- Email: [email protected]
Radical consequences of redox active metals on α-synuclein assemblies
A. Newman / Free Radical Biology and Medicine 128 (2018) S130–S135
320
Loss of Thioredoxin-1 in central nervous system neurons results in cerebellar ataxic behavioral deficits Rachel Laufmann*, Cassandra Aegerter, Brandon Meyerink, Jill Weimer, Peter Vitiello Sanford Research, USA
Reversible oxidation of protein thiols is an important signaling paradigm to modulate protein activity during redox perturbations. Protein thiol oxidation and reduction are regulated by the thioredoxin and glutathione enzyme superfamilies. Thioredoxin 1 (Trx1) is ubiquitous to all life and its oxidoreductase activity is associated with resistance from a myriad of oxidative insults. Although the physiological relevance of thiol switch regulation is largely unknown, Trx1 activity has been implicated in the pathogenesis of numerous diseases including neurodegeneration and peripheral neuropathy. Utilizing Cre-lox transgenic systems, we are able to drive a neuron specific loss of Trx1 in brain, spinal cord, and dorsal root ganglion (DRGs), allowing us to investigate the effects of a loss of Trx1 in the central nervous system (CNS). Since neurons are highly susceptible to redox perturbations and oxidative stress, we will determine how knockdown of Trx1 activity in neurons influences neurodevelopment, neuropathology, and behavior. Trx1fl/fl; Syn1-Creþ mice exhibit seizures and ataxic behavioral deficits compared to Trx1fl/þ ; Syn1-Creþ and Syn1Creþ controls, suggesting perturbations in neural development and/or maintenance. Ledge, gait, hind limb and force plate testing at 4, 6 and 8 weeks shows increasing seizures and ataxic decline after a loss of Trx1, with Trx1fl/fl; Syn1-Creþ mice ultimately only surviving to approximately 10 weeks of age. In addition to behavioral analysis, we will perform immunohistochemistry (IHC) to investigate changes in interneuron makers, inhibitory signaling molecules, and expression of the neuronal structural protein CRMP2 – a known interactor of Trx1. Complimentary to IHC, Nissl staining will reveal any changes in brain morphology and cell numbers. Fundamental knowledge gained from this project will advance our understanding of Trx1-dependent signaling in neurons and provide new understanding for how reversible oxidation of protein thiols regulates physiological responses to redox perturbations during neurodevelopment and oxidative neurological disorders.
https://doi.org/10.1016/j.freeradbiomed.2018.10.350
321
Cyclic ozone exposure synergizes with aging leading to memory impairment in male apoE3, not apoE4, targeted replacement mice Rui-Ming Liu 1,*, Chunsun Jiang 1, Luke Stewart 1, Hui-Chien Kuo 1, William McGilberry 1, Stepahie Wall 1, Bill Liang 2, Thomas van Groen 1, Shannon Bailey 1, Trent Tipple 1, Dean Jones 2, Lori McMahon 1 1 2
University of Alabama at Birmingham, USA Emory University, USA
The etiology of Alzheimer’s disease (AD), an aging-related neurodegenerative disease, is unknown. Recent epidemiology studies suggest that exposure to unhealthy levels of ozone (O3), a highly reactive oxidant and
S133
one of most abundant urban pollutant, may be responsible. Whether O3 acts alone or synergizes with other risk factors, such as aging and APOEe4, leading to AD remains to be determined. To test whether O3 exposure, APOEe4, and aging negatively interact leading to AD, we exposed otherwise healthy male apoE4 targeted replacement (TR) mice to a cyclic O3 exposure protocol, which mimics human exposure scenarios, and compared the results to male apoE3 TR mice, which represent the majority of the human population who carries the APOE e3 gene. The results show, surprisingly, that O3 exposure impairs memory of old male apoE3, but not apoE4, mice nor young apoE3 or apoE4 mice. Further studies show that glutathione concentration decreases with increased age and with O3 exposure in the hippocampus of male apoE3 mice, although its concentration is low in male apE4 mice since young age. Cysteine concentration, on the other hand, increases with age in apoE4 mice. The activities of several antioxidant enzymes including thioredoxin 1 are also increased in the hippocampus of old apoE4 mice, compared to old apoE3 mice. This is associated with diminished lipid and protein oxidation, astrocyte activation/inflammation, and neurogenesis in O3 exposed old apoE4 mice. Our results show, for the first time, that APOEe4, a genetic risk factor for AD, actually protects old male mice from O3-induced memory impairment, probably because old apoE4 male mice have developed compensatory mechanisms to combat O3-induced oxidative stress
https://doi.org/10.1016/j.freeradbiomed.2018.10.351
322
Radical consequences of redox active metals on α-synuclein assemblies Heather Lucas* Virginia Commonwealth University, USA
Histopathological hallmarks of Parkinson’s disease (PD) include the development of granular inclusions known as Lewy bodies that are enriched with aggregates of the protein α-synuclein (αS). Historically, αS has been considered a natively unfolded protein prone to amyloidogenic behavior. A molecular understanding of how the structurally dynamic nature of this protein contributes to PD symptoms is still lacking, but metal dyshomeostasis has long been linked to PD. Thus, a deeper understanding of biometal-influenced dynamics and metal-mediated redox chemistry associated with each of the various conformational states of αS may shed light on PD pathways associated with cerebral metal miscompartmentalization. Work in our lab is dedicated to the overarching goal of elucidating the mechanistic roles of cerebral biometals, specifically iron and copper, as either protagonistic cofactors or antagonistic toxins. My lab has recently demonstrated the biophysical consequences of metal binding that influence αS structure and aggregation, including an FeII/O2dependent self-association of αS to a PD-relevant oligomer-locked A11 þ species with a high degree of right-twisted anti-parallel b-sheet structure. In this work, we further elaborate the downstream effects of metal-αS interactions, including how aerobically aggregated FeII-bound αS, in particular, promotes protein-protein interactions. Secondary structural deconvolution and identification of proteins with homologous structures have implicated protein partners for αS with significance to PD and other neurodegenerative diseases, and we have verified these interactions through immunological analyses and chemical cross-linking. We have also established the metal-induced generation of protein-based radicals that are distinct to the metal ion identity and that contribute to the structural heterogeneity of αS assemblies. Presentation of this work will offer a unique perspective on both the beneficial and consequential role of prevalent transition metals in the brain.
https://doi.org/10.1016/j.freeradbiomed.2018.10.352
320
Loss of Thioredoxin-1 in central nervous system neurons results in cerebellar ataxic behavioral deficits Rachel Laufmann*, Cassandra Aegerter, Brandon Meyerink, Jill Weimer, Peter Vitiello Sanford Research, USA
Reversible oxidation of protein thiols is an important signaling paradigm to modulate protein activity during redox perturbations. Protein thiol oxidation and reduction are regulated by the thioredoxin and glutathione enzyme superfamilies. Thioredoxin 1 (Trx1) is ubiquitous to all life and its oxidoreductase activity is associated with resistance from a myriad of oxidative insults. Although the physiological relevance of thiol switch regulation is largely unknown, Trx1 activity has been implicated in the pathogenesis of numerous diseases including neurodegeneration and peripheral neuropathy. Utilizing Cre-lox transgenic systems, we are able to drive a neuron specific loss of Trx1 in brain, spinal cord, and dorsal root ganglion (DRGs), allowing us to investigate the effects of a loss of Trx1 in the central nervous system (CNS). Since neurons are highly susceptible to redox perturbations and oxidative stress, we will determine how knockdown of Trx1 activity in neurons influences neurodevelopment, neuropathology, and behavior. Trx1fl/fl; Syn1-Creþ mice exhibit seizures and ataxic behavioral deficits compared to Trx1fl/þ ; Syn1-Creþ and Syn1Creþ controls, suggesting perturbations in neural development and/or maintenance. Ledge, gait, hind limb and force plate testing at 4, 6 and 8 weeks shows increasing seizures and ataxic decline after a loss of Trx1, with Trx1fl/fl; Syn1-Creþ mice ultimately only surviving to approximately 10 weeks of age. In addition to behavioral analysis, we will perform immunohistochemistry (IHC) to investigate changes in interneuron makers, inhibitory signaling molecules, and expression of the neuronal structural protein CRMP2 – a known interactor of Trx1. Complimentary to IHC, Nissl staining will reveal any changes in brain morphology and cell numbers. Fundamental knowledge gained from this project will advance our understanding of Trx1-dependent signaling in neurons and provide new understanding for how reversible oxidation of protein thiols regulates physiological responses to redox perturbations during neurodevelopment and oxidative neurological disorders.
https://doi.org/10.1016/j.freeradbiomed.2018.10.350
321
Cyclic ozone exposure synergizes with aging leading to memory impairment in male apoE3, not apoE4, targeted replacement mice Rui-Ming Liu 1,*, Chunsun Jiang 1, Luke Stewart 1, Hui-Chien Kuo 1, William McGilberry 1, Stepahie Wall 1, Bill Liang 2, Thomas van Groen 1, Shannon Bailey 1, Trent Tipple 1, Dean Jones 2, Lori McMahon 1 1 2
University of Alabama at Birmingham, USA Emory University, USA
The etiology of Alzheimer’s disease (AD), an aging-related neurodegenerative disease, is unknown. Recent epidemiology studies suggest that exposure to unhealthy levels of ozone (O3), a highly reactive oxidant and
S133
one of most abundant urban pollutant, may be responsible. Whether O3 acts alone or synergizes with other risk factors, such as aging and APOEe4, leading to AD remains to be determined. To test whether O3 exposure, APOEe4, and aging negatively interact leading to AD, we exposed otherwise healthy male apoE4 targeted replacement (TR) mice to a cyclic O3 exposure protocol, which mimics human exposure scenarios, and compared the results to male apoE3 TR mice, which represent the majority of the human population who carries the APOE e3 gene. The results show, surprisingly, that O3 exposure impairs memory of old male apoE3, but not apoE4, mice nor young apoE3 or apoE4 mice. Further studies show that glutathione concentration decreases with increased age and with O3 exposure in the hippocampus of male apoE3 mice, although its concentration is low in male apE4 mice since young age. Cysteine concentration, on the other hand, increases with age in apoE4 mice. The activities of several antioxidant enzymes including thioredoxin 1 are also increased in the hippocampus of old apoE4 mice, compared to old apoE3 mice. This is associated with diminished lipid and protein oxidation, astrocyte activation/inflammation, and neurogenesis in O3 exposed old apoE4 mice. Our results show, for the first time, that APOEe4, a genetic risk factor for AD, actually protects old male mice from O3-induced memory impairment, probably because old apoE4 male mice have developed compensatory mechanisms to combat O3-induced oxidative stress
https://doi.org/10.1016/j.freeradbiomed.2018.10.351
322
Radical consequences of redox active metals on α-synuclein assemblies Heather Lucas* Virginia Commonwealth University, USA
Histopathological hallmarks of Parkinson’s disease (PD) include the development of granular inclusions known as Lewy bodies that are enriched with aggregates of the protein α-synuclein (αS). Historically, αS has been considered a natively unfolded protein prone to amyloidogenic behavior. A molecular understanding of how the structurally dynamic nature of this protein contributes to PD symptoms is still lacking, but metal dyshomeostasis has long been linked to PD. Thus, a deeper understanding of biometal-influenced dynamics and metal-mediated redox chemistry associated with each of the various conformational states of αS may shed light on PD pathways associated with cerebral metal miscompartmentalization. Work in our lab is dedicated to the overarching goal of elucidating the mechanistic roles of cerebral biometals, specifically iron and copper, as either protagonistic cofactors or antagonistic toxins. My lab has recently demonstrated the biophysical consequences of metal binding that influence αS structure and aggregation, including an FeII/O2dependent self-association of αS to a PD-relevant oligomer-locked A11 þ species with a high degree of right-twisted anti-parallel b-sheet structure. In this work, we further elaborate the downstream effects of metal-αS interactions, including how aerobically aggregated FeII-bound αS, in particular, promotes protein-protein interactions. Secondary structural deconvolution and identification of proteins with homologous structures have implicated protein partners for αS with significance to PD and other neurodegenerative diseases, and we have verified these interactions through immunological analyses and chemical cross-linking. We have also established the metal-induced generation of protein-based radicals that are distinct to the metal ion identity and that contribute to the structural heterogeneity of αS assemblies. Presentation of this work will offer a unique perspective on both the beneficial and consequential role of prevalent transition metals in the brain.
https://doi.org/10.1016/j.freeradbiomed.2018.10.352