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1.
J Neurochem ; 157(6): 2055-2069, 2021 06.
Article in English | MEDLINE | ID: mdl-33220080

ABSTRACT

Rarefaction of the dendritic tree leading to neuronal dysfunction is a hallmark of many neurodegenerative diseases and we have shown previously that heat shock protein B5 (HspB5)/αB-crystallin is able to increase dendritic complexity in vitro. The aim of this study was to investigate if this effect is also present in vivo, if HspB5 can counteract dendritic rarefaction under pathophysiological conditions and the impact of phosphorylation of HspB5 in this process. HspB5 and eight mutants inhibiting or mimicking phosphorylation at the three phosphorylation sites serine (S)19, S45, and S59 were over-expressed in cultured rat hippocampal neurons with subsequent investigation of the complexity of the dendritic tree. Sholl analysis revealed significant higher complexity of the dendritic tree after over-expression of wild-type HspB5 and the mutant HspB5-AEE. All other mutants showed no or minor effects. For in vivo investigation in utero electroporation of mouse embryos was applied. At embryonal day E15.5 the respective plasmids were injected, cornu ammonis 1 (CA1) pyramidal cells transfected by electroporation and their basal dendritic trees were analyzed at post-natal day P15. In vivo, HspB5 and HspB5-AEE led to an increase of total dendritic length as well as a higher complexity. Finally, the dendritic effect of HspB5 was investigated under a pathophysiological condition, that is, iron deficiency which reportedly results in dendritic rarefaction. HspB5 and HspB5-AEE but not the non-phosphorylatable mutant HspB5-AAA significantly counteracted the dendritic rarefaction. Thus, our data suggest that up-regulation and selective phosphorylation of HspB5 in neurodegenerative diseases may preserve dendritic morphology and counteract neuronal dysfunction.


Subject(s)
Crystallins/metabolism , Dendrites/metabolism , Hippocampus/metabolism , Microtubule-Associated Proteins/metabolism , Neurons/metabolism , Animals , Cells, Cultured , Dendrites/pathology , Female , Hippocampus/cytology , Hippocampus/pathology , Male , Mice , Mice, Inbred C57BL , Neurons/pathology , Phosphorylation/physiology , Pregnancy , Rats , Rats, Sprague-Dawley
2.
Cell Mol Life Sci ; 73(19): 3761-75, 2016 10.
Article in English | MEDLINE | ID: mdl-27085702

ABSTRACT

The small heat shock protein ΗspΒ5 (αB-crystallin) exhibits generally cytoprotective functions and possesses powerful neuroprotective capacity in the brain. However, little is known about the mode of action of ΗspΒ5 or other members of the HspB family particularly in neurons. To get clues of the neuronal function of HspBs, we overexpressed several HspBs in cultured rat hippocampal neurons and investigated their effect on neuronal morphology and stress resistance. Whereas axon length and synapse density were not affected by any HspB, dendritic complexity was enhanced by HspB5 and, to a lesser extent, by HspB6. Furthermore, we could show that this process was dependent on phosphorylation, since a non-phosphorylatable mutant of HspB5 did not show this effect. Rarefaction of the dendritic arbor is one hallmark of several neurodegenerative diseases. To investigate if HspB5, which is upregulated at pathophysiological conditions, might be able to protect dendrites during such situations, we exposed HspB5 overexpressing neuronal cultures to heat shock. HspB5 prevented heat shock-induced rarefaction of dendrites. In conclusion, we identified regulation of dendritic complexity as a new function of HspB5 in hippocampal neurons.


Subject(s)
Crystallins/metabolism , Dendrites/metabolism , Heat-Shock Response , Hippocampus/cytology , Microtubule-Associated Proteins/metabolism , Neuroprotection , Animals , Cells, Cultured , Lentivirus/metabolism , Mutagenesis, Site-Directed , Phosphorylation , Rats , Synapses/metabolism , Transduction, Genetic
3.
Pest Manag Sci ; 72(11): 2166-2175, 2016 Nov.
Article in English | MEDLINE | ID: mdl-26842010

ABSTRACT

BACKGROUND: The increasing structural diversity of the neonicotinoid class of insecticides presently used in crop protection calls for a more detailed analysis of their mode of action at their cellular targets, the nicotinic acetylcholine receptors. RESULTS: Comparative radioligand binding studies using membranes of Myzus persicae (Sulzer) and representatives of the chloropyridyl subclass (imidacloprid), the chlorothiazolyl subclass (thiamethoxam), the tetrahydrofuranyl subclass (dinotefuran), as well as the novel sulfoximine type (sulfoxaflor), which is not a neonicotinoid, reveal significant differences in the number of binding sites, the displacing potencies and the mode of binding interference. Furthermore, the mode of interaction of [3 H]thiamethoxam and the nicotinic antagonists methyllycaconitine and dihydro-ß-erythroidine is unique, with Hill values of >1, clearly different to the values of around unity for [3 H]imidacloprid and [3 H]N-desmethylthiamethoxam. The interaction of [3 H]N-desmethylthiamethoxam with the agonist (-)nicotine is also characterised by a Hill value of >1. CONCLUSIONS: There is no single conserved site or mode of binding of neonicotinoids and related nicotinic ligands to their target receptor, but a variety of binding pockets depending on the combination of receptor subunits, the receptor subtype, its functional state, as well as the structural flexibility of both the binding pockets and the ligands. © 2016 Society of Chemical Industry.


Subject(s)
Aphids/metabolism , Insect Proteins/genetics , Insecticides/pharmacology , Receptors, Nicotinic/genetics , Animals , Guanidines/pharmacology , Imidazoles/pharmacology , Insect Proteins/metabolism , Neonicotinoids , Nicotinic Agonists/metabolism , Nicotinic Antagonists/metabolism , Nitro Compounds/pharmacology , Oxazines/pharmacology , Receptors, Nicotinic/metabolism , Thiamethoxam , Thiazoles/pharmacology
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