Akt2 Regulates Cardiac Metabolism and Cardiomyocyte Survival
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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.Keywords:
AKT2
AKT3
The kinase Akt is a key downstream mediator of the phosphoinositide-3-kinase signaling pathway and participates in a variety of cellular processes. Akt comprises three isoforms each encoded by a separate gene. There is evidence to indicate that Akt is involved in the survival and protection of auditory hair cells in vitro. However, little is known about the physiological role of Akt in the inner ear—especially in the intact animal. To elucidate this issue, we first analyzed the mRNA expression of the three Akt isoforms in the inner ear of C57/BL6 mice by real-time PCR. Next, we tested the susceptibility to gentamicin-induced auditory hair cell loss in isoform-specific Akt knockout mice compared to wild-types (C57/BL6) in vitro. To analyze the effect of gene deletion in vivo, hearing and cochlear microanatomy were evaluated in Akt isoform knockout animals. In this study, we found that all three Akt isoforms are expressed in the cochlea. Our results further indicate that Akt2 and Akt3 enhance hair cell resistance to ototoxicity, while Akt1 does not. Finally, we determined that untreated Akt1 and Akt2/Akt3 double knockout mice display significant hearing loss, indicating a role for these isoforms in normal hearing. Taken together, our results indicate that each of the Akt isoforms plays a distinct role in the mammalian inner ear.
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Akt (PKB) is a serine/threonine protein kinase that plays an important role in the transduction of signals affecting apoptosis, cell proliferation and survival. The Akt gene is frequently hyperactivated in tumors and has been shown to be amplified in a number of types of human cancers. Furthermore, Akt activity is elevated in cell lines with the mutated PTEN tumor suppressor gene. These studies establish Akt as an attractive target for cancer therapy. To determine the roles of Akt1, Akt2 and Akt3 in signal transduction, constitutively active Akt1, Akt2 and Akt3 was ectopically overexpressed in human pancreatic MiaPaCa-2 cells. The three Akt stable clones were characterized to determine their effects on transformation and proliferation. Compared to a vector control, the three Akt clones were able to drive cellular proliferation even in reduced serum conditions. Furthermore, in soft-agar assays, the Akt clones showed an 25-38% increase in colony formation in 2% serum. Our results indicate that all three forms of Akt may have protective effects within the cell depending on the type of apoptotic stimuli. Using 2D-PAGE comparisons between parental and Akt overexpressing cells, we attempted to determine novel targets of Akt phosphorylation. In this study, we identified prohibitin as a substrate for Akt both in vitro and in vivo. These studies suggest that Akt may regulate the cellular function of prohibitin via its phosphorylation.
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Proto-Oncogene Proteins c-akt
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This article describes recent advances in the development and biological evaluation of allosteric and ATP-competitive small molecule inhibitors for the serine/threonine kinase Akt (protein kinase B, PKB). Unregulated activation of the PI3K/Akt/PTEN pathway is a prominent feature of many human cancers and Akt is over-expressed or activated in all major cancers making Akt an exciting new target for cancer therapy. The development of Akt inhibitors has been complicated and hampered by the presence of three Akt isozymes, (Akt1, Akt2 and Akt3) which differ in function and tissue distribution, as well as a lack of Akt specific inhibitors. In the past 18 months, a large number of reports have appeared describing the discovery and development of allosteric Akt kinase inhibitors and classical ATP-competitive Akt kinase inhibitors. This review will discuss the PI3K/Akt/PTEN pathway, allosteric and ATP-competitive Akt kinase inhibitors, their biological evaluation and progress towards target validation.
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Akt/PKB is a serine/threonine protein kinase that functions as a critical regulator of cell survival and proliferation. Akt/PKB family comprises three highly homologous members known as PKBα/Akt1, PKBβ/Akt2 and PKBγ/Akt3 in mammalian cells. Similar to many other protein kinases, Akt/PKB contains a conserved domain structure including a specific PH domain, a central kinase domain and a carboxyl-terminal regulatory domain that mediates the interaction between signaling molecules. Akt/PKB plays important roles in the signaling pathways in response to growth factors and other extracellular stimuli to regulate several cellular functions including nutrient metabolism, cell growth, apoptosis and survival. This review surveys recent developments in understanding the molecular mechanisms of Akt/PKB activation and its roles in cell survival in normal and cancer cells.
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Protein kinase domain
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The serine/threonine protein kinase Akt is a major transducer of the phosphoinositide 3-kinase pathway and plays a crucial role in regulation of cellular processes such as growth, metabolism, survival and proliferation. Mammalian cells are characterized by the expression of three different Akt isoforms (Akt1, Akt2, Akt3), encoded by distinct genes. Increased expression and activation of Akt observed in many human cancers is usually caused by amplification or mutation of Akt genes, amplification and activating mutation of the catalytic subunit of PI3K or deletion and mutations of phosphatidylinositol-3,4,5-triphosphate phosphatase--PTEN. Although activation of Akt alone is believed to be insufficient for tumorigenesis, it contributes to cancer progression by inhibiting apoptosis, promoting changes in metabolism and proliferation of cells and regulating their migration and invasion capabilities. Recent studies have provided evidence that depending on the cell type each specific Akt isoform may play a positive or negative role in cell migration and invasion. Akt is also involved in regulation of tumor angiogenesis.
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AbstractAKT is a key serine/threonine kinase in the PTEN/PI3K/AKT pathway1 and activation of AKT is often observed in human cancers. To explore the role of AKT in cell survival in different tumor cells, we tested 20 human tumor cell lines for response to knockdown of AKT by small interference RNA (siRNA) and/or a kinase-dead mutant AKT. siRNA-mediated knockdown of all three AKT isoforms in tumor cell lines led to a reduction of phosphorylation of AKT substrates. Knockdown of AKT resulted in apoptosis in 6 out of 11 tumor cells with activated AKT. In contrast, knockdown of AKT induced apoptosis in 3 out of 9 cell lines with a low level of active AKT. The responsiveness of the cells to knockdown of AKT was not affected by mutational status of p53 but appeared correlated with overexpression of HER2. To assess the role of individual AKT isoforms, five of the cell lines responsive to knockdown of AKT were further characterized. In ZR-75 cells, AKT1 is the predominant isoform responsible for cell proliferation and survival. Conversely, in IGROV1 cells, AKT2 plays a major role in cell proliferation, but no single isoform is essential for cell survival. Thus, the relative importance of the AKT isoforms is cell linespecific. Our data suggest that inhibiting all three AKT isoforms is necessary to elicit maximal apoptotic response in tumor cells, and the level of activated AKT is a favorable but not always reliable biomarker for pre-selection of responsive tumor cell lines to AKT inhibitors.
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Objective To study the molecular mechanism of phosphatidylinositol 3-kinase (PI3K)/Akt active pathway that involve in the regulation of apoptosis. Methods Different tumor cells were seeded into either 96 well or 6 well tissue culture plates and incubated with isoform specific Akt inhibitor for 6 hours. Akt phosphorylation and its activity were evaluated with immunoprecipitation; the activation of phosphorylated Akt were detected by immunoprecipitation and western blotting; Caspase assay has been performed to quantitate apoptosis. Results ①The inhibitors could reduce the phosphorylations of threonine 308 and serine 473 of isoform specific Akt ;②the inhibition of any isoform specific Akt could significantly downregulate the activation and activity of its substrate(GSK3) , but it was not the same case if blocking two isoforms of both Akt1 and Akt2 at the same time;③ PI3K involves in the regulation of apoptosis mainly through its downstream gene(AKT) , specifically the isoforms of Aktl and Akt2. Conclusion PI3K/Akt involves in the regulation of apoptosis, two isoforms of Akt(Akt1 and Akt2) are mainly the regulator of PI3K in terms of the regulation of apoptosis. It is required to inhibit two isoforms of both Akt1 and Akt2 to initiate apoptosis.
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Proto-Oncogene Proteins c-akt
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Recent studies indicate that dysregulation of the Akt/PKB family of serine/threonine kinases is a prominent feature of many human cancers. The Akt/PKB family is composed of three members termed Akt1/PKBalpha, Akt2/PKBbeta, and Akt3/PKBgamma. It is currently not known to what extent there is functional overlap between these family members. We have recently identified small molecule inhibitors of Akt. These compounds have pleckstrin homology domain-dependent, isozyme-specific activity. In this report, we present data showing the relative contribution that inhibition of the different isozymes has on the apoptotic response of tumor cells to a variety of chemotherapies. In multiple cell backgrounds, maximal induction of caspase-3 activity is achieved when both Akt1 and Akt2 are inhibited. This induction is not reversed by overexpression of functionally active Akt3. The level of caspase-3 activation achieved under these conditions is equivalent to that observed with the phosphatidylinositol-3-kinase inhibitor LY294002. We also show that in different tumor cell backgrounds inhibition of mammalian target of rapamycin, a downstream substrate of Akt, is less effective in inducing caspase-3 activity than inhibition of Akt1 and Akt2. This shows that the survival phenotype conferred by Akt can be mediated by signaling pathways independent of mammalian target of rapamycin in some tumor cell backgrounds. Finally, we show that inhibition of both Akt1 and Akt2 selectively sensitizes tumor cells, but not normal cells, to apoptotic stimuli.
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Proto-Oncogene Proteins c-akt
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Tumor progression
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The Akt (PKB) protein kinases are critical regulators of human physiology that control an impressive array of diverse cellular functions, including the modulation of growth, survival, proliferation and metabolism. The Akt kinase family is comprised of three highly homologous isoforms: Akt1 (PKBα), Akt2 (PKBβ) and Akt3 (PKBγ). Phenotypic analyses of Akt isoform knockout mice documented Akt isoform specific functions in the regulation of cellular growth, glucose homeostasis and neuronal development. Those studies establish that the functions of the different Akt kinases are not completely overlapping and that isoform-specific signaling contributes to the diversity of Akt activities. However, despite these important advances, a thorough understanding about the specific roles of Akt family members and the molecular mechanisms that determine Akt isoform functional specificity will be essential to elucidate the complexity of Akt regulated cellular processes and how Akt isoform-specific deregulation might contribute to disease states. Here, we summarize recent advances in understanding the roles of Akt isoforms in the regulation of metabolism and cancer, and possible mechanisms contributing to Akt isoform functional specificity.
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Proto-Oncogene Proteins c-akt
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