Na+/Ca2+ exchanger 1 on nuclear envelope controls PTEN/Akt pathway via nucleoplasmic Ca2+ regulation during neuronal differentiation

Agnese Secondo, Alba Esposito, Tiziana Petrozziello, Francesca Boscia, Pasquale Molinaro, Valentina Tedeschi, Anna Pannaccione, Roselia Ciccone, Natascia Guida, Gianfranco Di Renzo, Lucio Annunziato

Research output: Contribution to journalArticle

Abstract

Nuclear envelope (NE) is a Ca2+-storing organelle controlling neuronal differentiation through nuclear Ca2+ concentrations ([Ca2+]n). However, how [Ca2+]n regulates this important function remains unknown. Here, we investigated the role of the nuclear form of the Na+/Ca2+ exchanger 1(nuNCX1) during the different stages of neuronal differentiation and the involvement of PTEN/PI3'K/Akt pathway. In neuronal cells, nuNCX1 was detected on the inner membrane of the NE where protein expression and activity of the exchanger increased during NGF-induced differentiation. nuNCX1 activation by Na+-free perfusion induced a time-dependent activation of nuclear-resident PI3K/Akt pathway in isolated nuclei. To discriminate the contribution of nuNCX1 from those of plasma membrane NCX, we generated a chimeric protein composed of the fluorophore EYFP, the exchanger inhibitory peptide, and the nuclear localization signal, named XIP-NLS. Fura-2 measurements on single nuclei and patch-clamp experiments in whole-cell configuration showed that XIP-NLS selectively inhibited nuNCX1. Once it reached the nuclear compartment, XIP-NLS increased the nucleoplasmic Ca2+ peak elicited by ATP and reduced Akt phosphorylation, GAP-43 and MAP-2 expression through nuclear-resident PTEN induction. Furthermore, in accordance with the prevention of the neuronal phenotype, XIP-NLS significantly reduced TTX-sensitive Na+ currents and membrane potential during neuronal differentiation. The selective inhibition of nuNCX1 by XIP-NLS increased the percentage of β III tubulin-positive immature neurons in mature cultures of MAP-2-positive cortical neurons, thus unraveling a new function for nuNCX1 in regulating neuronal differentiation through [Ca2+]n-dependent PTEN/PI3K/Akt pathway.

Original languageEnglish
Pages (from-to)12
JournalCell Death Discovery
Volume4
DOIs
Publication statusPublished - Dec 2018

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Nuclear Envelope
Phosphatidylinositol 3-Kinases
GAP-43 Protein
Neurons
Nuclear Localization Signals
Fura-2
Nerve Growth Factor
Tubulin
Nuclear Proteins
Organelles
Membrane Potentials
Perfusion
Adenosine Triphosphate
Phosphorylation
Cell Membrane
Phenotype
Membranes
Proteins
exchanger inhibitory peptide

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Na+/Ca2+ exchanger 1 on nuclear envelope controls PTEN/Akt pathway via nucleoplasmic Ca2+ regulation during neuronal differentiation. / Secondo, Agnese; Esposito, Alba; Petrozziello, Tiziana; Boscia, Francesca; Molinaro, Pasquale; Tedeschi, Valentina; Pannaccione, Anna; Ciccone, Roselia; Guida, Natascia; Di Renzo, Gianfranco; Annunziato, Lucio.

In: Cell Death Discovery, Vol. 4, 12.2018, p. 12.

Research output: Contribution to journalArticle

Secondo, A, Esposito, A, Petrozziello, T, Boscia, F, Molinaro, P, Tedeschi, V, Pannaccione, A, Ciccone, R, Guida, N, Di Renzo, G & Annunziato, L 2018, 'Na+/Ca2+ exchanger 1 on nuclear envelope controls PTEN/Akt pathway via nucleoplasmic Ca2+ regulation during neuronal differentiation', Cell Death Discovery, vol. 4, pp. 12. https://doi.org/10.1038/s41420-017-0018-1
Secondo, Agnese ; Esposito, Alba ; Petrozziello, Tiziana ; Boscia, Francesca ; Molinaro, Pasquale ; Tedeschi, Valentina ; Pannaccione, Anna ; Ciccone, Roselia ; Guida, Natascia ; Di Renzo, Gianfranco ; Annunziato, Lucio. / Na+/Ca2+ exchanger 1 on nuclear envelope controls PTEN/Akt pathway via nucleoplasmic Ca2+ regulation during neuronal differentiation. In: Cell Death Discovery. 2018 ; Vol. 4. pp. 12.
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AU - Secondo, Agnese

AU - Esposito, Alba

AU - Petrozziello, Tiziana

AU - Boscia, Francesca

AU - Molinaro, Pasquale

AU - Tedeschi, Valentina

AU - Pannaccione, Anna

AU - Ciccone, Roselia

AU - Guida, Natascia

AU - Di Renzo, Gianfranco

AU - Annunziato, Lucio

PY - 2018/12

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N2 - Nuclear envelope (NE) is a Ca2+-storing organelle controlling neuronal differentiation through nuclear Ca2+ concentrations ([Ca2+]n). However, how [Ca2+]n regulates this important function remains unknown. Here, we investigated the role of the nuclear form of the Na+/Ca2+ exchanger 1(nuNCX1) during the different stages of neuronal differentiation and the involvement of PTEN/PI3'K/Akt pathway. In neuronal cells, nuNCX1 was detected on the inner membrane of the NE where protein expression and activity of the exchanger increased during NGF-induced differentiation. nuNCX1 activation by Na+-free perfusion induced a time-dependent activation of nuclear-resident PI3K/Akt pathway in isolated nuclei. To discriminate the contribution of nuNCX1 from those of plasma membrane NCX, we generated a chimeric protein composed of the fluorophore EYFP, the exchanger inhibitory peptide, and the nuclear localization signal, named XIP-NLS. Fura-2 measurements on single nuclei and patch-clamp experiments in whole-cell configuration showed that XIP-NLS selectively inhibited nuNCX1. Once it reached the nuclear compartment, XIP-NLS increased the nucleoplasmic Ca2+ peak elicited by ATP and reduced Akt phosphorylation, GAP-43 and MAP-2 expression through nuclear-resident PTEN induction. Furthermore, in accordance with the prevention of the neuronal phenotype, XIP-NLS significantly reduced TTX-sensitive Na+ currents and membrane potential during neuronal differentiation. The selective inhibition of nuNCX1 by XIP-NLS increased the percentage of β III tubulin-positive immature neurons in mature cultures of MAP-2-positive cortical neurons, thus unraveling a new function for nuNCX1 in regulating neuronal differentiation through [Ca2+]n-dependent PTEN/PI3K/Akt pathway.

AB - Nuclear envelope (NE) is a Ca2+-storing organelle controlling neuronal differentiation through nuclear Ca2+ concentrations ([Ca2+]n). However, how [Ca2+]n regulates this important function remains unknown. Here, we investigated the role of the nuclear form of the Na+/Ca2+ exchanger 1(nuNCX1) during the different stages of neuronal differentiation and the involvement of PTEN/PI3'K/Akt pathway. In neuronal cells, nuNCX1 was detected on the inner membrane of the NE where protein expression and activity of the exchanger increased during NGF-induced differentiation. nuNCX1 activation by Na+-free perfusion induced a time-dependent activation of nuclear-resident PI3K/Akt pathway in isolated nuclei. To discriminate the contribution of nuNCX1 from those of plasma membrane NCX, we generated a chimeric protein composed of the fluorophore EYFP, the exchanger inhibitory peptide, and the nuclear localization signal, named XIP-NLS. Fura-2 measurements on single nuclei and patch-clamp experiments in whole-cell configuration showed that XIP-NLS selectively inhibited nuNCX1. Once it reached the nuclear compartment, XIP-NLS increased the nucleoplasmic Ca2+ peak elicited by ATP and reduced Akt phosphorylation, GAP-43 and MAP-2 expression through nuclear-resident PTEN induction. Furthermore, in accordance with the prevention of the neuronal phenotype, XIP-NLS significantly reduced TTX-sensitive Na+ currents and membrane potential during neuronal differentiation. The selective inhibition of nuNCX1 by XIP-NLS increased the percentage of β III tubulin-positive immature neurons in mature cultures of MAP-2-positive cortical neurons, thus unraveling a new function for nuNCX1 in regulating neuronal differentiation through [Ca2+]n-dependent PTEN/PI3K/Akt pathway.

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