title: Pathways for plasmalemmal repair mediated by PKA, Epac, and cytosolic oxidation in rat B104 cells in vitro and rat sciatic axons ex vivo.
abstract: Plasmalemmal repair (sealing) is necessary for survival of damaged eukaryotic cells. Ca(2+) influx through plasmalemmal disruptions activates pathways that initiate sealing, which is commonly assessed by exclusion of extracellular dye. These sealing pathways include PKA, Epac, and cytosolic oxidation. In this article, we investigate whether PKA, Epac, and/or cytosolic oxidation, activate specific proteins required to produce a plasmalemmal seal. We report that toxin cleavage of proteins required for neurotransmitter release (SNAP-25), inhibition of Golgi trafficking (with Brefeldin A: Bref A) or inhibition of N-ethylmaleimide sensitive factor (NSF) all decrease sealing of rat B104 hippocampal cells with transected neuritis in vitro. Epac, but not PKA or cytosolic oxidation, partly overcomes the decrease in sealing produced by cleavage of SNAP-25. PKA and increased cytosolic oxidation, but not Epac, can partly overcome the decrease in sealing due to Bref A. PKA, Epac, and/or cytosolic oxidation cannot overcome NSF inhibition. Substances that affect plasmalemmal sealing of B104 neurites in vitro have similar effects on plasmalemmal sealing in rat sciatic axons ex vivo. From these and other data, we propose a model of plasmalemmal sealing having three redundant, evolutionarily conserved, parallel pathways that all converge on NSF.
authors.affiliation: Institute for Cellular and Molecular Biology, University of Texas at Austin, Austin, Texas 78712, USA.
authors.forename: Christopher S
authors.initials: CS
authors.lastname: Spaeth
authors.forename: Elaine B
authors.initials: EB
authors.lastname: Spaeth
authors.forename: Robert W
authors.initials: RW
authors.lastname: Wilcott
authors.forename: Jerry D
authors.initials: JD
authors.lastname: Fan
authors.forename: Taylor
authors.initials: T
authors.lastname: Robison
authors.forename: George D
authors.initials: GD
authors.lastname: Bittner
mesh_terms: D000818:Animals; D001369:Axons / Q000502:physiology*; D019771:Axotomy; D002118:Calcium / Q000502:physiology; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000502:physiology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000502:physiology; D003600:Cytosol / Q000502:physiology; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology; D006624:Hippocampus / Q000502:physiology; D066298:In Vitro Techniques; D052066:N-Ethylmaleimide-Sensitive Proteins / Q000502:physiology; D009416:Nerve Regeneration / Q000502:physiology*; D016501:Neurites / Q000502:physiology; D010084:Oxidation-Reduction; D051381:Rats; D012584:Sciatic Nerve / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*; D050825:Synaptosomal-Associated Protein 25 / Q000502:physiology
chemical_list: D020662:Guanine Nucleotide Exchange Factors; C526973:Rapgef3 protein, rat; C531571:Rapgef4 protein, rat; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D017868:Cyclic AMP-Dependent Protein Kinases; D052066:N-Ethylmaleimide-Sensitive Proteins; C498737:Nsf protein, rat; D002118:Calcium