DSpace Repository :: Browsing by Author "Caldwell, Kim A."

Browsing by Author "Caldwell, Kim A."

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Alterations in Double-Stranded Rna-Mediated Gene Silencing Modulate Α-Synuclein-Associated Neurodegeneration in C. Elegans Parkinson's Disease Models
(University of Alabama Libraries, 2022) Gaeta, Anthony Louis; Caldwell, Guy A.; Caldwell, Kim A.; University of Alabama Tuscaloosa
Parkinson's disease (PD), the second most common neurodegenerative disease in the world, is characterized by the progressive loss of dopamine (DA) neurons over time, which leads to worsening dysfunctions in movement and cognition. Although much research and effort has been spent trying to develop treatments and therapies for PD, accomplishments in this regard have been few and far between. Identifying targetable and modifiable factors that can lead to amelioration of symptoms, inhibition of progression, and ultimately a cure, is undoubtedly the end goal of research relating to PD. Model organisms have been used as a tool for discovery of such factors and have resulted in great strides in understanding of the molecular mechanisms that contribute to the disease.This work utilizes the nematode roundworm, Caenorhabditis elegans, to model PD by overexpressing the pathological hallmark, α-synuclein (α-syn), in the DA neurons of the animals. This results in progressive, age-dependent DA neurodegeneration, just as is seen in humans with PD. These α-syn expressing worms were used to enrich for populations resilient to the stress of α-syn, which resulted in the identification of genes that function to edit RNA and regulate G-proteins as factors of neuronal resilience. Additionally, the outcomes of this research have positioned Systemic RNA Interference Defective (SID) genes as modulators of α-syn-mediated neurodegeneration. A pharmacological compound which inhibits the activity of a SID protein, SID-3, was shown to reduce enhancement of α-syn-mediated neurodegeneration stemming from overexpression of a microRNA, mir-2, potentially positioning it and its targets as therapeutic candidates. Lastly, a non-standard bacterial diet for C. elegans was found to be neuroprotective in a transgenerational manner, and hundreds of novel factors were identified that contribute to this diet-induced benefit. Strikingly, the neuroprotection seen by the factors identified in this study were dependent on one common process: dsRNA-mediated gene silencing, and their ability to be imported into cells. With the help of C. elegans, the outcomes of this research have provided novel insights into factors that regulate neurodegeneration related to PD.
Amelioration of Alzheimer's disease pathology by mitophagy inducers identified via machine learning and a cross-species workflow
(Nature Portfolio, 2022) Xie, Chenglong; Zhuang, Xu-Xu; Niu, Zhangming; Ai, Ruixue; Lautrup, Sofie; Zheng, Shuangjia; Jiang, Yinghui; Han, Ruiyu; Sen Gupta, Tanima; Cao, Shuqin; Lagartos-Donate, Maria Jose; Cai, Cui-Zan; Xie, Li-Ming; Caponio, Domenica; Wang, Wen-Wen; Schmauck-Medina, Tomas; Zhang, Jianying; Wang, He-ling; Lou, Guofeng; Xiao, Xianglu; Zheng, Wenhua; Palikaras, Konstantinos; Yang, Guang; Caldwell, Kim A.; Caldwell, Guy A.; Shen, Han-Ming; Nilsen, Hilde; Lu, Jia-Hong; Fang, Evandro F.; Wenzhou Medical University; University of Oslo; Oujiang Laboratory; University of Macau; Sun Yat Sen University; Athens Medical School; National & Kapodistrian University of Athens; Royal Brompton Hospital; Imperial College London; University of Alabama Tuscaloosa; University of Alabama Birmingham; National University of Singapore; Zhengzhou University
A reduced removal of dysfunctional mitochondria is common to aging and age-related neurodegenerative pathologies such as Alzheimer's disease (AD). Strategies for treating such impaired mitophagy would benefit from the identification of mitophagy modulators. Here we report the combined use of unsupervised machine learning (involving vector representations of molecular structures, pharmacophore fingerprinting and conformer fingerprinting) and a cross-species approach for the screening and experimental validation of new mitophagy-inducing compounds. From a library of naturally occurring compounds, the workflow allowed us to identify 18 small molecules, and among them two potent mitophagy inducers (Kaempferol and Rhapontigenin). In nematode and rodent models of AD, we show that both mitophagy inducers increased the survival and functionality of glutamatergic and cholinergic neurons, abrogated amyloid-beta and tau pathologies, and improved the animals' memory. Our findings suggest the existence of a conserved mechanism of memory loss across the AD models, this mechanism being mediated by defective mitophagy. The computational-experimental screening and validation workflow might help uncover potent mitophagy modulators that stimulate neuronal health and brain homeostasis.
ApoE-associated modulation of neuroprotection from A beta-mediated neurodegeneration in transgenic Caenorhabditis elegans
(Company of Biologists, 2019) Griffin, Edward F.; Scopel, Samuel E.; Stephen, Cayman A.; Holzhauer, Adam C.; Vaji, Madeline A.; Tuckey, Ryan A.; Berkowitz, Laura A.; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
Allele-specific distinctions in the human apolipoprotein E (APOE) locus represent the best-characterized genetic predictor of Alzheimer's disease (AD) risk. Expression of isoform APOE epsilon 2 is associated with reduced risk, while APOE epsilon 3 is neutral and APOE epsilon 4 carriers exhibit increased susceptibility. Using Caenorhabditis elegans, we generated a novel suite of humanized transgenic nematodes to facilitate neuronal modeling of amyloid-beta peptide (A beta) co-expression in the context of distinct human APOE alleles. We found that co-expression of human APOE epsilon 2 with A beta attenuated A beta-induced neurodegeneration, whereas expression of the APOE epsilon 4 allele had no effect on neurodegeneration, indicating a loss of neuroprotective capacity. Notably, the APOE epsilon 3 allele displayed an intermediate phenotype; it was not neuroprotective in young adults but attenuated neurodegeneration in older animals. There was no functional impact from the three APOE isoforms in the absence of A beta co-expression. Pharmacological treatment that examined neuroprotective effects of APOE alleles on calcium homeostasis showed allele-specific responses to changes in ER-associated calcium dynamics in the A beta background. Additionally, A beta suppressed survival, an effect that was rescued by APOE epsilon 2 and APOE epsilon 3, but not APOE epsilon 4. Expression of the APOE alleles in neurons, independent of A beta, exerted no impact on survival. Taken together, these results illustrate that C. elegans provides a powerful in vivo platform with which to explore how AD-associated neuronal pathways are modulated by distinct APOE gene products in the context of A beta-associated neurotoxicity. The significance of both ApoE and A beta to AD highlights the utility of this new pre-clinical model as a means to dissect their functional inter-relationship.
Attenuation of Dopaminergic Neurodegeneration in a C. elegans Parkinson's Model through Regulation of Xanthine Dehydrogenase (XDH-1) Expression by the RNA Editase, ADR-2
(MDPI, 2023) Starr, Lindsey A.; McKay, Luke E.; Peter, Kylie N.; Seyfarth, Lena M.; Berkowitz, Laura A.; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
Differential RNA editing by adenosine deaminases that act on RNA (ADARs) has been implicated in several neurological disorders, including Parkinson's disease (PD). Here, we report results of a RNAi screen of genes differentially regulated in adr-2 mutants, normally encoding the only catalytically active ADAR in Caenorhabditis elegans, ADR-2. Subsequent analysis of candidate genes that alter the misfolding of human alpha-synuclein (alpha-syn) and dopaminergic neurodegeneration, two PD pathologies, reveal that reduced expression of xdh-1, the ortholog of human xanthine dehydrogenase (XDH), is protective against alpha-synuclein-induced dopaminergic neurodegeneration. Further, RNAi experiments show that WHT-2, the worm ortholog of the human ABCG2 transporter and a predicted interactor of XDH-1, is the rate-limiting factor in the ADR-2, XDH-1, WHT-2 system for dopaminergic neuroprotection. In silico structural modeling of WHT-2 indicates that the editing of one nucleotide in the wht-2 mRNA leads to the substitution of threonine with alanine at residue 124 in the WHT-2 protein, changing hydrogen bonds in this region. Thus, we propose a model where wht-2 is edited by ADR-2, which promotes optimal export of uric acid, a known substrate of WHT-2 and a product of XDH-1 activity. In the absence of editing, uric acid export is limited, provoking a reduction in xdh-1 transcription to limit uric acid production and maintain cellular homeostasis. As a result, elevation of uric acid is protective against dopaminergic neuronal cell death. In turn, increased levels of uric acid are associated with a decrease in ROS production. Further, downregulation of xdh-1 is protective against PD pathologies because decreased levels of XDH-1 correlate to a concomitant reduction in xanthine oxidase (XO), the form of the protein whose by-product is superoxide anion. These data indicate that modifying specific targets of RNA editing may represent a promising therapeutic strategy for PD.
Bcl-xL Is Required by Primary Hippocampal Neurons during Development to Support Local Energy Metabolism at Neurites
(MDPI, 2021) Jansen, Joseph; Scott, Madison; Amjad, Emma; Stumpf, Allison; Lackey, Kimberly H.; Caldwell, Kim A.; Park, Han-A; University of Alabama Tuscaloosa
Simple Summary B-cell lymphoma-extra large (Bcl-xL) is an anti-apoptotic protein that regulates energy metabolism in neurons. In this study, we found that primary hippocampal neurons transduced with Bcl-xL shRNA or treated with a pharmacological inhibitor of Bxl-xL had a decrease in the population of motile mitochondria. Primary hippocampal neurons lacking Bcl-xL failed to retain ATP at their neurites, which hindered the formation of complex neurite arbors, and ultimately had enhanced vulnerability to excitotoxic challenge. B-cell lymphoma-extra large (Bcl-xL) is a mitochondrial protein known to inhibit mitochondria-dependent intrinsic apoptotic pathways. An increasing number of studies have demonstrated that Bcl-xL is critical in regulating neuronal energy metabolism and has a protective role in pathologies associated with an energy deficit. However, it is less known how Bcl-xL regulates physiological processes of the brain. In this study, we hypothesize that Bcl-xL is required for neurite branching and maturation during neuronal development by improving local energy metabolism. We found that the absence of Bcl-xL in rat primary hippocampal neurons resulted in mitochondrial dysfunction. Specifically, the ATP/ADP ratio was significantly decreased in the neurites of Bcl-xL depleted neurons. We further found that neurons transduced with Bcl-xL shRNA or neurons treated with ABT-263, a pharmacological inhibitor of Bcl-xL, showed impaired mitochondrial motility. Neurons lacking Bcl-xL had significantly decreased anterograde and retrograde movement of mitochondria and an increased stationary mitochondrial population when Bcl-xL was depleted by either means. These mitochondrial defects, including loss of ATP, impaired normal neurite development. Neurons lacking Bcl-xL showed significantly decreased neurite arborization, growth and complexity. Bcl-xL depleted neurons also showed impaired synapse formation. These neurons showed increased intracellular calcium concentration and were more susceptible to excitotoxic challenge. Bcl-xL may support positioning of mitochondria at metabolically demanding regions of neurites like branching points. Our findings suggest a role for Bcl-xL in physiological regulation of neuronal growth and development.
Calcineurin determines toxic versus beneficial responses to alpha-synuclein
(National Academy of the Sciences, 2014) Caraveo, Gabriela; Auluck, Pavan K.; Whitesell, Luke; Chung, Chee Yeun; Baru, Valeriya; Mosharov, Eugene V.; Yan, Xiaohui; Ben-Johny, Manu; Soste, Martin; Picotti, Paola; Kim, Hanna; Caldwell, Kim A.; Caldwell, Guy A.; Sulzer, David; Yue, David T.; Lindquist, Susan; Massachusetts Institute of Technology (MIT); Whitehead Institute; Howard Hughes Medical Institute; Harvard University; Massachusetts General Hospital; Harvard Medical School; Columbia University; University of Alabama Tuscaloosa; Johns Hopkins University
Calcineurin (CN) is a highly conserved Ca2+-calmodulin (CaM)dependent phosphatase that senses Ca2+ concentrations and transduces that information into cellular responses. Ca2+ homeostasis is disrupted by alpha-synuclein (alpha-syn), a small lipid binding protein whose misfolding and accumulation is a pathological hallmark of several neurodegenerative diseases. We report that alpha-syn, from yeast to neurons, leads to sustained highly elevated levels of cytoplasmic Ca2+, thereby activating a CaM-CN cascade that engages substrates that result in toxicity. Surprisingly, complete inhibition of CN also results in toxicity. Limiting the availability of CaM shifts CN's spectrum of substrates toward protective pathways. Modulating CN or CN's substrates with highly selective genetic and pharmacological tools (FK506) does the same. FK506 crosses the blood brain barrier, is well tolerated in humans, and is active in neurons and glia. Thus, a tunable response to CN, which has been conserved for a billion years, can be targeted to rebalance the phosphatase's activities from toxic toward beneficial substrates. These findings have immediate therapeutic implications for synucleinopathies.
Chemical Compensation of Mitochondrial Phospholipid Depletion in Yeast and Animal Models of Parkinson's Disease
(PLOS, 2016) Wang, Shaoxiao; Zhang, Siyuan; Xu, Chuan; Barron, Addie; Galiano, Floyd; Patel, Dhaval; Lee, Yong Joo; Caldwell, Guy A.; Caldwell, Kim A.; Witt, Stephan N.; Louisiana State University Health Sciences Center at Shreveport; University of Alabama Tuscaloosa
We have been investigating the role that phosphatidylethanolamine (PE) and phosphatidylcholine (PC) content plays in modulating the solubility of the Parkinson's disease protein alpha-synuclein (alpha-syn) using Saccharomyces cerevisiae and Caenorhabditis elegans. One enzyme that synthesizes PE is the conserved enzyme phosphatidylserine decarboxylase (Psd1/yeast; PSD-1/worms), which is lodged in the inner mitochondrial membrane. We previously found that decreasing the level of PE due to knockdown of Psd1/psd-1 affects the homeostasis of alpha-syn in vivo. In S. cerevisiae, the co-occurrence of low PE and alpha-syn in psd1 Delta cells triggers mitochondrial defects, stress in the endoplasmic reticulum, misprocessing of glycosylphosphatidylinositol-anchored proteins, and a 3-fold increase in the level of alpha-syn. The goal of this study was to identify drugs that rescue this phenotype. We screened the Prestwick library of 1121 Food and Drug Administration-approved drugs using psd1 Delta + alpha-syn cells and identified cyclosporin A, meclofenoxate hydrochloride, and sulfaphenazole as putative protective compounds. The protective activity of these drugs was corroborated using C. elegans in which a-syn is expressed specifically in the dopaminergic neurons, with psd-1 depleted by RNAi. Worm populations were examined for dopaminergic neuron survival following psd-1 knockdown. Exposure to cyclosporine, meclofenoxate, and sulfaphenazole significantly enhanced survival at day 7 in alpha-syn-expressing worm populations whereby 50-55% of the populations displayed normal neurons, compared to only 10-15% of untreated animals. We also found that all three drugs rescued worms expressing alpha-syn in dopaminergic neurons that were deficient in the phospholipid cardiolipin following cardiolipin synthase (crls-1) depletion by RNAi. We discuss how these drugs might block a-syn pathology in dopaminergic neurons.
Clioquinol promotes the degradation of metal-dependent amyloid-beta (A beta) oligomers to restore endocytosis and ameliorate A beta toxicity
(National Academy of the Sciences, 2014) Matlack, Kent E. S.; Tardiff, Daniel F.; Narayan, Priyanka; Hamamichi, Shusei; Caldwell, Kim A.; Caldwell, Guy A.; Lindquist, Susan; Massachusetts Institute of Technology (MIT); Whitehead Institute; University of Alabama Tuscaloosa; Howard Hughes Medical Institute
Alzheimer's disease (AD) is a common, progressive neurodegenerative disorder without effective disease-modifying therapies. The accumulation of amyloid-beta peptide (A beta) is associated with AD. However, identifying new compounds that antagonize the underlying cellular pathologies caused by A beta has been hindered by a lack of cellular models amenable to high-throughput chemical screening. To address this gap, we use a robust and scalable yeast model of A beta toxicity where the A beta peptide transits through the secretory and endocytic compartments as it does in neurons. The pathogenic A beta 1-42 peptide forms more oligomers and is more toxic than A beta 1-40 and genome-wide genetic screens identified genes that are known risk factors for AD. Here, we report an unbiased screen of similar to 140,000 compounds for rescue of A beta toxicity. Of similar to 30 hits, several were 8-hydroxyquinolines (8-OHQs). Clioquinol (CQ), an 8-OHQ previously reported to reduce A beta burden, restore metal homeostasis, and improve cognition in mouse AD models, was also effective and rescued the toxicity of A beta secreted from glutamatergic neurons in Caenorhabditis elegans. In yeast, CQ dramatically reduced A beta peptide levels in a copper-dependent manner by increasing degradation, ultimately restoring endocytic function. This mirrored its effects on copper-dependent oligomer formation in vitro, which was also reversed by CQ. This unbiased screen indicates that copper-dependent A beta oligomer formation contributes to A beta toxicity within the secretory/endosomal pathways where it can be targeted with selective metal binding compounds. Establishing the ability of the A beta yeast model to identify disease-relevant compounds supports its further exploitation as a validated early discovery platform.
A conformational switch driven by phosphorylation regulates the activity of the evolutionarily conserved SNARE Ykt6
(National Academy of the Sciences, 2021) McGrath, Kaitlyn; Agarwal, Shivani; Tonelli, Marco; Dergai, Mykola; Gaeta, Anthony L.; Shum, Andrew K.; Lacoste, Jessica; Zhang, Yongbo; Wen, Wenyu; Chung, Daayun; Wiersum, Grant; Shevade, Aishwarya; Zaichick, Sofia; van Rossum, Damian B.; Shuvalova, Ludmilla; Savas, Jeffrey N.; Kuchin, Sergei; Taipale, Mikko; Caldwell, Kim A.; Caldwell, Guy A.; Fasshauer, Dirk; Caraveo, Gabriela; Northwestern University; Feinberg School of Medicine; University of Wisconsin Madison; University of Lausanne; University of Alabama Tuscaloosa; University of Toronto; Fudan University; University of Wisconsin Milwaukee; Pennsylvania State University; Penn State Health; University of Alabama Birmingham; University of Pennsylvania
Ykt6 is a soluble N-ethylmaleimide sensitive factor activating protein receptor (SNARE) critically involved in diverse vesicular fusion pathways. While most SNAREs rely on transmembrane domains for their activity, Ykt6 dynamically cycles between the cytosol and membrane-bound compartments where it is active. The mechanism that regulates these transitions and allows Ykt6 to achieve specificity toward vesicular pathways is unknown. Using a Parkinson's disease (PD) model, we found that Ykt6 is phosphorylated at an evolutionarily conserved site which is regulated by Ca2+ signaling. Through a multidisciplinary approach, we show that phosphorylation triggers a conformational change that allows Ykt6 to switch from a closed cytosolic to an open membrane-bound form. In the phosphorylated open form, the spectrum of protein interactions changes, leading to defects in both the secretory and autophagy pathways, enhancing toxicity in PD models. Our studies reveal a mechanism by which Ykt6 conformation and activity are regulated with potential implications for PD.
Conserved nicotine-activated neuroprotective pathways involve mitochondrial stress
(Cell Press, 2021) Nourse, J. Brucker, Jr.; Harshefi, Gilad; Marom, Adi; Karmi, Abdelrahaman; Ben-Ami, Hagit Cohen; Caldwell, Kim A.; Caldwell, Guy A.; Treinin, Millet; University of Alabama Tuscaloosa; University of Alabama Birmingham; Hebrew University of Jerusalem
Tobacco smoking is a risk factor for several human diseases. Conversely, smoking also reduces the prevalence of Parkinson's disease, whose hallmark is degeneration of substantia nigra dopaminergic neurons (DNs). We use C. elegans as a model to investigate whether tobacco-derived nicotine activates nicotinic acetylcholine receptors (nAChRs) to selectively protect DNs. Using this model, we demonstrate conserved functions of DN-expressed nAChRs. We find that DOP-2, a D3-receptor homolog; MCU-1, a mitochondrial calcium uniporter; PINK-1 (PTEN-induced kinase 1); and PDR-1 (Parkin) are required for nicotine-mediated protection of DNs. Together, our results support involvement of a calcium-modulated, mitochondrial stress-activated PINK1/Parkin-dependent pathway in nicotine-induced neuroprotection. This suggests that nicotine-selective protection of substantia nigra DNs is due to the confluence of two factors: first, their unique vulnerability to mitochondrial stress, which is mitigated by increased mitochondrial quality control due to PINK1 activation, and second, their specific expression of D3-receptors.
Cyclized NDGA modifies dynamic alpha-synuclein monomers preventing aggregation and toxicity
(Nature Portfolio, 2019) Daniels, Malcolm J.; Nourse, J. Brucker, Jr.; Kim, Hanna; Sainati, Valerio; Schiavina, Marco; Murrali, Maria Grazia; Pan, Buyan; Ferrie, John J.; Haney, Conor M.; Moons, Rani; Gould, Neal S.; Natalello, Antonino; Grandori, Rita; Sobott, Frank; Petersson, E. James; Rhoades, Elizabeth; Pierattelli, Roberta; Felli, Isabella; Uversky, Vladimir N.; Caldwell, Kim A.; Caldwell, Guy A.; Krol, Edward S.; Ischiropoulos, Harry; University of Pennsylvania; Pennsylvania Medicine; University of Alabama Tuscaloosa; University of Florence; University of Antwerp; Childrens Hospital of Philadelphia; University of Milano-Bicocca; University of Leeds; University of South Florida; Russian Academy of Sciences; University of Saskatchewan
Growing evidence implicates alpha-synuclein aggregation as a key driver of neurodegeneration in Parkinson's disease (PD) and other neurodegenerative disorders. Herein, the molecular and structural mechanisms of inhibiting alpha-synuclein aggregation by novel analogs of nordihydroguaiaretic acid (NDGA), a phenolic dibenzenediol lignan, were explored using an array of biochemical and biophysical methodologies. NDGA analogs induced modest, progressive compaction of monomeric alpha-synuclein, preventing aggregation into amyloid-like fibrils. This conformational remodeling preserved the dynamic adoption of alpha-helical conformations, which are essential for physiological membrane interactions. Oxidation-dependent NDGA cyclization was required for the interaction with monomeric alpha-synuclein. NDGA analog-pretreated alpha-synuclein did not aggregate even without NDGA-analogs in the aggregation mixture. Strikingly, NDGA-pretreated alpha-synuclein suppressed aggregation of naive untreated aggregation-competent monomeric a-synuclein. Further, cyclized NDGA reduced alpha-synuclein-driven neurodegeneration in Caenorhabditis elegans. The cyclized NDGA analogs may serve as a platform for the development of small molecules that stabilize aggregation-resistant alpha-synuclein monomers without interfering with functional conformations yielding potential therapies for PD and related disorders.
Different 8-Hydroxyquinolines Protect Models of TDP-43 Protein, alpha-Synuclein, and Polyglutamine Proteotoxicity through Distinct Mechanisms
(American Society of Biochemistry and Molecular Biology, 2012) Tardiff, Daniel F.; Tucci, Michelle L.; Caldwell, Kim A.; Caldwell, Guy A.; Lindquist, Susan; Massachusetts Institute of Technology (MIT); Whitehead Institute; University of Alabama Tuscaloosa; Howard Hughes Medical Institute
No current therapies target the underlying cellular pathologies of age-related neurodegenerative diseases. Model organisms provide a platform for discovering compounds that protect against the toxic, misfolded proteins that initiate these diseases. One such protein, TDP-43, is implicated in multiple neurodegenerative diseases, including amyotrophic lateral sclerosis and frontotemporal lobar degeneration. In yeast, TDP-43 expression is toxic, and genetic modifiers first discovered in yeast have proven to modulate TDP-43 toxicity in both neurons and humans. Here, we describe a phenotypic screen for small molecules that reverse TDP-43 toxicity in yeast. One group of hit compounds was 8-hydroxyquinolines (8-OHQ), a class of clinically relevant bioactive metal chelators related to clioquinol. Surprisingly, in otherwise wild-type yeast cells, different 8-OHQs had selectivity for rescuing the distinct toxicities caused by the expression of TDP-43, alpha-synuclein, or polyglutamine proteins. In fact, each 8-OHQ synergized with the other, clearly establishing that they function in different ways. Comparative growth and molecular analyses also revealed that 8-OHQs have distinct metal chelation and ionophore activities. The diverse bioactivity of 8-OHQs indicates that altering different aspects of metal homeostasis and/or metalloprotein activity elicits distinct protective mechanisms against several neurotoxic proteins. Indeed, phase II clinical trials of an 8-OHQ has produced encouraging results in modifying Alzheimer disease. Our unbiased identification of 8-OHQs in a yeast TDP-43 toxicity model suggests that tailoring 8-OHQ activity to a particular neurodegenerative disease may be a viable therapeutic strategy.
Dihydropyrimidine-Thiones and Clioquinol Synergize To Target beta-Amyloid Cellular Pathologies through a Metal-Dependent Mechanism
(American Chemical Society, 2017) Tardiff, Daniel F.; Brown, Lauren E.; Yan, Xiaohui; Trilles, Richard; Jui, Nathan T.; Barrasa, M. Inmaculada; Caldwell, Kim A.; Caldwell, Guy A.; Schaus, Scott E.; Lindquist, Susan; Massachusetts Institute of Technology (MIT); Whitehead Institute; Boston University; University of Alabama Tuscaloosa; Howard Hughes Medical Institute; Emory University
The lack of therapies for neurodegenerative diseases arises from our incomplete understanding of their underlying cellular toxicities and the limited number of predictive model systems. It is critical that we develop approaches to identify novel targets and lead compounds. Here, a phenotypic screen of yeast proteinopathy models identified dihydropyrimidine-thiones (DHPM-thiones) that selectively rescued the toxicity caused by beta-amyloid (Afi), the peptide implicated in Alzheimer's disease. Rescue of A beta toxicity by DHPM-thiones occurred through a metal dependent mechanism of action. The bioactivity was distinct, however, from that of the 8-hydroxyquinoline clioquinol (CQ). These structurally dissimilar compounds strongly synergized at concentrations otherwise not competent to reduce toxicity. Cotreatment ameliorated Afi toxicity by reducing Afi levels and restoring functional vesicle trafficking. Notably, these low doses significantly reduced deleterious off-target effects caused by CQon mitochondria at higher concentrations. Both single and combinatorial treatments also reduced death of neurons expressing A beta in a nematode, indicating that DHPM-thiones target a conserved protective mechanism. Furthermore, this conserved activity suggests that expression of the A beta peptide causes similar cellular pathologies from yeast to neurons. Our identification of a new cytoprotective scaffold that requires metal-binding underscores the critical role of metal phenomenology in mediating Afi toxicity. Additionally, our findings demonstrate the valuable potential of synergistic compounds to enhance on-target activities, while mitigating deleterious off-target effects. The identification and prosecution of synergistic compounds could prove useful for developing AD therapeutics where combination therapies may be required to antagonize diverse pathologies.
Distinct functional roles of Vps41-mediated neuroprotection in Alzheimer's and Parkinson's disease models of neurodegeneration
(Oxford University Press, 2018) Griffin, Edward F.; Yan, Xiaohui; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
Commonalities and, in some cases, pathological overlap between neurodegenerative diseases have led to speculation that targeting of underlying mechanisms might be of potentially shared therapeutic benefit. Alzheimer's disease is characterized by the formation of plaques, composed primarily of the amyloid-beta 1-42 (A beta) peptide in the brain, resulting in neurodegeneration. Previously, we have shown that overexpression of the lysosomal-trafficking protein, human Vps41 (hVps41), is neuroprotective in a transgenic worm model of Parkinson's disease, wherein progressive dopaminergic neurodegeneration is induced by alpha-synuclein overexpression. Here, we report the results of a systematic comparison of hVps41-mediated neuroprotection between alpha-synuclein and A beta in transgenic nematode models of Caenorhabditis elegans. Our results indicate that an ARF-like GTPase gene product, ARL-8, mitigates endocytic neurodegeneration in a VPS-41-dependent manner, rather than through RAB-7 and AP3 as with alpha-synuclein. Furthermore, the neuroprotective effect of ARL-8 or hVps41 appears to be dependent on their colocalization and the activity of ARL-8. Additionally, we demonstrate that the LC3 orthologue, LGG-2, plays a critical role in A beta toxicity with ARL-8. Further analysis of functional effectors of A beta protein processing via the lysosomal pathway will assist in the elucidation of the underlying mechanism involving VPS-41-mediated neuroprotection. These results reveal functional distinctions in the intracellular management of neurotoxic proteins that serve to better inform the path for development of therapeutic interventions to halt neurodegeneration.
Dopamine induces soluble alpha-synuclein oligomers and nigrostriatal degeneration
(Nature Portfolio, 2017) Mor, Danielle E.; Tsika, Elpida; Mazzulli, Joseph R.; Gould, Neal S.; Kim, Hanna; Daniels, Malcolm J.; Doshi, Shachee; Gupta, Preetika; Grossman, Jennifer L.; Tan, Victor X.; Kalb, Robert G.; Caldwell, Kim A.; Caldwell, Guy A.; Wolfe, John H.; Ischiropoulos, Harry; University of Pennsylvania; Pennsylvania Medicine; Swiss Federal Institutes of Technology Domain; Ecole Polytechnique Federale de Lausanne; Northwestern University; Feinberg School of Medicine; Childrens Hospital of Philadelphia; University of Alabama Tuscaloosa; State University of New York (SUNY) Downstate Medical Center
Parkinson's disease (PD) is defined by the loss of dopaminergic neurons in the substantia nigra and the formation of Lewy body inclusions containing aggregated alpha-synuclein. Efforts to explain dopamine neuron vulnerability are hindered by the lack of dopaminergic cell death in a-synuclein transgenic mice. To address this, we manipulated both dopamine levels and alpha-synuclein expression. Nigrally targeted expression of mutant tyrosine hydroxylase with enhanced catalytic activity increased dopamine levels without damaging neurons in non-transgenic mice. In contrast, raising dopamine levels in mice expressing human A53T mutant alpha-synuclein induced progressive nigrostriatal degeneration and reduced locomotion. Dopamine elevation in A53T mice increased levels of potentially toxic alpha-synuclein oligomers, resulting in conformationally and functionally modified species. Moreover, in genetically tractable Caenorhabditis elegans models, expression of alpha-synuclein mutated at the site of interaction with dopamine prevented dopamine-induced toxicity. These data suggest that a unique mechanism links two cardinal features of PD: dopaminergic cell death and alpha-synuclein aggregation.
Dysregulation of the Mitochondrial Unfolded Protein Response Induces Non-Apoptotic Dopaminergic Neurodegeneration in C-elegans Models of Parkinson's Disease
(Society for Neuroscience, 2017) Martinez, Bryan A.; Petersen, Daniel A.; Gaeta, Anthony L.; Stanley, Samuel P.; Caldwell, Guy A.; Caldwell, Kim A.; University of Alabama Tuscaloosa
Due to environmental insult or innate genetic deficiency, protein folding environments of the mitochondrial matrix are prone to dysregulation, prompting the activation of a specific organellar stress-response mechanism, the mitochondrial unfolded protein response (UPRMT). In Caenorhabditis elegans, mitochondrial damage leads to nuclear translocation of the ATFS-1 transcription factor to activate the UPRMT. After short-term acute stress has been mitigated, the UPRMT is eventually suppressed to restore homeostasis to C. elegans hermaphrodites. In contrast, and reflective of the more chronic nature of progressive neurodegenerative disorders such as Parkinson's disease (PD), here, we report the consequences of prolonged, cell-autonomous activation of the UPRMT in C. elegans dopaminergic neurons. We reveal that neuronal function and integrity decline rapidly with age, culminating in activity-dependent, non-apoptotic cell death. In a PD-like context wherein transgenic nematodes express the Lewy body constituent protein alpha-synuclein(alpha S), we not only find that this protein and its PD-associated disease variants have the capacity to induce the UPRMT, but also that coexpression of alpha S and ATFS-1-associated dysregulation of the UPRMT synergistically potentiate dopaminergic neurotoxicity. This genetic interaction is in parallel to mitophagic pathways dependent on the C. elegans PINK1 homolog, which is necessary for cellular resistance to chronic malfunction of the UPRMT. Given the increasingly recognized role of mitochondrial quality control in neurodegenerative diseases, these studies illustrate, for the first time, an insidious aspect of mitochondrial signaling in which the UPRMT pathway, under diseaseassociated, context-specific dysregulation, exacerbates disruption of dopaminergic neurons in vivo, resulting in the neurodegeneration characteristic of PD.
The effects of pdr1, djr1.1 and pink1 loss in manganese-induced toxicity and the role of alpha-synuclein in C-elegans
(Royal Society of Chemistry, 2014) Bornhorst, Julia; Chakraborty, Sudipta; Meyer, Soeren; Lohren, Hanna; Brinkhaus, Sigrid Grosse; Knight, Adam L.; Caldwell, Kim A.; Caldwell, Guy A.; Karst, Uwe; Schwerdtle, Tanja; Bowman, Aaron; Aschner, Michael; University of Munster; Vanderbilt University; University of Alabama Tuscaloosa; Yeshiva University; Albert Einstein College of Medicine
Parkinson's disease (PD) is a neurodegenerative brain disorder characterized by selective dopaminergic (DAergic) cell loss that results in overt motor and cognitive deficits. Current treatment options exist to combat PD symptomatology, but are unable to directly target its pathogenesis due to a lack of knowledge concerning its etiology. Several genes have been linked to PD, including three genes associated with an early-onset familial form: parkin, pink1 and dj1. All three genes are implicated in regulating oxidative stress pathways. Another hallmark of PD pathophysiology is Lewy body deposition, associated with the gain-of-function genetic risk factor a-synuclein. The function of a-synuclein is poorly understood, as it shows both neurotoxic and neuroprotective activities in PD. Using the genetically tractable invertebrate Caenorhabditis elegans (C. elegans) model system, the neurotoxic or neuroprotective role of a-synuclein upon acute Mn exposure in the background of mutated pdr1, pink1 or djr1.1 was examined. The pdr1 and djr1.1 mutants showed enhanced Mn accumulation and oxidative stress that was reduced by a-synuclein. Moreover, DAergic neurodegeneration, while unchanged with Mn exposure, returned to wild-type (WT) levels for pdr1, but not djr1.1 mutants expressing a-synuclein. Taken together, this study uncovers a novel, neuroprotective role for WT human a-synuclein in attenuating Mn-induced toxicity in the background of PD-associated genes, and further supports the role of extracellular dopamine in exacerbating Mn neurotoxicity.
Found in Translation: The Utility of C. elegans Alpha-Synuclein Models of Parkinson's Disease
(MDPI, 2019) Gaeta, Anthony L.; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
Parkinson's Disease (PD) is the second-most common neurodegenerative disease in the world, yet the fundamental and underlying causes of the disease are largely unknown, and treatments remain sparse and impotent. Several biological systems have been employed to model the disease but the nematode roundworm Caenorhabditis elegans (C. elegans) shows unique promise among these to disinter the elusive factors that may prevent, halt, and/or reverse PD phenotypes. Some of the most salient of these C. elegans models of PD are those that position the misfolding-prone protein alpha-synuclein (-syn), a hallmark pathological component of PD, as the primary target for scientific interrogation. By transgenic expression of human -syn in different tissues, including dopamine neurons and muscle cells, the primary cellular phenotypes of PD in humans have been recapitulated in these C. elegans models and have already uncovered multifarious genetic factors and chemical compounds that attenuate dopaminergic neurodegeneration. This review describes the paramount discoveries obtained through the application of different -syn models of PD in C. elegans and highlights their established utility and respective promise to successfully uncover new conserved genetic modifiers, functional mechanisms, therapeutic targets and molecular leads for PD with the potential to translate to humans.
From bugs to bedside: functional annotation of human genetic variation for neurological disorders using invertebrate models
(Oxford University Press, 2022) Mew, Melanie; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
The exponential accumulation of DNA sequencing data has opened new avenues for discovering the causative roles of single-nucleotide polymorphisms (SNPs) in neurological diseases. The opportunities emerging from this are staggering, yet only as good as our abilities to glean insights from this surplus of information. Whereas computational biology continues to improve with respect to predictions and molecular modeling, the differences between in silico and in vivo analysis remain substantial. Invertebrate in vivo model systems represent technically advanced, experimentally mature, high-throughput, efficient and cost-effective resources for investigating a disease. With a decades-long track record of enabling investigators to discern function from DNA, fly (Drosophila) and worm (Caenorhabditis elegans) models have never been better poised to serve as living engines of discovery. Both of these animals have already proven useful in the classification of genetic variants as either pathogenic or benign across a range of neurodevelopmental and neurodegenerative disorders-including autism spectrum disorders, ciliopathies, amyotrophic lateral sclerosis, Alzheimer's and Parkinson's disease. Pathogenic SNPs typically display distinctive phenotypes in functional assays when compared with null alleles and frequently lead to protein products with gain-of-function or partial loss-of-function properties that contribute to neurological disease pathogenesis. The utility of invertebrates is logically limited by overt differences in anatomical and physiological characteristics, and also the evolutionary distance in genome structure. Nevertheless, functional annotation of disease-SNPs using invertebrate models can expedite the process of assigning cellular and organismal consequences to mutations, ascertain insights into mechanisms of action, and accelerate therapeutic target discovery and drug development for neurological conditions.
Functional Analysis of VPS41-Mediated Neuroprotection in Caenorhabditis elegans and Mammalian Models of Parkinson's Disease
(Society for Neuroscience, 2012) Harrington, Adam J.; Yacoubian, Talene A.; Slone, Sunny R.; Caldwell, Kim A.; Caldwell, Guy A.; University of Alabama Tuscaloosa; University of Alabama Birmingham
Disruption of the lysosomal system has emerged as a key cellular pathway in the neurotoxicity of alpha-synuclein (alpha-syn) and the progression of Parkinson's disease (PD). A large-scale RNA interference (RNAi) screen using Caenorhabditis elegans identified VPS-41, a multidomain protein involved in lysosomal protein trafficking, as a modifier of alpha-syn accumulation and dopaminergic neuron degeneration (Hamamichi et al., 2008). Previous studies have shown a conserved neuroprotective function of human VPS41 (hVPS41) against PD-relevant toxins in mammalian cells and C. elegans neurons (Ruan et al., 2010). Here, we report that both the AP-3 (heterotetrameric adaptor protein complex) interaction domain and clathrin heavy-chain repeat domain are required for protecting C. elegans dopaminergic neurons from alpha-syn-induced neurodegeneration, as well as to prevent alpha-syn inclusion formation in an H4 human neuroglioma cell model. Using mutant C. elegans and neuron-specific RNAi, we revealed that hVPS41 requires both a functional AP-3 (heterotetrameric adaptor protein complex) and HOPS (homotypic fusion and vacuole protein sorting)-tethering complex to elicit neuroprotection. Interestingly, two nonsynonymous single-nucleotide polymorphisms found within the AP-3 interacting domain of hVPS41 attenuated the neuroprotective property, suggestive of putative susceptibility factors for PD. Furthermore, we observed a decrease in alpha-syn protein level when hVPS41 was overexpressed in human neuroglioma cells. Thus, the neuroprotective capacity of hVPS41 may be a consequence of enhanced clearance of misfolded and aggregated proteins, including toxic alpha-syn species. These data reveal the importance of lysosomal trafficking in maintaining cellular homeostasis in the presence of enhanced alpha-syn expression and toxicity. Our results support hVPS41 as a potential novel therapeutic target for the treatment of synucleinopathies like PD.

Browsing by Author "Caldwell, Kim A."

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