title: Class I PI3-kinase or Akt inhibition do not impair axonal polarization, but slow down axonal elongation.
abstract: PI3K proteins family have multiple and essential functions in most cellular events. This family is composed of class I, class II and class III PI3Ks, which upstream and downstream elements are not completely elucidated. Previous studies using the broad PI3K inhibitor, LY294002 allowed to propose that PI3 kinase>Akt pathway is a key element in the determination of axonal polarity in hippocampal neurons. Recently, new inhibitors with a higher selectivity for class I PI3K have been characterized. In the present study we have examined this widely accepted theory using a new class I PI3K inhibitor (GDC-0941), as well as Akt inhibitors, and PTEN phosphatase constructs to reduce PIP3 levels. Our present data show that both, class I PI3K inhibitor and Akt inhibitor did not alter axon specification in hippocampal neurons, but greatly reduced axon length. However, in the same experiments LY294002 effectively impeded axonal polarization, as previously reported. Our biochemical data show that both, class I PI3K and Akt inhibitors, effectively block downstream elements from Akt to S6K1 activity. Both inhibitors are stable in culture medium along the time period analysed, maintaining the inhibition better than LY294002. Besides, we found evidence that LY294002 directly inhibits mTORC1. However, further analysis using an mTORC1 inhibitor showed no change in neuron polarity. Same result was obtained using a general class III PI3K inhibitor. Interestingly, we found that either, wild-type PTEN, or a phosphatase-dead form of PTEN, disrupted axonal polarization, strongly suggesting that the role of PTEN in axonal polarity can be independent of PIP3.
authors.affiliation: Centro de Biología Molecular "Severo Ochoa", CSIC-UAM, Univ. Autonoma de Madrid, 28049 Madrid, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain.
authors.forename: Héctor
authors.initials: H
authors.lastname: Diez
authors.affiliation: Centro de Biología Molecular "Severo Ochoa", CSIC-UAM, Univ. Autonoma de Madrid, 28049 Madrid, Spain; Dept. Quimica Fisica Aplicada, Univ. Autonoma de Madrid, Madrid 28049, Spain; Department of Molecular, Cellular and Developmental Neurobiology, Instituto Cajal, CSIC, Spain.
authors.forename: Ma José
authors.initials: MJ
authors.lastname: Benitez
authors.affiliation: Department of Functional and Systems Neurobiology, Instituto Cajal, CSIC, Madrid 28002, Spain.
authors.forename: Silvia
authors.initials: S
authors.lastname: Fernandez
authors.affiliation: Department of Functional and Systems Neurobiology, Instituto Cajal, CSIC, Madrid 28002, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Torres-Aleman
authors.affiliation: Department of Molecular, Cellular and Developmental Neurobiology, Instituto Cajal, CSIC, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain. Electronic address: jjgarrido@cajal.csic.es.
authors.forename: Juan José
authors.initials: JJ
authors.lastname: Garrido
authors.affiliation: Centro de Biología Molecular "Severo Ochoa", CSIC-UAM, Univ. Autonoma de Madrid, 28049 Madrid, Spain; Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Spain. Electronic address: fwandosell@cbm.csic.es.
authors.forename: Francisco
authors.initials: F
authors.lastname: Wandosell
mesh_terms: D000818:Animals; D001369:Axons / Q000187:drug effects* / Q000201:enzymology; D045744:Cell Line, Tumor; D016764:Cell Polarity / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D002867:Chromones / Q000494:pharmacology*; D058534:Class I Phosphatidylinositol 3-Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005865:Gestational Age; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D007191:Indazoles / Q000494:pharmacology*; D000076222:Mechanistic Target of Rapamycin Complex 1; D051379:Mice; D009025:Morpholines / Q000494:pharmacology*; D046912:Multiprotein Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D051059:PTEN Phosphohydrolase / Q000235:genetics / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013449:Sulfonamides / Q000494:pharmacology*; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D014162:Transfection
chemical_list: C532162:2-(1H-indazol-4-yl)-6-(4-methanesulfonylpiperazin-1-ylmethyl)-4-morpholin-4-ylthieno(3,2-d)pyrimidine; D002867:Chromones; D007191:Indazoles; C549075:LY 290042; D009025:Morpholines; D046912:Multiprotein Complexes; D018129:Phosphatidylinositol Phosphates; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D058534:Class I Phosphatidylinositol 3-Kinases; D000076222:Mechanistic Target of Rapamycin Complex 1; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D051059:PTEN Phosphohydrolase
keywords: Akt; Axon establishment; PI3K; Polarity; Signalling; mTORC1