Nucleofection induces transient eIF2α phosphorylation by GCN2 and PERK
Identifieur interne : 004A51 ( Ncbi/Curation ); précédent : 004A50; suivant : 004A52Nucleofection induces transient eIF2α phosphorylation by GCN2 and PERK
Auteurs : Bart R. Anderson [États-Unis] ; Katalin Karik [États-Unis] ; Drew Weissman [États-Unis]Source :
- Gene therapy [ 0969-7128 ] ; 2012.
Abstract
Nucleofection permits efficient transfection even with difficult cell types such as primary and non-dividing cells, and is used to deliver various nucleic acids including DNA, mRNA, and siRNA. Unlike DNA and siRNA, mRNA is subject to rapid degradation, which necessitates instant early translation following mRNA delivery. We examined factors important in translation following nucleofection and observed rapid phosphorylation of eukaryotic initiation factor 2 alpha (eIF2α) following nucleofection, which occurred in the absence of delivered nucleic acid. We studied the involvement of 3 ubiquitous kinases capable of phosphorylating eIF2α in mammalian cells and identified that nucleofection-mediated phosphorylation of eIF2α was dependent on general control non-derepressible 2 (GCN2) and protein kinase RNA-activated (PKR)-like ER kinase (PERK) but not PKR. A reduction in translation due to eIF2α phosphorylation was observed post nucleofection demonstrating functional significance. Understanding the impact of nucleofection on translational machinery has important implications for therapeutics currently under development based on the delivery of mRNA, DNA, and siRNA. Strategies to circumvent eIF2α phosphorylation and other downstream effects of activating GCN2 and PERK will facilitate further advancement of nucleic acid-based therapies.
Url:
DOI: 10.1038/gt.2012.5
PubMed: 22301437
PubMed Central: 3345295
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Nucleofection induces transient eIF2α phosphorylation by GCN2 and PERK</title><author><name sortKey="Anderson, Bart R" sort="Anderson, Bart R" uniqKey="Anderson B" first="Bart R." last="Anderson">Bart R. Anderson</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Medicine, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Medicine, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author><author><name sortKey="Karik, Katalin" sort="Karik, Katalin" uniqKey="Karik K" first="Katalin" last="Karik">Katalin Karik</name><affiliation wicri:level="1"><nlm:aff id="A2">Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author><author><name sortKey="Weissman, Drew" sort="Weissman, Drew" uniqKey="Weissman D" first="Drew" last="Weissman">Drew Weissman</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Medicine, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Medicine, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">22301437</idno><idno type="pmc">3345295</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3345295</idno><idno type="RBID">PMC:3345295</idno><idno type="doi">10.1038/gt.2012.5</idno><date when="2012">2012</date><idno type="wicri:Area/Pmc/Corpus">004711</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">004711</idno><idno type="wicri:Area/Pmc/Curation">004710</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Curation">004710</idno><idno type="wicri:Area/Pmc/Checkpoint">002854</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Checkpoint">002854</idno><idno type="wicri:Area/Ncbi/Merge">004A51</idno><idno type="wicri:Area/Ncbi/Curation">004A51</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Nucleofection induces transient eIF2α phosphorylation by GCN2 and PERK</title><author><name sortKey="Anderson, Bart R" sort="Anderson, Bart R" uniqKey="Anderson B" first="Bart R." last="Anderson">Bart R. Anderson</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Medicine, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Medicine, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author><author><name sortKey="Karik, Katalin" sort="Karik, Katalin" uniqKey="Karik K" first="Katalin" last="Karik">Katalin Karik</name><affiliation wicri:level="1"><nlm:aff id="A2">Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author><author><name sortKey="Weissman, Drew" sort="Weissman, Drew" uniqKey="Weissman D" first="Drew" last="Weissman">Drew Weissman</name><affiliation wicri:level="1"><nlm:aff id="A1">Department of Medicine, University of Pennsylvania, Philadelphia, PA, USA</nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Medicine, University of Pennsylvania, Philadelphia, PA</wicri:regionArea><wicri:noRegion>PA</wicri:noRegion></affiliation></author></analytic><series><title level="j">Gene therapy</title><idno type="ISSN">0969-7128</idno><idno type="eISSN">1476-5462</idno><imprint><date when="2012">2012</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p id="P1">Nucleofection permits efficient transfection even with difficult cell types such as primary and non-dividing cells, and is used to deliver various nucleic acids including DNA, mRNA, and siRNA. Unlike DNA and siRNA, mRNA is subject to rapid degradation, which necessitates instant early translation following mRNA delivery. We examined factors important in translation following nucleofection and observed rapid phosphorylation of eukaryotic initiation factor 2 alpha (eIF2α) following nucleofection, which occurred in the absence of delivered nucleic acid. We studied the involvement of 3 ubiquitous kinases capable of phosphorylating eIF2α in mammalian cells and identified that nucleofection-mediated phosphorylation of eIF2α was dependent on general control non-derepressible 2 (GCN2) and protein kinase RNA-activated (PKR)-like ER kinase (PERK) but not PKR. A reduction in translation due to eIF2α phosphorylation was observed post nucleofection demonstrating functional significance. Understanding the impact of nucleofection on translational machinery has important implications for therapeutics currently under development based on the delivery of mRNA, DNA, and siRNA. 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Hp" uniqKey="Harding H">HP Harding</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kawagishi Kobayashi, M" uniqKey="Kawagishi Kobayashi M">M Kawagishi-Kobayashi</name></author><author><name sortKey="Silverman, Jb" uniqKey="Silverman J">JB Silverman</name></author><author><name sortKey="Ung, Tl" uniqKey="Ung T">TL Ung</name></author><author><name sortKey="Dever, Te" uniqKey="Dever T">TE Dever</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kariko, K" uniqKey="Kariko K">K Kariko</name></author><author><name sortKey="Buckstein, M" uniqKey="Buckstein M">M Buckstein</name></author><author><name sortKey="Ni, H" uniqKey="Ni H">H Ni</name></author><author><name sortKey="Weissman, D" uniqKey="Weissman D">D Weissman</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Kariko, K" uniqKey="Kariko K">K Kariko</name></author><author><name sortKey="Muramatsu, H" uniqKey="Muramatsu H">H Muramatsu</name></author><author><name sortKey="Welsh, Fa" uniqKey="Welsh F">FA Welsh</name></author><author><name sortKey="Ludwig, J" uniqKey="Ludwig J">J Ludwig</name></author><author><name sortKey="Kato, H" uniqKey="Kato H">H Kato</name></author><author><name sortKey="Akira, S" uniqKey="Akira S">S Akira</name></author></analytic></biblStruct></listBibl></div1></back></TEI></record>Pour manipuler ce document sous Unix (Dilib)
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