glucose metabolism, and cellular protein synthesis. All three known Akt family members, Akt1, Akt2 and Akt3, are expressed in the myocardium, although Akt1 and Akt2 are most abundant. Previous studies demonstrated that Akt1 and Akt3 overexpression results in enhanced myocardial size and function. Yet, little is known about the role of Akt2 in modulating cardiac metabolism, survival, and growth. Here, we utilize murine models with targeted disruption of the akt2 or the akt1 genes to demonstrate that Akt2, but not Akt1, is required for insulin-stimulated 2-[ 3 H]deoxyglucose uptake and metabolism. In contrast, akt2 ؊/؊ mice displayed normal cardiac growth responses to provocative stimulation, including ligand stimulation of cultured cardiomyocytes, pressure overload by transverse aortic constriction, and myocardial infarction. However, akt2 ؊/؊ mice were found to be sensitized to cardiomyocyte apoptosis in response to ischemic injury, and apoptosis was significantly increased in the peri-infarct zone of akt2 ؊/؊ hearts 7 days after occlusion of the left coronary artery. These results implicate Akt2 in the regulation of cardiomyocyte metabolism and survival. Cardiac growth and metabolism are coordinated through the integration of a complex array of extracellular and intracellular signals. Much recent work suggests that the Akt family of intracellular serine-threonine kinases regulates both cardiac growth and metabolism (1-3). The Akt family of serine-threonine kinases consists of three isoforms, Akt1, Akt2, and Akt3, each encoded by distinct, highly conserved genes. All three isoforms are expressed in the myocardium, although Akt1 and Akt2 comprise the vast majority of total Akt protein in the heart (2). Examination of numerous experimental models implicates both Akt1 and Akt3 in regulating pathological and physiological hypertrophy (4-6). Indeed, the hypothesis that the phosphatidylinositol 3-kinase (PI3K) 3 ␣-Akt1 cascade mediates physiological cardiac growth is now well founded (7). Cardiacspecific expression of constitutively active Akt1 (myristoylated Akt1) in transgenic mice results in massive cardiac hypertrophy and fibrosis consistent with pathological hypertrophy (4), and nuclear localization of Akt1 was recently shown to augment ventricular function and contractility (8). A comparable phenotype was observed in response to cardiac overexpression of activated Akt3, whereas no observable cardiac growth defects were detectable in Akt3-deficient mice at baseline (6). Akt family members are also key regulators of cellular metabolism. Indeed, GLUT4 translocation to the plasma membrane is a wortmannin-sensitive process (9), and Akt2-mediated phosphorylation of the syntaxin interacting protein (synip) results in docking and fusion of GLUT4-containing vesicles with the plasma membrane (10). Akt family members promote glycogen synthesis through phosphorylation and inhibition of glycogen synthase kinase 3 (GSK3), which itself inhibits glycogen synthesis (11). GSK3 phosphorylation results in the augmentation of glycogen synthesis, whereas Akt activation antagonizes the AMP-activated protein kinase (12), a key mediator of glycogenolysis and lipolysis. In addition, Akt kinases inhibit fatty acid metabolism by phosphorylating and inhibiting FOXO-1, a forkhead family transcription factor that positively modulates fatty acid oxidative gene expression (13). Although the role of Akt family members in cardiac growth and metabolism has been widely studied, the role of Akt2 in the development of physiological and pathological cardiac hypertrophy is unknown. Additionally, the role of individual Akt family members in the regulation of cardiac metabolism remains unexplored. In the current study, an Akt2 loss-of-function murine model was utilized to assess the role of Akt2 in cardiac growth, metabolism, and cardiomyocyte survival. Here, we show that Akt2 is dispensable in the development of cardiac hypertrophy in response to a variety of physiological and pathological provocative stimuli. Conversely, we demonstrate that
Pim-1 regulates cardiomyocyte survival downstream of Akt
Nature Medicine, 2007
The serine-threonine kinases Pim-1 and Akt regulate cellular proliferation and survival. Although Akt is known to be a crucial signaling protein in the myocardium, the role of Pim-1 has been overlooked. Pim-1 expression in the myocardium of mice decreased during postnatal development, re-emerged after acute pathological injury in mice and was increased in failing hearts of both mice and humans. Cardioprotective stimuli associated with Akt activation induced Pim-1 expression, but compensatory increases in Akt abundance and phosphorylation after pathological injury by infarction or pressure overload did not protect the myocardium in Pim-1-deficient mice. Transgenic expression of Pim-1 in the myocardium protected mice from infarction injury, and Pim-1 expression inhibited cardiomyocyte apoptosis with concomitant increases in Bcl-2 and Bcl-X L protein levels, as well as in Bad phosphorylation levels. Relative to nontransgenic controls, calcium dynamics were significantly enhanced in Pim-1-overexpressing transgenic hearts, associated with increased expression of SERCA2a, and were depressed in Pim-1-deficient hearts. Collectively, these data suggest that Pim-1 is a crucial facet of cardioprotection downstream of Akt.
A kinase interacting protein (AKIP1) is a key regulator of cardiac stress
Proceedings of the National Academy of Sciences, 2013
cAMP-dependent protein kinase (PKA) regulates a myriad of functions in the heart, including cardiac contractility, myocardial metabolism, and gene expression. However, a molecular integrator of the PKA response in the heart is unknown. Here, we show that the PKA adaptor A-kinase interacting protein 1 (AKIP1) is up-regulated in cardiac myocytes in response to oxidant stress. Mice with cardiac gene transfer of AKIP1 have enhanced protection to ischemic stress. We hypothesized that this adaptation to stress was mitochondrialdependent. AKIP1 interacted with the mitochondrial localized apoptosis inducing factor (AIF) under both normal and oxidant stress. When cardiac myocytes or whole hearts are exposed to oxidant and ischemic stress, levels of both AKIP1 and AIF were enhanced. AKIP1 is preferentially localized to interfibrillary mitochondria and up-regulated in this cardiac mitochondrial subpopulation on ischemic injury. Mitochondria isolated from AKIP1 genetransferred hearts showed increased mitochondrial localization of AKIP1, decreased reactive oxygen species generation, enhanced calcium tolerance, decreased mitochondrial cytochrome C release, and enhance phosphorylation of mitochondrial PKA substrates on ischemic stress. These observations highlight AKIP1 as a critical molecular regulator and a therapeutic control point for stress adaptation in the heart. ischemia/reperfusion | oxidative stress
PHLPP-1 Negatively Regulates Akt Activity and Survival in the Heart
Circulation Research, 2010
Rationale: The recently discovered PHLPP-1 (PH domain leucine-rich repeat protein phosphatase-1) selectively dephosphorylates Akt at Ser473 and terminates Akt signaling in cancer cells. The regulatory role of PHLPP-1 in the heart has not been considered. Objective: To test the hypothesis that blockade/inhibition of PHLPP-1 could constitute a novel way to enhance Akt signals and provide cardioprotection. Methods and Results: PHLPP-1 is expressed in neonatal rat ventricular myocytes (NRVMs) and in adult mouse ventricular myocytes (AMVMs). PHLPP-1 knockdown by small interfering RNA significantly enhances phosphorylation of Akt (p-Akt) at Ser473, but not at Thr308, in NRVMs stimulated with leukemia inhibitory factor (LIF). The increased phosphorylation is accompanied by greater Akt catalytic activity. PHLPP-1 knockdown enhances LIF-mediated cardioprotection against doxorubicin and also protects cardiomyocytes against H 2 O 2. Direct Akt effects at mitochondria have been implicated in cardioprotection and mitochondria/ cytosol fractionation revealed a significant enrichment of PHLPP-1 at mitochondria. The ability of PHLPP-1 knockdown to potentiate LIF-mediated increases in p-Akt at mitochondria and an accompanying increase in mitochondrial hexokinase-II was demonstrated. We generated PHLPP-1 knockout (KO) mice and demonstrate that AMVMs isolated from KO mice show potentiated p-Akt at Ser473 in response to agonists. When isolated perfused hearts are subjected to ischemia/reperfusion, p-Akt in whole-heart homogenates and in the mitochondrial fraction is significantly increased. Additionally in PHLPP-1 KO hearts, the increase in p-Akt elicited by ischemia/reperfusion is potentiated and, concomitantly, infarct size is significantly reduced. Conclusions: These results implicate PHLPP-1 as an endogenous negative regulator of Akt activity and cell survival in the heart. (Circ Res. 2010;107:476-484.
Disruption of Protein Kinase A Interaction with A-kinase-anchoring Proteins in the Heart in Vivo
Journal of Biological Chemistry, 2008
Protein kinase A (PKA)-dependent phosphorylation is regulated by targeting of PKA to its substrate as a result of binding of regulatory subunit, R, to A-kinase-anchoring proteins (AKAPs). We investigated the effects of disrupting PKA targeting to AKAPs in the heart by expressing the 24-amino acid regulatory subunit RII-binding peptide, Ht31, its inactive analog, Ht31P, or enhanced green fluorescent protein by adenoviral gene transfer into rat hearts in vivo. Ht31 expression resulted in loss of the striated staining pattern of type II PKA (RII), indicating loss of PKA from binding sites on endogenous AKAPs. In the absence of isoproterenol stimulation, Ht31-expressing hearts had decreased ؉dP/dt max and ؊dP/dt min but no change in left ventricular ejection fraction or stroke volume and decreased end diastolic pressure versus controls. This suggests that cardiac output is unchanged despite decreased ؉dP/dt and ؊dP/dt. There was also no difference in PKA phosphorylation of cardiac troponin I (cTnI), phospholamban, or ryanodine receptor (RyR 2). Upon isoproterenol infusion, ؉dP/dt max and-dP/dt min did not differ between Ht31 hearts and controls. At higher doses of isoproterenol, left ventricular ejection fraction and stroke volume increased versus isoproterenol-stimulated controls. This occurred in the context of decreased PKA phosphorylation of cTnI, RyR 2 , and phospholamban versus controls. We previously showed that expression of N-terminal-cleaved cTnI (cTnI-ND) in transgenic mice improves cardiac function. Increased cTnI N-terminal truncation was also observed in Ht31-expressing hearts versus controls. Increased cTnI-ND may help compensate for reduced PKA phosphorylation as occurs in heart failure.
Muscle ring finger protein-1 inhibits PKC activation and prevents cardiomyocyte hypertrophy
The Journal of Cell Biology, 2004
uch effort has focused on characterizing the signal transduction cascades that are associated with cardiac hypertrophy. In spite of this, we still know little about the mechanisms that inhibit hypertrophic growth. We define a novel anti-hypertrophic signaling pathway regulated by muscle ring finger protein-1 (MURF1) that inhibits the agonist-stimulated PKC-mediated signaling response in neonatal rat ventricular myocytes. MURF1 interacts with receptor for activated protein kinase C (RACK1) and colocalizes with RACK1 after activation with phenylephrine or PMA. Coincident with this agonist-M stimulated interaction, MURF1 blocks PKC ε translocation to focal adhesions, which is a critical event in the hypertrophic signaling cascade. MURF1 inhibits focal adhesion formation, and the activity of downstream effector ERK1/2 is also inhibited in the presence of MURF1. MURF1 inhibits phenylephrine-induced (but not IGF-1-induced) increases in cell size. These findings establish that MURF1 is a key regulator of the PKC-dependent hypertrophic response and can blunt cardiomyocyte hypertrophy, which may have important implications in the pathophysiology of clinical cardiac hypertrophy.