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Robert A. Neff - One of the best experts on this subject based on the ideXlab platform.
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Susceptibility of ATP-sensitive K+ channels to cell stress through mediation of phosphoinositides as examined by photoirradiation.
The Journal of physiology, 2000Co-Authors: Robert A. NeffAbstract:Cell stress is implicated in a number of pathological states of metabolism, such as ischaemia, reperfusion and apoptosis in heart, neurons and other tissues. While it is known that the ATP-sensitive K+ (KATP) channel plays a role during metabolic abnormality, little information is available about the direct response of this channel to cell stress. Using photoirradiation stimulation, we studied the effects of cell stress on both native and cloned KATP channels. Single KATP channel currents were recorded from cell-attached and inside-out patches of rat ventricular myocytes and COS-1 cells coexpressing SUR2 and Kir6.2. KATP channel activity increased within < 1 min upon irradiation. The activity resulted from increased maximal open probability and decreased ATP inhibition. The effects remained after the irradiation was stopped. Irradiation also affected the channels formed only by Kir6.2DeltaC35. The irradiation-induced activation was comparable to that induced by phosphoinositides. Analysis of Phosphatidylinositol composition revealed an elevated Phosphatidylinositol bisphosphate level with irradiation. Wortmannin, an inhibitor of Phosphatidylinositol Kinases, decreased both the irradiation-induced channel activity and the production of Phosphatidylinositol bisphosphates. Radical scavengers also reduced the irradiation-induced activation, suggesting a role for free radicals, an immediate product of photoirradiation. We conclude that photoirradiation can modify the single-channel properties of KATP, which appears to be mediated by phosphoinositides. Our study suggests that cellular stress may be linked with KATP channels, and we offer a putative mechanism for such a linkage.
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Susceptibility of ATP‐sensitive K+ channels to cell stress through mediation of phosphoinositides as examined by photoirradiation
The Journal of Physiology, 2000Co-Authors: Robert A. NeffAbstract:1 Cell stress is implicated in a number of pathological states of metabolism, such as ischaemia, reperfusion and apoptosis in heart, neurons and other tissues. While it is known that the ATP-sensitive K+ (KATP) channel plays a role during metabolic abnormality, little information is available about the direct response of this channel to cell stress. Using photoirradiation stimulation, we studied the effects of cell stress on both native and cloned KATP channels. 2 Single KATP channel currents were recorded from cell-attached and inside-out patches of rat ventricular myocytes and COS-1 cells coexpressing SUR2 and Kir6.2. KATP channel activity increased within < 1 min upon irradiation. The activity resulted from increased maximal open probability and decreased ATP inhibition. The effects remained after the irradiation was stopped. Irradiation also affected the channels formed only by Kir6.2ΔC35. 3 The irradiation-induced activation was comparable to that induced by phosphoinositides. Analysis of Phosphatidylinositol composition revealed an elevated Phosphatidylinositol bisphosphate level with irradiation. Wortmannin, an inhibitor of Phosphatidylinositol Kinases, decreased both the irradiation-induced channel activity and the production of Phosphatidylinositol bisphosphates. 4 Radical scavengers also reduced the irradiation-induced activation, suggesting a role for free radicals, an immediate product of photoirradiation. 5 We conclude that photoirradiation can modify the single-channel properties of KATP, which appears to be mediated by phosphoinositides. Our study suggests that cellular stress may be linked with KATP channels, and we offer a putative mechanism for such a linkage.
Brooke M. Emerling - One of the best experts on this subject based on the ideXlab platform.
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Abstract A078: Towards understanding noncanonical Phosphatidylinositol Kinases in the maintenance of prostate metabolism
Poster Presentations - Proffered Abstracts, 2018Co-Authors: Joanna Triscott, Matteo Benelli, Verena Sailer, Davide Prandi, Brooke M. Emerling, Francesca Demichelis, Lewis C. Cantley, Mark A. RubinAbstract:An estimated 1 in 7 men will develop prostate cancer (PCa) with many progressing to advanced castrate-resistant disease. Unlike other tissue types, normal prostate cell growth and development is heavily dependent on the androgen receptor (AR) signaling pathway. While the introduction of novel AR antagonists for clinical treatment has improved outcomes, most castration-resistant prostate cancer (CRPC) patients ultimately develop resistance to these therapies. A need exists to better understand the mechanisms that control the transition of prostate cells from a hormone-dependent to castrate-resistant state. Androgens strongly influence the metabolic state of PCa cells to favor sustained cellular growth. We hypothesize there are effectors working in conjunction with AR to coordinate alterations to androgen-dependent metabolism that are linchpins in the orchestration of the transition to CRPC. Leading candidates are members of phosphoinositol (PI) pathways, which have a high frequency of alteration in PCa (i.e phosphoinositide 3-kinase (PI3K)). Herein we explore a family of poorly understood lipid Kinases called the type II Phosphatidylinositol-5-phosphate 4-Kinases (PI5P4Ks) and predict them to be critical regulators of cancer cell survival. PI5P4Ks are druggable targets that act by phosphorylating the lipid Phosphatidylinositol-5-phosphate (PI 5-P) at the 4 position of the inositol ring to generate Phosphatidylinositol-4,5-bisphosphate (PI-4,5-P2; PIP2). We implicate the three PI5P4K isoforms (PI5P4Kα, PI5P4Kβ, and PI5P4Kγ) encoded by the genes PIP4K2A, B, and C, to be important regulators of cancer metabolism that play a role in the maintenance of prostate biology and oncogenesis. Analysis of transcript data revealed expression of PIP4K2A, B, and C in primary PCa patient samples, which was correlated with an AR activation gene signature and hotspot tumor suppressor deletion. As well, isoform expression was assessed for differential expression in relation to an integrated neuroendocrine prostate cancer mRNA score (TCGA; n=333). PI5P4Kα and PI5P4Kβ protein was detected in primary and advanced prostate cancer using optimized antibodies of patient tissue TMAs (n= 72). Using in vitro LNCaP cell models, siRNA knockdown systems were tested to evaluate the molecular consequence of targeting PIP4K2A and PIP4K2B in androgen-dependent systems. Stable knockdown using fluorescently labeled lentiviral shRNA constructs significantly reduced proliferation of shPIP4K2 treated cells. As well, we have produced a prostate-specific PI5P4K knockout mouse model by expressing probasin-driven Cre in a homozygous 129/SvEv Pip4k2aflx/flx murine strain. Finally, implementation of a discovery-based metabolomic platform (Metabolon HD4) was used to profile the overall shift in metabolite species that results from downregulating the expression of PIP4K2A in androgen-dependent cell models. In summary, we have developed novel insights into the role of a family of noncanonical PI Kinases in prostate biology. There are a growing number of PI3K/AKT inhibitors being tested in combination with androgen deprivation therapy in clinical trials, but there is still almost nothing known about the potential crosstalk of the greater PI kinase network. These data convincingly implicate a fundamental role for PI5P4Ks in PCa androgen signaling and metabolism, as well as lay the foundation of phenotypic understanding of what PI5P4K is responsible for in the prostate. Citation Format: Joanna Triscott, Matteo Benelli, Verena Sailer, Davide Prandi, Brooke Emerling, Francesca Demichelis, Lewis Cantley, Mark A. Rubin. Towards understanding noncanonical Phosphatidylinositol Kinases in the maintenance of prostate metabolism [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A078.
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abstract 4714 targeting p53 mutant cancers through inhibition of the Phosphatidylinositol 5 phosphate 4 Kinases
Cancer Research, 2015Co-Authors: Brooke M. Emerling, Zhiwei Yang, Ryan M Loughran, Jonathan T Yang, Jared L Johnson, Rajan Pragani, Mindy I Davis, Min Shen, Matthew B Boxer, Anton SimeonovAbstract:The bulk of cellular Phosphatidylinositol-4,5-bisphosphate (PI-4,5-P2) is generated by the canonical pathway in which a 5-kinase converts Phosphatidylinositol-4-phosphate (PI-4-P) to PI-4,5-P2. However, several years ago we discovered that PI-4, 5-P2 can also be generated at intracellular sites by a family of Kinases that phosphorylate the 4 position of Phosphatidylinositol-5-phosphate (PI-5-P), a lipid that was not previously known to exist in nature. These enzymes are called the type 2 Phosphatidylinositol-5-phosphate 4-Kinases (PI5P4Ks). To date, we have shown that a subset of breast cancers express high levels of PI5P4Kα and/or β, and have provided evidence that these Kinases are essential for growth in the absence of p53 (Emerling et al., 2013). Furthermore, we showed that PI5P4Kα and β play critical roles in mediating changes in metabolism in response to cellular stress, in particular, stress that occurs in the absence of p53. Here, we disclose the discovery of potent and selective inhibitors of PI5P4K (Kis in the range of 10 to 200 nM). These novel PI5P4K inhibitors have provided us with powerful tools to further investigate the role of the PI5P4K enzymes in cancer metabolism. Through pharmacological inhibition of PI5P4K, we now have support that PI5P4K provides an alternative pathway to p53 in regard to mediating responses to metabolic and oxidative stress, thereby suggesting that the PI5P4K enzymes are essential for survival mechanisms when p53 function is lost. Above all, these inhibitors may be effective therapies not only for breast cancers with genetic aberrations in TP53, but in all cancer types with loss of p53 function. Supported by DOD Breast Research Program Breakthrough Award to B.M.E. References Emerling BM, Hurov JB, Poulogiannis G, Tsukazawa KS, Wulf G, Bell EL, Shim H, Choo-Wing R, Bellinger G, Lamia KA, Rameh LE, Sasaki A, Asara JM, Yuan X, Bullock A, Brown V, Signoretti S, and Cantley LC. (2013) Depletion of a Putatively Druggable Class of Phosphatidylinositol Kinases Inhibits Growth of p53 Null Tumors. Cell. Nov 7; 155 (4): 844-57. Citation Format: Brooke M. Emerling, Zhiwei Yang, Ryan Loughran, T.Jonathan Yang, Jared Johnson, Rajan Pragani, Mindy Davis, Min Shen, Matthew Boxer, Anton Simeonov, Lewis C. Cantley. Targeting p53 mutant cancers through inhibition of the Phosphatidylinositol-5-phosphate 4-Kinases. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4714. doi:10.1158/1538-7445.AM2015-4714
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depletion of a putatively druggable class of Phosphatidylinositol Kinases inhibits growth of p53 null tumors
Cell, 2013Co-Authors: Brooke M. Emerling, Jonathan Hurov, George Poulogiannis, Kazumi S Tsukazawa, Rayman ChoowingAbstract:Summary Here, we show that a subset of breast cancers express high levels of the type 2 Phosphatidylinositol-5-phosphate 4-Kinases α and/or β (PI5P4Kα and β) and provide evidence that these Kinases are essential for growth in the absence of p53. Knocking down PI5P4Kα and β in a breast cancer cell line bearing an amplification of the gene encoding PI5P4K β and deficient for p53 impaired growth on plastic and in xenografts. This growth phenotype was accompanied by enhanced levels of reactive oxygen species (ROS) leading to senescence. Mice with homozygous deletion of both TP53 and PIP4K2B were not viable, indicating a synthetic lethality for loss of these two genes. Importantly however, PIP4K2A −/− , PIP4K2B +/− , and TP53 −/− mice were viable and had a dramatic reduction in tumor formation compared to TP53 −/− littermates. These results indicate that inhibitors of PI5P4Ks could be effective in preventing or treating cancers with mutations in TP53.
Rayman Choowing - One of the best experts on this subject based on the ideXlab platform.
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depletion of a putatively druggable class of Phosphatidylinositol Kinases inhibits growth of p53 null tumors
Cell, 2013Co-Authors: Brooke M. Emerling, Jonathan Hurov, George Poulogiannis, Kazumi S Tsukazawa, Rayman ChoowingAbstract:Summary Here, we show that a subset of breast cancers express high levels of the type 2 Phosphatidylinositol-5-phosphate 4-Kinases α and/or β (PI5P4Kα and β) and provide evidence that these Kinases are essential for growth in the absence of p53. Knocking down PI5P4Kα and β in a breast cancer cell line bearing an amplification of the gene encoding PI5P4K β and deficient for p53 impaired growth on plastic and in xenografts. This growth phenotype was accompanied by enhanced levels of reactive oxygen species (ROS) leading to senescence. Mice with homozygous deletion of both TP53 and PIP4K2B were not viable, indicating a synthetic lethality for loss of these two genes. Importantly however, PIP4K2A −/− , PIP4K2B +/− , and TP53 −/− mice were viable and had a dramatic reduction in tumor formation compared to TP53 −/− littermates. These results indicate that inhibitors of PI5P4Ks could be effective in preventing or treating cancers with mutations in TP53.
Joseph P. Albanesi - One of the best experts on this subject based on the ideXlab platform.
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A Novel Family of Phosphatidylinositol 4-Kinases Conserved from Yeast to Humans
The Journal of biological chemistry, 2001Co-Authors: Barbara Barylko, Stefan H. Gerber, Derk D. Binns, Nikolai Grichine, Mikhail Khvotchev, Thomas C. Südhof, Joseph P. AlbanesiAbstract:Abstract Phosphatidylinositolpolyphosphates (PIPs) are centrally involved in many biological processes, ranging from cell growth and organization of the actin cytoskeleton to endo- and exocytosis. Phosphorylation of Phosphatidylinositol at the D-4 position, an essential step in the biosynthesis of PIPs, appears to be catalyzed by two biochemically distinct enzymes. However, only one of these two enzymes has been molecularly characterized. We now describe a novel class of Phosphatidylinositol 4-Kinases that probably corresponds to the missing element in Phosphatidylinositol metabolism. These Kinases are highly conserved evolutionarily, but unrelated to previously characterized Phosphatidylinositol Kinases, and thus represent the founding members of a new family. The novel Phosphatidylinositol 4-Kinases, which are widely expressed in cells, only phosphorylate Phosphatidylinositol, are potently inhibited by adenosine, but are insensitive to wortmannin or phenylarsine oxide. Although they lack an obvious transmembrane domain, they are strongly attached to membranes by palmitoylation. Our data suggest that independent pathways for Phosphatidylinositol 4-phosphate synthesis emerged during evolution, possibly to allow tight temporal and spatial control over the production of this key signaling molecule.
H. U. Häring - One of the best experts on this subject based on the ideXlab platform.
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The insulin receptor: signalling mechanism and contribution to the pathogenesis of insulin resistance
Diabetologia, 1991Co-Authors: H. U. HäringAbstract:The insulin receptor is a heterotetrameric structure consisting of two α-subunits of M_r135 kilodalton on the outside of the plasma membrane connected by disulphide bonds to β-subunits of M_r95 kilodalton which are transmembrane proteins. Insulin binding to the α-subunit induces conformational changes which are transduced to the β-subunit. This leads to the activation of a tyrosine kinase activity which is intrinsic to the cytoplasmatic domains of the β-subunit. Activation of the tyrosine kinase activity of the insulin receptor represents an essential step in the transduction of an insulin signal across the plasma membrane of target cells. Signal transduction on the post-kinase level is not yet understood in detail, possible mechanisms involve phosphorylation of substrate proteins at tyrosine residues, activation of serine Kinases, the interaction with G-proteins, phospholipases and Phosphatidylinositol Kinases. Studies in multiple insulin-resistant cell models have demonstrated that an impaired response of the tyrosine kinase to insulin stimulation is one potential mechanism causing insulin resistance. An impairment of the insulin effect on tyrosine kinase activation in all major target tissues of insulin, in particular the skeletal muscle was demonstrated in Type 2 (non-insulin-dependent) diabetic patients. There is no evidence that the impaired tyrosine kinase response in the skeletal muscle is a primary defect, however, it is likely that this abnormality of insulin signal transduction contributes significantly to the pathogenesis of the insulin-resistant state in Type 2 diabetes.