Journal articles: 'Phosphatidylinositol 3-Kinases'

1

Abraham, Robert T. "Phosphatidylinositol 3-kinase related kinases." Current Opinion in Immunology 8, no. 3 (June 1996): 412–18. http://dx.doi.org/10.1016/s0952-7915(96)80132-4.

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Suzuki, Takahiro, Osamu Hazeki, and Michio Ui. "Phosphatidylinositol 3-Kinases." membrane 21, no. 3 (1996): 158–64. http://dx.doi.org/10.5360/membrane.21.158.

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Muftuoglu, Yagmur, Yi Xue, Xiang Gao, Dianqing Wu, and Ya Ha. "Mechanism of substrate specificity of phosphatidylinositol phosphate kinases." Proceedings of the National Academy of Sciences 113, no. 31 (July 20, 2016): 8711–16. http://dx.doi.org/10.1073/pnas.1522112113.

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The phosphatidylinositol phosphate kinase (PIPK) family of enzymes is primarily responsible for converting singly phosphorylated phosphatidylinositol derivatives to phosphatidylinositol bisphosphates. As such, these kinases are central to many signaling and membrane trafficking processes in the eukaryotic cell. The three types of phosphatidylinositol phosphate kinases are homologous in sequence but differ in catalytic activities and biological functions. Type I and type II kinases generate phosphatidylinositol 4,5-bisphosphate from phosphatidylinositol 4-phosphate and phosphatidylinositol 5-phosphate, respectively, whereas the type III kinase produces phosphatidylinositol 3,5-bisphosphate from phosphatidylinositol 3-phosphate. Based on crystallographic analysis of the zebrafish type I kinase PIP5Kα, we identified a structural motif unique to the kinase family that serves to recognize the monophosphate on the substrate. Our data indicate that the complex pattern of substrate recognition and phosphorylation results from the interplay between the monophosphate binding site and the specificity loop: the specificity loop functions to recognize different orientations of the inositol ring, whereas residues flanking the phosphate binding Arg244 determine whether phosphatidylinositol 3-phosphate is exclusively bound and phosphorylated at the 5-position. This work provides a thorough picture of how PIPKs achieve their exquisite substrate specificity.

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Rane, M. J., S. L. Carrithers, J. M. Arthur, J. B. Klein, and K. R. McLeish. "Formyl peptide receptors are coupled to multiple mitogen-activated protein kinase cascades by distinct signal transduction pathways: role in activation of reduced nicotinamide adenine dinucleotide oxidase." Journal of Immunology 159, no. 10 (November 15, 1997): 5070–78. http://dx.doi.org/10.4049/jimmunol.159.10.5070.

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Abstract Formyl peptide receptor activation of three mitogen-activated protein kinase (MAPK) cascades, extracellular signal-regulated kinases (ERKs), N-terminal kinases (JNKs), and p38 MAPK was examined in differentiated HL-60 granulocytes. FMLP stimulated a concentration- and time-dependent increase in ERK, JNK, and p38 MAPK activities, all of which were dependent on a pertussis toxin-sensitive G protein. Pharmacologic inhibitors were used to examine the roles of tyrosine kinases, phosphatidylinositol 3-kinase, protein kinase C, and phospholipase C. FMLP-stimulated ERK activity was dependent on tyrosine kinases, phosphatidylinositol 3-kinase, protein kinase C, and phospholipase C; p38 MAPK activation was dependent on phosphatidylinositol 3-kinase and phospholipase C; while JNK activation was independent of all of these signaling components. The mitogen-activated protein kinase/ERK kinase inhibitor PD098059 reduced ERK activation by 90%, while an inhibitor of p38 MAPK, SB203580, inhibited p38 MAPK activation by 80%. Both PD098059 and SB203580 inhibited FMLP-stimulated superoxide release, as did inhibitors directed against protein kinase C, tyrosine kinases, and phosphatidylinositol 3-kinase. We conclude that formyl peptide receptors are coupled to three MAPK cascades by Gi proteins. ERKs, p38 MAPK, and JNKs are each activated by distinct proximal signal transduction pathways. Activation of p38 MAPK is necessary for FMLP stimulation of respiratory burst activity; however, a second signal that may involve ERK is also required for this activity.

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Roymans, Dirk, and Herman Slegers. "Phosphatidylinositol 3-kinases in tumor progression." European Journal of Biochemistry 268, no. 3 (February 2001): 487–98. http://dx.doi.org/10.1046/j.1432-1327.2001.01936.x.

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Imseng, Stefan, Christopher HS Aylett, and Timm Maier. "Architecture and activation of phosphatidylinositol 3-kinase related kinases." Current Opinion in Structural Biology 49 (April 2018): 177–89. http://dx.doi.org/10.1016/j.sbi.2018.03.010.

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7

Shibasaki, F., Y. Fukui, and T. Takenawa. "Different properties of monomer and heterodimer forms of phosphatidylinositol 3-kinases." Biochemical Journal 289, no. 1 (January 1, 1993): 227–31. http://dx.doi.org/10.1042/bj2890227.

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Phosphatidylinositol (PI) 3-kinase plays an important role in the signalling of cell growth. We previously purified two types of PI 3-kinase from bovine thymus, a monomer from (PI 3-kinase I) and a heterodimer form (PI 3-kinase II) [Shibasaki, Homma and Takenawa (1991) J. Biol. Chem. 266, 8108-8114]. Here we examine the properties of these purified PI 3-kinases. Both PI 3-kinases were inhibited strongly by quercetin and isoquercetin. The inhibition of PI 3-kinase I and PI 3-kinase II by quercetin appears to be non-competitive, with apparent Ki values of 4 microM and 2.5 microM respectively. PI 3-kinase II, but not PI 3-kinase I, co-immunoprecipitates with pp60v-src and polyoma middle T (mT)/pp60c-src, even under conditions where the PI 3-kinases are not phosphorylated, suggesting that non-phosphorylated PI 3-kinase recognizes autophosphorylated pp60v-src. PI 3-kinase II is phosphorylated by pp60v-src and binds to it. Anti-p85 (85 kDa subunit of PI 3-kinase II) antibody precipitates not only PI 3-kinase II but also co-immunoprecipitates pp60v-src in src-transformed cells, suggesting that PI 3-kinase II binds to pp60v-src in vivo. These data suggest that the two PI 3-kinases may be regulated independently.

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Yamboliev, Ilia A., Kevin M. Wiesmann, Cherie A. Singer, Jason C. Hedges, and William T. Gerthoffer. "Phosphatidylinositol 3-kinases regulate ERK and p38 MAP kinases in canine colonic smooth muscle." American Journal of Physiology-Cell Physiology 279, no. 2 (August 1, 2000): C352—C360. http://dx.doi.org/10.1152/ajpcell.2000.279.2.c352.

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In canine colon, M2/M3 muscarinic receptors are coupled to extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein (MAP) kinases. We tested the hypothesis that this coupling is mediated by enzymes of the phosphatidylinositol (PI) 3-kinase family. RT-PCR and Western blotting demonstrated expression of two isoforms, PI 3-kinase-α and PI 3-kinase-γ. Muscarinic stimulation of intact muscle strips (10 μM ACh) activated PI 3-kinase-γ, ERK and p38 MAP kinases, and MAP kinase-activated protein kinase-2, whereas PI 3-kinase-α activation was not detected. Wortmannin (25 μM) abolished the activation of PI 3-kinase-γ, ERK, and p38 MAP kinases. MAP kinase inhibition was a PI 3-kinase-γ-specific effect, since wortmannin did not inhibit recombinant activated murine ERK2 MAP kinase, protein kinase C, Raf-1, or MAP kinase kinase. In cultured muscle cells, newborn calf serum (3%) activated PI 3-kinase-α and PI 3-kinase-γ isoforms, ERK and p38 MAP kinases, and stimulated chemotactic cell migration. Using wortmannin and LY-294002 to inhibit PI 3-kinase activity and PD-098059 and SB-203580 to inhibit ERK and p38 MAP kinases, we established that these enzymes are functionally important for regulation of chemotactic migration of colonic myocytes.

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Choi, Suyong, Xander Houdek, and Richard A. Anderson. "Phosphoinositide 3-kinase pathways and autophagy require phosphatidylinositol phosphate kinases." Advances in Biological Regulation 68 (May 2018): 31–38. http://dx.doi.org/10.1016/j.jbior.2018.02.003.

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Christoforidis, Savvas, Marta Miaczynska, Keith Ashman, Matthias Wilm, Liyun Zhao, Shu-Chin Yip, Michael D. Waterfield, Jonathan M. Backer, and Marino Zerial. "Phosphatidylinositol-3-OH kinases are Rab5 effectors." Nature Cell Biology 1, no. 4 (July 15, 1999): 249–52. http://dx.doi.org/10.1038/12075.

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11

Takegawa, K., D. B. DeWald, and S. D. Emr. "Schizosaccharomyces pombe Vps34p, a phosphatidylinositol-specific PI 3-kinase essential for normal cell growth and vacuole morphology." Journal of Cell Science 108, no. 12 (December 1, 1995): 3745–56. http://dx.doi.org/10.1242/jcs.108.12.3745.

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We have cloned the gene, vps34+, from the fission yeast Schizosaccharomyces pombe which encodes an 801 amino acid protein with phosphatidylinositol 3-kinase activity. The S. pombe Vps34 protein shares 43% amino acid sequence identity with the Saccharomyces cerevisiae Vps34 protein and 28% identity with the p110 catalytic subunit of the mammalian phosphatidylinositol 3-kinase. When the vps34+ gene is disrupted, S.pombe strains are temperature-sensitive for growth and the mutant cells contain enlarged vacuoles. Furthermore, while wild-type strains exhibit substantial levels of phosphatidylinositol 3-kinase activity, this activity is not detected in the vps34 delta strain. S.pombe Vps34p-specific antiserum detects a single protein in cells of -90 kDa that fractionates almost exclusively with the crude membrane fraction. Phosphatidylinositol 3-kinase activity also is localized mainly in the membrane fraction of wild-type cells. Immunoisolated Vps34p specifically phosphorylates phosphatidylinositol on the D-3 position of the inositol ring to yield phosphatidylinositol(3)phosphate. but does not utilize phosphatidylinositol(4)phosphate or phosphatidylinositol(4,5)bisphosphate as substrates. In addition, when compared to the mammalian p110 phosphatidylinositol 3-kinase, S. pombe Vps34p is relatively insensitive to the inhibitors wortmannin and LY294002. Together, these results indicate that S. pombe Vps34 is more similar to the phosphatidylinositol-specific 3-kinase, Vps34p from S. cerevisiae, and is distinct from the p110/p85 and G protein-coupled phosphatidylinositol 3-kinases from mammalian cells. These data are discussed in relation to the possible role of Vps34p in vesicle-mediated protein sorting to the S. pombe vacuole.

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12

Min, Sang H., and Charles S. Abrams. "Why do phosphatidylinositol kinases have so many isoforms?" Biochemical Journal 423, no. 1 (September 14, 2009): e5-e8. http://dx.doi.org/10.1042/bj20091274.

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Macromolecules can be transported into the cells by endocytosis, either by phagocytosis or by pinocytosis. Typically, phagocytosis involves the uptake of solid large particles mediated by cell-surface receptors, whereas pinocytosis takes up fluid and solutes. The synthesis of PtdIns(4,5)P2 and PtdIns(3,4,5)P3 plays fundamental roles in all forms of endocytosis. Curiously, almost all eukaryotic cells have multiple isoforms of the kinases that synthesize these critical phosphatidylinositols. In this issue of the Biochemical Journal, Namiko Tamura, Osamu Hazeki and co-workers report that the subunit p110α of the type I PI3K (phosphoinositide 3-kinase) is implicated in the phagocytosis and the pinocytosis of large molecules, whereas the receptor-mediated pinocytosis and micropinocytosis of small molecules do not seem to be controlled by this mechanism. The present commentary discusses recent literature that has begun to unravel why cells need so many phosphatidylinositol kinase isoforms, which were previously believed to be redundant.

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Vieira, Otilia V., Roberto J. Botelho, Lucia Rameh, Saskia M. Brachmann, Tsuyoshi Matsuo, Howard W. Davidson, Alan Schreiber, Jonathan M. Backer, Lewis C. Cantley, and Sergio Grinstein. "Distinct roles of class I and class III phosphatidylinositol 3-kinases in phagosome formation and maturation." Journal of Cell Biology 155, no. 1 (October 1, 2001): 19–26. http://dx.doi.org/10.1083/jcb.200107069.

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Phagosomes acquire their microbicidal properties by fusion with lysosomes. Products of phosphatidylinositol 3-kinase (PI 3-kinase) are required for phagosome formation, but their role in maturation is unknown. Using chimeric fluorescent proteins encoding tandem FYVE domains, we found that phosphatidylinositol 3-phosphate (PI[3]P) accumulates greatly but transiently on the phagosomal membrane. Unlike the 3′-phosphoinositides generated by class I PI 3-kinases which are evident in the nascent phagosomal cup, PI(3)P is only detectable after the phagosome has sealed. The class III PI 3-kinase VPS34 was found to be responsible for PI(3)P synthesis and essential for phagolysosome formation. In contrast, selective ablation of class I PI 3-kinase revealed that optimal phagocytosis, but not maturation, requires this type of enzyme. These results highlight the differential functional role of the two families of kinases, and raise the possibility that PI(3)P production by VPS34 may be targeted during the maturation arrest induced by some intracellular parasites.

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14

Mitra, Prasenjit, Yingjie Zhang, Lucia E. Rameh, Maria P. Ivshina, Dannel McCollum, John J. Nunnari, Gregory M. Hendricks, et al. "A novel phosphatidylinositol(3,4,5)P3 pathway in fission yeast." Journal of Cell Biology 166, no. 2 (July 12, 2004): 205–11. http://dx.doi.org/10.1083/jcb.200404150.

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The mammalian tumor suppressor, phosphatase and tensin homologue deleted on chromosome 10 (PTEN), inhibits cell growth and survival by dephosphorylating phosphatidylinositol-(3,4,5)-trisphosphate (PI[3,4,5]P3). We have found a homologue of PTEN in the fission yeast, Schizosaccharomyces pombe (ptn1). This was an unexpected finding because yeast (S. pombe and Saccharomyces cerevisiae) lack the class I phosphoinositide 3-kinases that generate PI(3,4,5)P3 in higher eukaryotes. Indeed, PI(3,4,5)P3 has not been detected in yeast. Surprisingly, upon deletion of ptn1 in S. pombe, PI(3,4,5)P3 became detectable at levels comparable to those in mammalian cells, indicating that a pathway exists for synthesis of this lipid and that the S. pombe ptn1, like mammalian PTEN, suppresses PI(3,4,5)P3 levels. By examining various mutants, we show that synthesis of PI(3,4,5)P3 in S. pombe requires the class III phosphoinositide 3-kinase, vps34p, and the phosphatidylinositol-4-phosphate 5-kinase, its3p, but does not require the phosphatidylinositol-3-phosphate 5-kinase, fab1p. These studies suggest that a pathway for PI(3,4,5)P3 synthesis downstream of a class III phosphoinositide 3-kinase evolved before the appearance of class I phosphoinositide 3-kinases.

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15

Tichý, Aleš, Kamila Ďurišová, Eva Novotná, Lenka Zárybnická, Jiřina Vávrová, Jaroslav Pejchal, and Zuzana Šinkorová. "PHOSPHATIDYLINOSITOL-3-KINASE RELATED KINASES (PIKKS) IN RADIATION-INDUCED DNA DAMAGE." Military Medical Science Letters 81, no. 4 (December 7, 2012): 177–87. http://dx.doi.org/10.31482/mmsl.2012.025.

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16

Gutkind, J. S., P. M. Lacal, and K. C. Robbins. "Thrombin-dependent association of phosphatidylinositol-3 kinase with p60c-src and p59fyn in human platelets." Molecular and Cellular Biology 10, no. 7 (July 1990): 3806–9. http://dx.doi.org/10.1128/mcb.10.7.3806-3809.1990.

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Recent studies have shown that ligand-activated growth factor receptors as well as transforming versions of nonreceptor protein-tyrosine kinases physically associate with phosphatidylinositol-3 kinase (PI-3 kinase). Reasoning that PI-3 kinase might also play a role in the normal functions of nonreceptor kinases, we sought to determine whether association with PI-3 kinase might serve as a measure of nonreceptor protein-tyrosine kinase activation under physiological conditions. We found that p60c-src as well as p59fyn, the product of another member of the src family of proto-oncogenes, physically associated with a PI kinase activity within 5 s after exposure to thrombin. Furthermore, PI kinase reaction products generated in p60v-src, p60c-src or p59fyn containing immunoprecipitates were indistinguishable, demonstrating the identity of the associated enzyme as PI-3 kinase. These findings demonstrate a thrombin-dependent interaction between p60c-src or p59fyn and PI-3 kinase and suggest a role for nonreceptor protein-tyrosine kinases in human platelet signal transduction.

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17

Gutkind, J. S., P. M. Lacal, and K. C. Robbins. "Thrombin-dependent association of phosphatidylinositol-3 kinase with p60c-src and p59fyn in human platelets." Molecular and Cellular Biology 10, no. 7 (July 1990): 3806–9. http://dx.doi.org/10.1128/mcb.10.7.3806.

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Recent studies have shown that ligand-activated growth factor receptors as well as transforming versions of nonreceptor protein-tyrosine kinases physically associate with phosphatidylinositol-3 kinase (PI-3 kinase). Reasoning that PI-3 kinase might also play a role in the normal functions of nonreceptor kinases, we sought to determine whether association with PI-3 kinase might serve as a measure of nonreceptor protein-tyrosine kinase activation under physiological conditions. We found that p60c-src as well as p59fyn, the product of another member of the src family of proto-oncogenes, physically associated with a PI kinase activity within 5 s after exposure to thrombin. Furthermore, PI kinase reaction products generated in p60v-src, p60c-src or p59fyn containing immunoprecipitates were indistinguishable, demonstrating the identity of the associated enzyme as PI-3 kinase. These findings demonstrate a thrombin-dependent interaction between p60c-src or p59fyn and PI-3 kinase and suggest a role for nonreceptor protein-tyrosine kinases in human platelet signal transduction.

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Takeda, Kosuke, Atsuo T. Sasaki, Hyunjung Ha, Hyun-A. Seung, and Richard A. Firtel. "Role of Phosphatidylinositol 3-Kinases in Chemotaxis inDictyostelium." Journal of Biological Chemistry 282, no. 16 (March 1, 2007): 11874–84. http://dx.doi.org/10.1074/jbc.m610984200.

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McNulty, Shannon, William Bornmann, Jill Schriewer, Chas Werner, Scott K. Smith, Victoria A. Olson, Inger K. Damon, R. Mark Buller, John Heuser, and Daniel Kalman. "Multiple Phosphatidylinositol 3-Kinases Regulate Vaccinia Virus Morphogenesis." PLoS ONE 5, no. 5 (May 28, 2010): e10884. http://dx.doi.org/10.1371/journal.pone.0010884.

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20

Li, Z., M. I. Wahl, A. Eguinoa, L. R. Stephens, P. T. Hawkins, and O. N. Witte. "Phosphatidylinositol 3-kinase- activates Bruton's tyrosine kinase in concert with Src family kinases." Proceedings of the National Academy of Sciences 94, no. 25 (December 9, 1997): 13820–25. http://dx.doi.org/10.1073/pnas.94.25.13820.

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21

Gillooly, David J., Anne Simonsen, and Harald Stenmark. "Phosphoinositides and phagocytosis." Journal of Cell Biology 155, no. 1 (October 1, 2001): 15–18. http://dx.doi.org/10.1083/jcb.200109001.

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Phosphoinositide 3 kinases (PI3Ks)**Abbreviation used in this paper: PI3K, phosphoinositide 3 kinase. are known as regulators of phagocytosis. Recent results demonstrate that class I and III PI3Ks act consecutively in phagosome formation and maturation, and that their respective products, phosphatidylinositol 3,4,5-trisphosphate (PI[3,4,5]P3) and phosphatidylinositol 3-phosphate (PI[3]P), accumulate transiently at different stages. Phagosomes containing Mycobacterium tuberculosis do not acquire the PI(3)P-binding protein EEA1, which is required for phagosome maturation. This suggests a possible mechanism of how this microorganism evades degradation in phagolysosomes.

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NAKAGAWA, Tamotsu, Kaoru GOTO, and Hisatake KONDO. "Cloning and characterization of a 92 kDa soluble phosphatidylinositol 4-kinase." Biochemical Journal 320, no. 2 (December 1, 1996): 643–49. http://dx.doi.org/10.1042/bj3200643.

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A phosphatidylinositol (PtdIns) 4-kinase cDNA cloned from a rat brain cDNA library encoded a protein of 816 amino acids with a calculated molecular mass of 91654 Da. This molecule contained a lipid-kinase-unique domain and a presumed lipid/protein kinase homology domain that are found in other PtdIns 4-kinases and PtdIns 3-kinases. Furthermore, this kinase molecule had 43.3% shared identity with the presumed catalytic domain of yeast PtdIns 4-kinase, PtdInsK1, and the two molecules had a region of similarity that is not conserved in other lipid kinases. By examining PtdIns kinase activity in transfected COS-7 cells using epitope tag immunoprecipitation as well as conventional methods, the product PtdIns phosphate was identified as phosphatidylinositol 4-phosphate (PtdIns4P), but not phosphatidylinositol 3-phosphate (PtdIns3P). The PtdIns 4-kinase activity was recovered predominantly from the soluble fraction and the activity was markedly enhanced in the presence of Triton X-100 and was relatively insensitive to inhibition by adenosine. In addition, the PtdIns 4-kinase activity was completely inhibited in the presence of 10 µM wortmannin. When examined by epitope tag immunocytochemistry, the immunoreactivity for the PtdIns 4-kinase molecule was dominantly aggregated in a cytoplasmic region juxtaposed to the nuclei and was faintly but widely dispersed in the cytoplasm. By in situ hybridization analysis, the mRNA for PtdIns 4-kinase was expressed ubiquitously and was detected in most neurons throughout the grey matter of the brain, with higher expression intensity found in fetal than in adult brain.

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Shin, Hye-Won, Mitsuko Hayashi, Savvas Christoforidis, Sandra Lacas-Gervais, Sebastian Hoepfner, Markus R. Wenk, Jan Modregger, et al. "An enzymatic cascade of Rab5 effectors regulates phosphoinositide turnover in the endocytic pathway." Journal of Cell Biology 170, no. 4 (August 15, 2005): 607–18. http://dx.doi.org/10.1083/jcb.200505128.

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Generation and turnover of phosphoinositides (PIs) must be coordinated in a spatial- and temporal-restricted manner. The small GTPase Rab5 interacts with two PI 3-kinases, Vps34 and PI3Kβ, suggesting that it regulates the production of 3-PIs at various stages of the early endocytic pathway. Here, we discovered that Rab5 also interacts directly with PI 5- and PI 4-phosphatases and stimulates their activity. Rab5 regulates the production of phosphatidylinositol 3-phosphate (PtdIns[3]P) through a dual mechanism, by directly phosphorylating phosphatidylinositol via Vps34 and by a hierarchical enzymatic cascade of phosphoinositide-3-kinaseβ (PI3Kβ), PI 5-, and PI 4-phosphatases. The functional importance of such an enzymatic pathway is demonstrated by the inhibition of transferrin uptake upon silencing of PI 4-phosphatase and studies in weeble mutant mice, where deficiency of PI 4-phosphatase causes an increase of PtdIns(3,4)P2 and a reduction in PtdIns(3)P. Activation of PI 3-kinase at the plasma membrane is accompanied by the recruitment of Rab5, PI 4-, and PI 5-phosphatases to the cell cortex. Our data provide the first evidence for a dual role of a Rab GTPase in regulating both generation and turnover of PIs via PI kinases and phosphatases to coordinate signaling functions with organelle homeostasis.

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Thi, E. P., and N. E. Reiner. "Phosphatidylinositol 3-kinases and their roles in phagosome maturation." Journal of Leukocyte Biology 92, no. 3 (May 8, 2012): 553–66. http://dx.doi.org/10.1189/jlb.0212053.

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Todorov, Alex G., Marcelo Einicker-Lamas, Solange L. de Castro, Mecia M. Oliveira, and Adilson Guilherme. "Activation of Host Cell Phosphatidylinositol 3-Kinases byTrypanosoma cruziInfection." Journal of Biological Chemistry 275, no. 41 (July 25, 2000): 32182–86. http://dx.doi.org/10.1074/jbc.m909440199.

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Zhou, Q. L., J. G. Park, Z. Y. Jiang, J. J. Holik, P. Mitra, S. Semiz, A. Guilherme, et al. "Analysis of insulin signalling by RNAi-based gene silencing." Biochemical Society Transactions 32, no. 5 (October 26, 2004): 817–21. http://dx.doi.org/10.1042/bst0320817.

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Using siRNA-mediated gene silencing in cultured adipocytes, we have dissected the insulin-signalling pathway leading to translocation of GLUT4 glucose transporters to the plasma membrane. RNAi (RNA interference)-based depletion of components in the putative TC10 pathway (CAP, CrkII and c-Cbl plus Cbl-b) or the phospholipase Cγ pathway failed to diminish insulin signalling to GLUT4. Within the phosphoinositide 3-kinase pathway, loss of the 5′-phosphatidylinositol 3,4,5-trisphosphate phosphatase SHIP2 was also without effect, whereas depletion of the 3′-phosphatase PTEN significantly enhanced insulin action. Downstream of phosphatidylinositol 3,4,5-trisphosphate and PDK1, silencing the genes encoding the protein kinases Akt1/PKBα, or CISK(SGK3) or protein kinases Cλ/ζ had little or no effect, but loss of Akt2/PKBβ significantly attenuated GLUT4 regulation by insulin. These results show that Akt2/PKBβ is the key downstream intermediate within the phosphoinositide 3-kinase pathway linked to insulin action on GLUT4 in cultured adipocytes, whereas PTEN is a potent negative regulator of this pathway.

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Jeschke, Andreas, Nicole Zehethofer, Buko Lindner, Jessica Krupp, Dominik Schwudke, Ina Haneburger, Marko Jovic, et al. "Phosphatidylinositol 4-phosphate and phosphatidylinositol 3-phosphate regulate phagolysosome biogenesis." Proceedings of the National Academy of Sciences 112, no. 15 (March 30, 2015): 4636–41. http://dx.doi.org/10.1073/pnas.1423456112.

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Professional phagocytic cells ingest microbial intruders by engulfing them into phagosomes, which subsequently mature into microbicidal phagolysosomes. Phagosome maturation requires sequential fusion of the phagosome with early endosomes, late endosomes, and lysosomes. Although various phosphoinositides (PIPs) have been detected on phagosomes, it remained unclear which PIPs actually govern phagosome maturation. Here, we analyzed the involvement of PIPs in fusion of phagosomes with various endocytic compartments and identified phosphatidylinositol 4-phosphate [PI(4)P], phosphatidylinositol 3-phosphate [PI(3)P], and the lipid kinases that generate these PIPs, as mediators of phagosome–lysosome fusion. Phagosome–early endosome fusion required PI(3)P, yet did not depend on PI(4)P. Thus, PI(3)P regulates phagosome maturation at early and late stages, whereas PI(4)P is selectively required late in the pathway.

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Yamboliev, Ilia A., Jennifer Chen, and William T. Gerthoffer. "PI 3-kinases and Src kinases regulate spreading and migration of cultured VSMCs." American Journal of Physiology-Cell Physiology 281, no. 2 (August 1, 2001): C709—C718. http://dx.doi.org/10.1152/ajpcell.2001.281.2.c709.

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Pulmonary artery smooth muscle cell (PASMC) adhesion, spreading, and migration depend on matrix-stimulated reorganization of focal adhesions. Platelet-derived growth factor (PDGF) activates intracellular signal transduction cascades that also regulate adhesion, spreading, and migration, but the signaling molecules involved in these events are poorly defined. We hypothesized that phosphatidylinositol (PI) 3-kinases and Src tyrosine kinases translate matrix and PDGF-initiated signals into cell motility. In experiments with cultured canine PASMCs, inhibition of PI 3-kinases with wortmannin (0.3 μM) and LY-294002 (50 μM) and of Src kinase with PP1 (30 μM) did not decrease spontaneous (nonstimulated) or PDGF-stimulated (10 ng/ml) adhesion onto collagen. PI 3-kinase and Src kinase activities, however, were necessary for cell spreading: PP1 inhibited cell spreading and Src Tyr-418 phosphorylation in a concentration-dependent manner. Inhibition of PI 3-kinase and Src partially reduced cell migration, while at 10 and 30 μM, PP1 eliminated migration, likely due to inhibition of PDGF receptors. In conclusion, both PI 3-kinases and Src tyrosine kinases are components of pathways that mediate spreading and migration of cultured PASMCs on collagen.

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Graziani, A., L. E. Ling, G. Endemann, C. L. Carpenter, and L. C. Cantley. "Purification and characterization of human erythrocyte phosphatidylinositol 4-kinase. Phosphatidylinositol 4-kinase and phosphatidylinositol 3-monophosphate 4-kinase are distinct enzymes." Biochemical Journal 284, no. 1 (May 15, 1992): 39–45. http://dx.doi.org/10.1042/bj2840039.

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PtdIns 4-kinase has been purified 83,000-fold from human erythrocyte membranes. The major protein detected by SDS/PAGE is of molecular mass 56 kDa, and enzymic activity can be renatured from this band of the gel. The characteristics of this enzyme are similar to other type II PtdIns kinases previously described: PtdIns presented in Triton X-100 micelles is preferred as a substrate over PtdIns vesicles, the enzyme possesses a relatively low Km for ATP (20 microM), and adenosine is an effective inhibitor. A monoclonal antibody raised against bovine brain type II PtdIns 4-kinase is an effective inhibitor of the purified enzyme. PtdIns(4,5)P2 inhibits by approx. 50% when added in equimolar amounts with PtdIns; PtdIns4P has little effect on activity. A PtdIns3P 4-kinase activity has also been detected in erythrocyte lysates. Approximately two-thirds of this activity is in the cytosolic fraction and one-third in the membrane fraction. No PtdIns3P 4-kinase activity could be detected in the purified type II PtdIns 4-kinase preparation, nor could this activity be detected in a bovine brain type III PtdIns 4-kinase preparation. The monoclonal antibody that inhibits the type II PtdIns 4-kinase does not affect the PtdIns3P 4-kinase activity in the membrane fraction. The cytosolic PtdIns3P 4-kinase can be efficiently recovered from a 60%-satd.-(NH4)2SO4 precipitate that is virtually free of PtdIns 4-kinase activity. We conclude that PtdIns3P 4-kinase is a new enzyme distinct from previously characterized PtdIns 4-kinases, and that this enzyme prefers PtdIns3P over PtdIns as a substrate.

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Liu, X., L. E. Marengere, C. A. Koch, and T. Pawson. "The v-Src SH3 domain binds phosphatidylinositol 3'-kinase." Molecular and Cellular Biology 13, no. 9 (September 1993): 5225–32. http://dx.doi.org/10.1128/mcb.13.9.5225-5232.1993.

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Fibroblasts transformed by v-src or by related oncogenes encoding activated tyrosine kinases contain elevated levels of polyphosphoinositides with phosphate at the D-3 position of the inositol ring, as a result of the activation of phosphatidylinositol (PI) 3'-kinase. v-src-transformed cells also contain increased levels of PI 3'-kinase activity immunoprecipitable with anti-phosphotyrosine antibodies; furthermore, PI 3'-kinase can be detected in association with the v-Src tyrosine kinase. To identify regions of v-Src that can interact with PI 3'-kinase, the v-Src SH2 and SH3 domains were expressed in bacteria and incubated with lysates of normal chicken embryo fibroblasts. In vitro, the v-Src SH3 domain, but not the SH2 domain, bound PI 3'-kinase in lysates of uninfected chicken embryo fibroblasts. Substitutions of two highly conserved SH3 residues implicated in ligand binding abolished the ability of the v-Src SH3 domain to associate with PI 3'-kinase. Furthermore, the v-Src SH3 domain bound in vitro to the amino-terminal region of the p85 alpha subunit of PI 3'-kinase. These results suggest that the v-Src SH3 domain may mediate an interaction between the v-Src tyrosine kinase and PI 3'-kinase, by direct binding to p85.

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Liu, X., L. E. Marengere, C. A. Koch, and T. Pawson. "The v-Src SH3 domain binds phosphatidylinositol 3'-kinase." Molecular and Cellular Biology 13, no. 9 (September 1993): 5225–32. http://dx.doi.org/10.1128/mcb.13.9.5225.

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Fibroblasts transformed by v-src or by related oncogenes encoding activated tyrosine kinases contain elevated levels of polyphosphoinositides with phosphate at the D-3 position of the inositol ring, as a result of the activation of phosphatidylinositol (PI) 3'-kinase. v-src-transformed cells also contain increased levels of PI 3'-kinase activity immunoprecipitable with anti-phosphotyrosine antibodies; furthermore, PI 3'-kinase can be detected in association with the v-Src tyrosine kinase. To identify regions of v-Src that can interact with PI 3'-kinase, the v-Src SH2 and SH3 domains were expressed in bacteria and incubated with lysates of normal chicken embryo fibroblasts. In vitro, the v-Src SH3 domain, but not the SH2 domain, bound PI 3'-kinase in lysates of uninfected chicken embryo fibroblasts. Substitutions of two highly conserved SH3 residues implicated in ligand binding abolished the ability of the v-Src SH3 domain to associate with PI 3'-kinase. Furthermore, the v-Src SH3 domain bound in vitro to the amino-terminal region of the p85 alpha subunit of PI 3'-kinase. These results suggest that the v-Src SH3 domain may mediate an interaction between the v-Src tyrosine kinase and PI 3'-kinase, by direct binding to p85.

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SCHNYDER, Bruno, Paul C. MEUNIER, and Bruce D. CAR. "Inhibition of kinases impairs neutrophil activation and killing of Staphylococcus aureus." Biochemical Journal 331, no. 2 (April 15, 1998): 489–95. http://dx.doi.org/10.1042/bj3310489.

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Intracellular phosphorylations polymorphonuclear neutrophils are mediated by kinases, including mitogen activated-protein (MAP) kinases and phosphatidylinositol 3-kinase. In the present study we demonstrate their effector functions upon both ligation of cell-surface seven-transmembrane-spanning receptors by bacterial peptide formylmethionyl-leucylphenylalanine as well as in the process of destruction of Staphylococcus aureus. To regulate neutrophil MAP kinases p38 and p44/42, specifically, we made use of their specific inhibitors 10 µM SK&F 86002 (for p38) and PD 098059 (for activating kinase of p44/42). SK&F 86002 was a potent inhibitor (by 70%) of induced antimicrobial oxygen-radical generation compared with PD 098059 (by 20%). SK&F 86002 and PD 098059 inhibited mobilization of a dominant neutrophil adhesion molecule, β2 integrin, from cytoplasmic granules to the plasma membrane by 40 and 10% respectively, and the combination of the two drugs resulted in a 90% effect. The combined effect of both drugs was moderate inhibition of bacterial destruction, despite the fact that neither compound had detectable effect on bactericidal activity if applied individually. Bacterial destruction was also inhibited by wortmannin (0.1 µM), the specific inhibitor of phosphatidylinositol 3-kinase, which had previously been described to target various other activations of the neutrophil, including oxygen-radical generation. Although the relative contribution of p38 and p44/42 MAP kinases varied, the marked effects of the combined inhibition of the kinases revealed their concerted actions to be critical for normal neutrophil function.

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Velnati, Suresh, Sara Centonze, Federico Girivetto, Daniela Capello, Ricardo M. Biondi, Alessandra Bertoni, Roberto Cantello, et al. "Identification of Key Phospholipids That Bind and Activate Atypical PKCs." Biomedicines 9, no. 1 (January 6, 2021): 45. http://dx.doi.org/10.3390/biomedicines9010045.

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PKCζ and PKCι/λ form the atypical protein kinase C subgroup, characterised by a lack of regulation by calcium and the neutral lipid diacylglycerol. To better understand the regulation of these kinases, we systematically explored their interactions with various purified phospholipids using the lipid overlay assays, followed by kinase activity assays to evaluate the lipid effects on their enzymatic activity. We observed that both PKCζ and PKCι interact with phosphatidic acid and phosphatidylserine. Conversely, PKCι is unique in binding also to phosphatidylinositol-monophosphates (e.g., phosphatidylinositol 3-phosphate, 4-phosphate, and 5-phosphate). Moreover, we observed that phosphatidylinositol 4-phosphate specifically activates PKCι, while both isoforms are responsive to phosphatidic acid and phosphatidylserine. Overall, our results suggest that atypical Protein kinase C (PKC) localisation and activity are regulated by membrane lipids distinct from those involved in conventional PKCs and unveil a specific regulation of PKCι by phosphatidylinositol-monophosphates.

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Lingaraj, Trupti, John Donovan, Zhi Li, Ping Li, Amanda Doucette, Sean Harrison, Jeffrey A. Ecsedy, Lenny Dang, and Wenhai Zhang. "A High-Throughput Liposome Substrate Assay with Automated Lipid Extraction Process for PI 3-Kinase." Journal of Biomolecular Screening 13, no. 9 (September 23, 2008): 906–11. http://dx.doi.org/10.1177/1087057108324498.

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The signaling pathways involving lipid kinase class I phosphatidylinositol 3-kinases (PI 3-kinases) regulate cell growth, proliferation, and survival. Class I PI 3-kinases catalyze the conversion of PI (4,5)P2 to PI (3,4,5)P3, which acts as a lipid second messenger to activate mitogenic signaling cascades. Recently, p110α, a class IA PI 3-kinase, was found to be mutated frequently in many human cancers. Therefore, it is increasingly studied as an anticancer drug target. Traditionally, PI 3-kinase activities have been studied using liposome substrates. This method, however, is hampered significantly by the labor-intensive manual lipid extraction followed by a low-throughput thin-layer chromatography analysis. The authors describe a high-throughput liposome substrate-based assay based on an automated lipid extraction method that allows them to study PI 3-kinase enzyme mechanism and quantitatively measure inhibitor activity using liposome substrates in a high-throughput mode. This improved assay format can easily be extended to study other classes of phosphoinositide lipid kinases. ( Journal of Biomolecular Screening 2008:906-911)

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KRISTIANSEN, S⊘ren, Toolsie RAMLAL, and Amira KLIP. "Phosphatidylinositol 4-kinase, but not phosphatidylinositol 3-kinase, is present in GLUT4-containing vesicles isolated from rat skeletal muscle." Biochemical Journal 335, no. 2 (October 15, 1998): 351–56. http://dx.doi.org/10.1042/bj3350351.

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Insulin stimulates the rate of glucose transport into muscle and adipose cells by translocation of glucose transporter (GLUT4)-containing vesicles from an intracellular storage pool to the surface membrane. This event is mediated through the insulin receptor substrates (IRSs), which in turn activate phosphatidylinositol (PI) 3-kinase isoforms. It has been suggested that insulin causes attachment of PI 3-kinases to the intracellular GLUT4-containing vesicles in rat adipose cells. Furthermore, it has also been shown that GLUT4-containing vesicles in adipose cells contain a PI 4-kinase. In the present study we investigate whether GLUT4-containing vesicles isolated from rat skeletal muscle display PI 3-kinase and/or PI 4-kinase activities. Insulin stimulation caused a rapid increase (5–15-fold increase compared with control) in the intracellular cytosolic IRS-1-associated PI-3 kinase activity. This PI 3-kinase activity was also present in a membrane preparation containing the insulin-regulatable pool of GLUT4 transporters. However, when GLUT4-containing vesicles were isolated by immunoprecipitation from basal and insulin-stimulated (3 min) skeletal muscle, the vesicles displayed PI 4-kinase, but not PI 3-kinase, activity. Insulin did not regulate the PI 4-kinase activity in the GLUT4-containing vesicles. In conclusion, GLUT4-containing vesicles from rat skeletal muscle contain a PI 4-kinase, but not a PI 3-kinase. It is suggested that, in skeletal muscle, insulin causes activation of the IRS/PI 3-kinase complex in an intracellular membrane compartment associated closely with the GLUT4-containing vesicles, but not in the GLUT4-containing vesicles themselves.

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Sugimoto, Katsunori. "Branching the Tel2 pathway for exact fit on phosphatidylinositol 3-kinase-related kinases." Current Genetics 64, no. 5 (February 22, 2018): 965–70. http://dx.doi.org/10.1007/s00294-018-0817-9.

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37

Bony, Claire, Serge Roche, Ueno Shuichi, Takehiko Sasaki, Michael A. Crackower, Josef Penninger, Hiroyuki Mano, and Michel Pucéat. "A Specific Role of Phosphatidylinositol 3–Kinase γ." Journal of Cell Biology 152, no. 4 (February 19, 2001): 717–28. http://dx.doi.org/10.1083/jcb.152.4.717.

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Purinergic stimulation of cardiomyocytes turns on a Src family tyrosine kinase–dependent pathway that stimulates PLCγ and generates IP3, a breakdown product of phosphatidylinositol 4,5–bisphosphate (PIP2). This signaling pathway closely regulates cardiac cell autonomic activity (i.e., spontaneous cell Ca2+ spiking). PIP2 is phosphorylated on 3′ by phosphoinositide 3–kinases (PI3Ks) that belong to a broad family of kinase isoforms. The product of PI3K, phosphatidylinositol 3,4,5–trisphosphate, regulates activity of PLCγ. PI3Ks have emerged as crucial regulators of many cell functions including cell division, cell migration, cell secretion, and, via PLCγ, Ca2+ homeostasis. However, although PI3Kα and -β have been shown to mediate specific cell functions in nonhematopoietic cells, such a role has not been found yet for PI3Kγ. We report that neonatal rat cardiac cells in culture express PI3Kα, -β, and -γ. The purinergic agonist predominantly activates PI3Kγ. Both wortmannin and LY294002 prevent tyrosine phosphorylation, and membrane translocation of PLCγ as well as IP3 generation in ATP-stimulated cells. Furthermore, an anti-PI3Kγ, but not an anti-PI3Kβ, injected in the cells prevents the effect of ATP on cell Ca2+ spiking. A dominant negative mutant of PI3Kγ transfected in the cells also exerts the same action. The effect of ATP was observed on spontaneous Ca2+ spiking of wild-type but not of PI3Kγ2/2 embryonic stem cell–derived cardiomyocytes. ATP activates the Btk tyrosine kinase, Tec, and induces its association with PLCγ. A dominant negative mutant of Tec blocks the purinergic effect on cell Ca2+ spiking. Tec is translocated to the T-tubes upon ATP stimulation of cardiac cells. Both an anti-PI3Kγ antibody and a dominant negative mutant of PI3Kγ injected or transfected into cells prevent the latter event. We conclude that PI3Kγ activation is a crucial step in the purinergic regulation of cardiac cell spontaneous Ca2+ spiking. Our data further suggest that Tec works in concert with a Src family kinase and PI3Kγ to fully activate PLCγ in ATP-stimulated cardiac cells. This cluster of kinases provides the cardiomyocyte with a tight regulation of IP3 generation and thus cardiac autonomic activity.

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Endemann, G. C., A. Graziani, and L. C. Cantley. "A monoclonal antibody distinguishes two types of phosphatidylinositol 4-kinase." Biochemical Journal 273, no. 1 (January 1, 1991): 63–66. http://dx.doi.org/10.1042/bj2730063.

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A monoclonal antibody has been developed against the type II PtdIns 4-kinase from bovine brain. This antibody, 4C5G, causes greater than 90% inhibition of the type II PtdIns 4-kinase from bovine brain, rat brain and human erythrocytes. However, it fails to inhibit type III PtdIns 4-kinase from bovine brain or PtdIns 3-kinase from rat liver. These results suggest that type II and type III PtdIns 4-kinases are distinct gene products, and that 4C5G will be useful in studying the function of the type II PtdIns 4-kinase.

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Yoshioka, Kazuaki. "Class II phosphatidylinositol 3-kinase isoforms in vesicular trafficking." Biochemical Society Transactions 49, no. 2 (March 5, 2021): 893–901. http://dx.doi.org/10.1042/bst20200835.

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Phosphatidylinositol 3-kinases (PI3Ks) are critical regulators of many cellular processes including cell survival, proliferation, migration, cytoskeletal reorganization, and intracellular vesicular trafficking. They are a family of lipid kinases that phosphorylate membrane phosphoinositide lipids at the 3′ position of their inositol rings, and in mammals they are divided into three classes. The role of the class III PI3K Vps34 is well-established, but recent evidence suggests the physiological significance of class II PI3K isoforms in vesicular trafficking. This review focuses on the recently discovered functions of the distinct PI3K-C2α and PI3K-C2β class II PI3K isoforms in clathrin-mediated endocytosis and consequent endosomal signaling, and discusses recently reported data on class II PI3K isoforms in different physiological contexts in comparison with class I and III isoforms.

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40

Boronenkov, Igor V., Joost C. Loijens, Masato Umeda, and Richard A. Anderson. "Phosphoinositide Signaling Pathways in Nuclei Are Associated with Nuclear Speckles Containing Pre-mRNA Processing Factors." Molecular Biology of the Cell 9, no. 12 (December 1998): 3547–60. http://dx.doi.org/10.1091/mbc.9.12.3547.

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Phosphoinositide signal transduction pathways in nuclei use enzymes that are indistinguishable from their cytosolic analogues. We demonstrate that distinct phosphatidylinositol phosphate kinases (PIPKs), the type I and type II isoforms, are concentrated in nuclei of mammalian cells. The cytosolic and nuclear PIPKs display comparable activities toward the substrates phosphatidylinositol 4-phosphate and phosphatidylinositol 3-phosphate. Indirect immunofluorescence revealed that these kinases were associated with distinct subnuclear domains, identified as “nuclear speckles,” which also contained pre-mRNA processing factors. A pool of nuclear phosphatidylinositol bisphosphate (PIP2), the product of these kinases, was also detected at these same sites by monoclonal antibody staining. The localization of PIPKs and PIP2 to speckles is dynamic in that both PIPKs and PIP2 reorganize along with other speckle components upon inhibition of mRNA transcription. Because PIPKs have roles in the production of most phosphatidylinositol second messengers, these findings demonstrate that phosphatidylinositol signaling pathways are localized at nuclear speckles. Surprisingly, the PIPKs and PIP2 are not associated with invaginations of the nuclear envelope or any nuclear membrane structure. The putative absence of membranes at these sites suggests novel mechanisms for the generation of phosphoinositides within these structures.

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King, W. G., M. D. Mattaliano, T. O. Chan, P. N. Tsichlis, and J. S. Brugge. "Phosphatidylinositol 3-kinase is required for integrin-stimulated AKT and Raf-1/mitogen-activated protein kinase pathway activation." Molecular and Cellular Biology 17, no. 8 (August 1997): 4406–18. http://dx.doi.org/10.1128/mcb.17.8.4406.

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Cell attachment to fibronectin stimulates the integrin-dependent interaction of p85-associated phosphatidylinositol (PI) 3-kinase with integrin-dependent focal adhesion kinase (FAK) as well as activation of the Ras/mitogen-activated protein (MAP) kinase pathway. However, it is not known if this PI 3-kinase-FAK interaction increases the synthesis of the 3-phosphorylated phosphoinositides (3-PPIs) or what role, if any, is played by activated PI 3-kinase in integrin signaling. We demonstrate here the integrin-dependent accumulation of the PI 3-kinase products, PI 3,4-bisphosphate [PI(3,4)P2] and PI(3,4,5)P3, as well as activation of AKT kinase, a serine/threonine kinase that can be stimulated by binding of PI(3,4)P2. The PI 3-kinase inhibitors wortmannin and LY294002 significantly decreased the integrin-induced accumulation of the 3-PPIs and activation of AKT kinase, without having significant effects on the levels of PI(4,5)P2 or tyrosine phosphorylation of paxillin. These inhibitors also reduced cell adhesion/spreading onto fibronectin but had no effect on attachment to polylysine. Interestingly, integrin-mediated Erk-2, Mek-1, and Raf-1 activation, but not Ras-GTP loading, was inhibited at least 80% by wortmannin and LY294002. In support of the pharmacologic results, fibronectin activation of Erk-2 and AKT kinases was completely inhibited by overexpression of a dominant interfering p85 subunit of PI 3-kinase. We conclude that integrin-mediated adhesion to fibronectin results in the accumulation of the PI 3-kinase products PI(3,4)P2 and PI(3,4,5)P3 as well as the PI 3-kinase-dependent activation of the kinases Raf-1, Mek-1, Erk-2, and AKT and that PI 3-kinase may function upstream of Raf-1 but downstream of Ras in integrin activation of Erk-2 MAP and AKT kinases.

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Shen, Dadong, Jinlin Zhu, Guofeng Wu, Li Sheng, Haoling Gao, and Pu Wang. "Development of Synthesis of Phosphatidylinositol 3-Kinases Inhibitor Puquitinib Mesylate." Chinese Journal of Organic Chemistry 39, no. 9 (2019): 2676. http://dx.doi.org/10.6023/cjoc201902020.

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43

Gypakis, A., and H. K. Wasner. "Phosphatidylinositol 3-Kinase and Prostaglandylinositol Cyclic Phosphate (Cyclic PIP), a Mediator of Insulin Action, in the Signal Transduction of Insulin." Biological Chemistry 381, no. 11 (November 15, 2000): 1139–41. http://dx.doi.org/10.1515/bc.2000.140.

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Abstract It has been suggested that downstream signaling from the insulin receptor to the level of the protein kinases and protein phosphatases is accomplished by prostaglandylinositol cyclic phosphate (cyclic PIP), a proposed second messenger of insulin. However, evidence points also to both phosphatidylinositol 3-kinase, which binds to the tyrosine phosphorylated insulin receptor substrate-1, and the Ras complex in insulin's downstream signaling. We have examined whether a correlation exists between these various observations. It was found that wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase, prevented insulin-induced, as well as cyclic PIP-induced activation of glucose transport, indicating that PI 3-kinase action on glucose transport involves downstream signaling of both insulin and cyclic PIP. Wortmannin has no effect on cyclic PIP synthase activity nor on the substrate production for cyclic PIP synthesis either, indicating that the functional role of PI 3-kinase is exclusively downstream of cyclic PIP.

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COFFER, Paul J., Jing JIN, and James R. WOODGETT. "Protein kinase B (c-Akt): a multifunctional mediator of phosphatidylinositol 3-kinase activation." Biochemical Journal 335, no. 1 (October 1, 1998): 1–13. http://dx.doi.org/10.1042/bj3350001.

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While a plethora of extracellular molecules exist that modulate cellular functions via binding to membrane receptors inside the cell, their actions are mediated by relatively few signalling mechanisms. One of these is activation of phosphatidylinositol 3-kinase (PI-3K), which results in the generation of a membrane-restricted second messenger, polyphosphatidylinositides containing a 3´-phosphate. How these molecules transduced the effects of agonists of PI-3K was unclear until the recent discovery that several protein kinases become activated upon exposure to 3´-phosphorylated inositol lipids. These enzymes include protein kinase B (PKB)/AKT and PtdIns(3,4,5)P3-dependent kinases 1 and 2, the first two of which interact with 3´-phosphorylated phosphoinositides via pleckstrin homology domains. Once targeted to the membrane by this motif, PKB becomes phosphorylated at two residues, which relieves intermolecular inhibition, allowing the activated complex to dissociate and modify its targets. Identification of these substrates is the subject of intensive research, since at least one must play a key role in suppressing apoptosis, as demonstrated by expression of activated alleles of PKB. The generation of effective transdominant mutants, coupled with genetic analysis of the protein kinase in simpler organisms, should help in elucidating outstanding questions in the functions, targets and regulation of this important mediator of PI-3K signalling.

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Chapman, Carolyn Riley, Sarah Tyler Evans, Antony M. Carr, and Tamar Enoch. "Requirement of Sequences outside the Conserved Kinase Domain of Fission Yeast Rad3p for Checkpoint Control." Molecular Biology of the Cell 10, no. 10 (October 1999): 3223–38. http://dx.doi.org/10.1091/mbc.10.10.3223.

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The fission yeast Rad3p checkpoint protein is a member of the phosphatidylinositol 3-kinase-related family of protein kinases, which includes human ATMp. Mutation of the ATMgene is responsible for the disease ataxia-telangiectasia. The kinase domain of Rad3p has previously been shown to be essential for function. Here, we show that although this domain is necessary, it is not sufficient, because the isolated kinase domain does not have kinase activity in vitro and cannot complement a rad3 deletion strain. Using dominant negative alleles of rad3, we have identified two sites N-terminal to the conserved kinase domain that are essential for Rad3p function. One of these sites is the putative leucine zipper, which is conserved in other phosphatidylinositol 3-kinase-related family members. The other is a novel motif, which may also mediate Rad3p protein–protein interactions.

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46

Ravi, Srimadhavi, Bhanu Priya, Pankaj Dubey, Vijay Thiruvenkatam, and Sivapriya Kirubakaran. "Molecular Docking and Molecular Dynamics Simulation Studies of Quinoline-3-Carboxamide Derivatives with DDR Kinases–Selectivity Studies towards ATM Kinase." Chemistry 3, no. 2 (April 11, 2021): 511–24. http://dx.doi.org/10.3390/chemistry3020036.

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Quinoline-3-carboxamides are an essential class of drug-like small molecules that are known to inhibit the phosphatidylinositol 3-kinase-related kinases (PIKK) family kinases. The quinoline nitrogen is shown to bind to the hinge region of the kinases, making them competitive inhibitors of adenosine triphosphate (ATP). We have previously designed and synthesized quinoline-3-carboxamides as potential ataxia telangiectasia mutated (ATM) kinase inhibitors to function as an adjuvant treatment with DNA damaging agents. This article discusses the molecular docking studies performed with these derivatives with the DNA damage and response (DDR) kinases-ATM, ataxia telangiectasia and rad3 related (ATR), and DNA dependent protein kinase catalytic subunit (DNA-PKcs) and highlights their selectivity towards ATM kinase. Docking studies were also performed with mTOR and PI3Kγ, which are close homologs of the DDR kinases. Molecular dynamics simulations were performed for one of the inhibitors against all the enzymes to establish the stability of the interactions involved. Finally, the absorption, distribution, metabolism, and excretion (ADME) properties of the inhibitors were predicted using the QikProp manual in Maestro. In conclusion, the molecules synthesized showed high selectivity towards the ATM kinase in comparison with the other kinases, though the sequence similarity between them was relatively high.

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Gold, M. R., V. W. Chan, C. W. Turck, and A. L. DeFranco. "Membrane Ig cross-linking regulates phosphatidylinositol 3-kinase in B lymphocytes." Journal of Immunology 148, no. 7 (April 1, 1992): 2012–22. http://dx.doi.org/10.4049/jimmunol.148.7.2012.

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Abstract Cross-linking of the B cell AgR results in activation of mature B cells and tolerization of immature B cells. The initial signaling events stimulated by membrane immunoglobulin (mIg) cross-linking are tyrosine phosphorylation of a number of proteins. Among the targets of mIg-induced tyrosine phosphorylation are the tyrosine kinases encoded by the lyn, blk, fyn, and syk genes, the mIg-associated proteins MB-1 and Ig-beta, phospholipase C-gamma 1 and -gamma 2, as well as many unidentified proteins. In this report we show that mIg cross-linking also regulates phosphatidylinositol 3-kinase (PtdIns 3-kinase), an enzyme that phosphorylates inositol phospholipids and plays a key role in mediating the effects of tyrosine kinases on growth control in fibroblasts. Cross-linking mIg on B lymphocytes greatly increased the amount of PtdIns 3-kinase activity which could be immunoprecipitated with anti-phosphotyrosine (anti-tyr(P) antibodies. This response was observed after mIg cross-linking in mIgM- and mIgG-bearing B cell lines and after cross-linking either mIgM or mIgD in murine splenic B cells. Thus, regulation of PtdIns 3-kinase is a common feature of signaling by several different isotypes of mIg. This response was rapid and peaked 2 to 3 min after the addition of anti-Ig antibodies. The anti-Ig-stimulated increase in PtdIns 3-kinase activity associated with anti-Tyr(P) immunoprecipitates could reflect increased tyrosine phosphorylation of PtdIns 3-kinase, increased activity of the enzyme, or both. In favor of the first possibility, the tyrosine kinase inhibitor herbimycin A blocked the increase in ant-Tyr(P)-immunoprecipitated PtdIns 3-kinase activity as well as the anti-Ig-induced tyrosine phosphorylation. Moreover, this response was not secondary to phospholipase C activation but rather seemed to be a direct consequence of mIg-induced tyrosine phosphorylation. Activation of the phosphoinositide pathway by a transfected M1 muscarinic acetylcholine receptor expressed in WEHI-231 B lymphoma cells did not increase the amount of PtdIns 3-kinase activity which could be precipitated with anti-Tyr(P) antibodies. Similarly, inhibition of the phosphoinositide pathway did not abrogate the ability of mIg cross-linking to stimulate this response. Thus, mIg-induced tyrosine phosphorylation regulates PtdIns 3-kinase, an important mediator of growth control in fibroblasts and potentially an important regulatory component in B cells as well.

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Xu, Jun, Dan Liu, Gordon Gill, and Zhou Songyang. "Regulation of cytokine-independent survival kinase (CISK) by the Phox homology domain and phosphoinositides." Journal of Cell Biology 154, no. 4 (August 20, 2001): 699–706. http://dx.doi.org/10.1083/jcb.200105089.

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PKB/Akt and serum and glucocorticoid–regulated kinase (SGK) family kinases are important downstream targets of phosphatidylinositol 3 (PI-3) kinase and have been shown to mediate a variety of cellular processes, including cell growth and survival. Although regulation of Akt can be achieved through several mechanisms, including its phosphoinositide-binding Pleckstrin homology (PH) domain, how SGK kinases are targeted and regulated remains to be elucidated. Unlike Akt, cytokine-independent survival kinase (CISK)/SGK3 contains a Phox homology (PX) domain. PX domains have been implicated in several cellular events involving membrane trafficking. However, their precise function remains unknown. We demonstrate here that the PX domain of CISK interacts with phosphatidylinositol (PtdIns)(3,5)P2, PtdIns(3,4,5)P3, and to a lesser extent PtdIns(4,5)P2. The CISK PX domain is required for targeting CISK to the endosomal compartment. Mutation in the PX domain that abolished its phospholipid binding ability not only disrupted CISK localization, but also resulted in a decrease in CISK activity in vivo. These results suggest that the PX domain regulates CISK localization and function through its direct interaction with phosphoinositides. Therefore, CISK and Akt have evolved to utilize different lipid binding domains to accomplish a similar mechanism of activation in response to PI-3 kinase signaling.

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Klippel, A., C. Reinhard, W. M. Kavanaugh, G. Apell, M. A. Escobedo, and L. T. Williams. "Membrane localization of phosphatidylinositol 3-kinase is sufficient to activate multiple signal-transducing kinase pathways." Molecular and Cellular Biology 16, no. 8 (August 1996): 4117–27. http://dx.doi.org/10.1128/mcb.16.8.4117.

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Phosphatidylinositol (PI) 3-kinase is a cytoplasmic signaling molecule recruited to the membrane by activated growth factor receptors. The p85 subunit of PI 3-kinase links the catalytic p110 subunit to activated growth factor receptors and is required for enzymatic activity of p110. In this report, we describe the effects of expressing novel forms of p110 that are targeted to the membrane by either N-terminal myristoylation or C-terminal farnesylation. The expression of membrane-localized p110 is sufficient to trigger downstream responses characteristic of growth factor action, including the stimulation of pp70 S6 kinase, Akt/Rac, and Jun N-terminal kinase (JNK). These responses can also be triggered by expression of a form of p110 (p110*) that is cytosolic but exhibits a high specific activity. Finally, targeting of pl10* to the membrane results in maximal activation of downstream responses. Our data demonstrate that either membrane-targeted forms of p110 or a form of p110 with high specific activity can act as constitutively active PI 3-kinases and induce PI 3-kinase-dependent responses in the absence of growth factor stimulation. The results also show that PI 3-kinase activation is sufficient to stimulate several kinases that appear to function in different signaling pathways.

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Buhl, Anne Mette, and John C. Cambier. "Phosphorylation of CD19 Y484 and Y515, and Linked Activation of Phosphatidylinositol 3-Kinase, Are Required for B Cell Antigen Receptor-Mediated Activation of Bruton’s Tyrosine Kinase." Journal of Immunology 162, no. 8 (April 15, 1999): 4438–46. http://dx.doi.org/10.4049/jimmunol.162.8.4438.

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Abstract Bruton’s tyrosine kinase (Btk) plays a critical role in B cell Ag receptor (BCR) signaling, as indicated by the X-linked immunodeficiency and X-linked agammaglobulinemia phenotypes of mice and men that express mutant forms of the kinase. Although Btk activity can be regulated by Src-family and Syk tyrosine kinases, and perhaps by phosphatidylinositol 3,4,5-trisphosphate, BCR-coupled signaling pathways leading to Btk activation are poorly understood. In view of previous findings that CD19 is involved in BCR-mediated phosphatidylinositol 3-kinase (PI3-K) activation, we assessed its role in Btk activation. Using a CD19 reconstituted myeloma model and CD19 gene-ablated animals we found that BCR-mediated Btk activation and phosphorylation are dependent on the expression of CD19, while BCR-mediated activation of Lyn and Syk is not. Wortmannin preincubation inhibited the BCR-mediated activation and phosphorylation of Btk. Btk activation was not rescued in the myeloma by expression of a CD19 mutant in which tyrosine residues previously shown to mediate CD19 interaction with PI3-K, Y484 and Y515, were changed to phenylalanine. Taken together, the data presented indicate that BCR aggregation-driven CD19 phosphorylation functions to promote Btk activation via recruitment and activation of PI3-K. Resultant phosphatidylinositol 3,4,5-trisphosphate probably functions to localize Btk for subsequent phosphorylation and activation by Src and Syk family kinases.

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Journal articles on the topic 'Phosphatidylinositol 3-Kinases'

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