scholarly journals Neuron-based high-content assay and screen for CNS active mitotherapeutics

2020 ◽  
Vol 6 (2) ◽  
pp. eaaw8702 ◽  
Author(s):  
Boglarka H. Varkuti ◽  
Miklos Kepiro ◽  
Ze Liu ◽  
Kyle Vick ◽  
Yosef Avchalumov ◽  
...  
Keyword(s):  
Small Molecule ◽  
Hippocampal Neurons ◽  
Mitochondrial Dynamics ◽  
Synaptic Activity ◽  
Glutamate Toxicity ◽  
Primary Neurons ◽  
Screening Assay ◽  
Dynamics And Function ◽  
Small Molecule Modulators ◽  
And Function

Impaired mitochondrial dynamics and function are hallmarks of many neurological and psychiatric disorders, but direct screens for mitotherapeutics using neurons have not been reported. We developed a multiplexed and high-content screening assay using primary neurons and identified 67 small-molecule modulators of neuronal mitostasis (MnMs). Most MnMs that increased mitochondrial content, length, and/or health also increased mitochondrial function without altering neurite outgrowth. A subset of MnMs protected mitochondria in primary neurons from Aβ(1–42) toxicity, glutamate toxicity, and increased oxidative stress. Some MnMs were shown to directly target mitochondria. The top MnM also increased the synaptic activity of hippocampal neurons and proved to be potent in vivo, increasing the respiration rate of brain mitochondria after administering the compound to mice. Our results offer a platform that directly queries mitostasis processes in neurons, a collection of small-molecule modulators of mitochondrial dynamics and function, and candidate molecules for mitotherapeutics.

scholarly journals Synaptic activity controls autophagic vacuole motility and function in dendrites

2021 ◽  
Vol 220 (6) ◽  
Author(s):  
Vineet Vinay Kulkarni ◽  
Anip Anand ◽  
Jessica Brandt Herr ◽  
Christina Miranda ◽  
Maria Chalokh Vogel ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Degradation Pathway ◽  
Autophagic Vacuole ◽  
Synaptic Activity ◽  
Lysosomal Degradation ◽  
Autophagy Pathway ◽  
Dynamics And Function ◽  
And Function ◽  
Activity Dependent ◽  
Stimulation Of

Macroautophagy (hereafter “autophagy”) is a lysosomal degradation pathway that is important for learning and memory, suggesting critical roles for autophagy at the neuronal synapse. Little is known, however, about the molecular details of how autophagy is regulated with synaptic activity. Here, we used live-cell confocal microscopy to define the autophagy pathway in primary hippocampal neurons under various paradigms of synaptic activity. We found that synaptic activity regulates the motility of autophagic vacuoles (AVs) in dendrites. Stimulation of synaptic activity dampens AV motility, whereas silencing synaptic activity induces AV motility. Activity-dependent effects on dendritic AV motility are local and reversible. Importantly, these effects are compartment specific, occurring in dendrites and not in axons. Most strikingly, synaptic activity increases the presence of degradative autolysosomes in dendrites and not in axons. On the basis of our findings, we propose a model whereby synaptic activity locally controls AV dynamics and function within dendrites that may regulate the synaptic proteome.

scholarly journals Critical requirement of SOS1 RAS-GEF function for mitochondrial dynamics, metabolism, and redox homeostasis

Oncogene ◽  
2021 ◽  
Author(s):  
Rósula García-Navas ◽  
Pilar Liceras-Boillos ◽  
Carmela Gómez ◽  
Fernando C. Baltanás ◽  
Nuria Calzada ◽  
...  
Keyword(s):  
Mitochondrial Dynamics ◽  
Redox Homeostasis ◽  
Atp Production ◽  
Oxidative Energy Metabolism ◽  
Ras Activation ◽  
Mitochondrial Defects ◽  
Critical Requirement ◽  
Dynamics And Function ◽  
And Function ◽  
And Control

AbstractSOS1 ablation causes specific defective phenotypes in MEFs including increased levels of intracellular ROS. We showed that the mitochondria-targeted antioxidant MitoTEMPO restores normal endogenous ROS levels, suggesting predominant involvement of mitochondria in generation of this defective SOS1-dependent phenotype. The absence of SOS1 caused specific alterations of mitochondrial shape, mass, and dynamics accompanied by higher percentage of dysfunctional mitochondria and lower rates of electron transport in comparison to WT or SOS2-KO counterparts. SOS1-deficient MEFs also exhibited specific alterations of respiratory complexes and their assembly into mitochondrial supercomplexes and consistently reduced rates of respiration, glycolysis, and ATP production, together with distinctive patterns of substrate preference for oxidative energy metabolism and dependence on glucose for survival. RASless cells showed defective respiratory/metabolic phenotypes reminiscent of those of SOS1-deficient MEFs, suggesting that the mitochondrial defects of these cells are mechanistically linked to the absence of SOS1-GEF activity on cellular RAS targets. Our observations provide a direct mechanistic link between SOS1 and control of cellular oxidative stress and suggest that SOS1-mediated RAS activation is required for correct mitochondrial dynamics and function.

Does metformin modulate mitochondrial dynamics and function in type 2 diabetic patients?

2021 ◽  
Author(s):  
Aranzazu Martinez de Marañón ◽  
Francisco Gerardo Canet ◽  
Zaida Abad-Jimenez ◽  
Ana Jover ◽  
Carlos Morillas ◽  
...  
Keyword(s):  
Mitochondrial Dynamics ◽  
Diabetic Patients ◽  
Type 2 Diabetic Patients ◽  
Dynamics And Function ◽  
And Function ◽  
Type 2 Diabetic

scholarly journals Detection of Small-Molecule Modulators of Actin-Myosin Structure and Function using High-Throughput Time-Resolved FRET

2017 ◽  
Vol 112 (3) ◽  
pp. 237a
Author(s):  
Piyali Guhathakurta ◽  
Ewa Prochniewicz ◽  
Kurt C. Peterson ◽  
Benjamin D. Grant ◽  
Gregory D. Gillispie ◽  
...  
Keyword(s):  
Small Molecule ◽  
High Throughput ◽  
Structure And Function ◽  
Throughput Time ◽  
Time Resolved ◽  
Small Molecule Modulators ◽  
And Function

scholarly journals Corticotropin-releasing hormone (CRH) alters mitochondrial morphology and function by activating the NF-kB-DRP1 axis in hippocampal neurons

2020 ◽  
Vol 11 (11) ◽  
Author(s):  
Chiara R. Battaglia ◽  
Silvia Cursano ◽  
Enrico Calzia ◽  
Alberto Catanese ◽  
Tobias M. Boeckers
Keyword(s):  
Hippocampal Neurons ◽  
Morphological Changes ◽  
Mitochondrial Dynamics ◽  
Synaptic Activity ◽  
Mitochondrial Activity ◽  
Mitochondrial Morphology ◽  
Significant Shift ◽  
Corticotropin Releasing Hormone ◽  
Releasing Hormone

AbstractNeuronal stress-adaptation combines multiple molecular responses. We have previously reported that thorax trauma induces a transient loss of hippocampal excitatory synapses mediated by the local release of the stress-related hormone corticotropin-releasing hormone (CRH). Since a physiological synaptic activity relies also on mitochondrial functionality, we investigated the direct involvement of mitochondria in the (mal)-adaptive changes induced by the activation of neuronal CRH receptors 1 (CRHR1). We observed, in vivo and in vitro, a significant shift of mitochondrial dynamics towards fission, which correlated with increased swollen mitochondria and aberrant cristae. These morphological changes, which are associated with increased NF-kB activity and nitric oxide concentrations, correlated with a pronounced reduction of mitochondrial activity. However, ATP availability was unaltered, suggesting that neurons maintain a physiological energy metabolism to preserve them from apoptosis under CRH exposure. Our findings demonstrate that stress-induced CRHR1 activation leads to strong, but reversible, modifications of mitochondrial dynamics and morphology. These alterations are accompanied by bioenergetic defects and the reduction of neuronal activity, which are linked to increased intracellular oxidative stress, and to the activation of the NF-kB/c-Abl/DRP1 axis.

scholarly journals The ALS disease-associated mutant TDP-43 impairs mitochondrial dynamics and function in motor neurons

2013 ◽  
Vol 22 (23) ◽  
pp. 4706-4719 ◽  
Author(s):  
Wenzhang Wang ◽  
Li Li ◽  
Wen-Lang Lin ◽  
Dennis W. Dickson ◽  
Leonard Petrucelli ◽  
...  
Keyword(s):  
Motor Neurons ◽  
Mitochondrial Dynamics ◽  
Dynamics And Function ◽  
And Function

scholarly journals Identification of a Small Molecule That Induces Mitotic Arrest Using a Simplified High-Content Screening Assay and Data Analysis Method

2005 ◽  
Vol 11 (1) ◽  
pp. 21-28 ◽  
Author(s):  
Christopher J. Wilson ◽  
Ying Si ◽  
Craig M. Thompsons ◽  
Andrew Smellie ◽  
Mark A. Ashwell ◽  
...  
Keyword(s):  
Small Molecule ◽  
Active Compound ◽  
Cell Biology ◽  
Mitotic Arrest ◽  
New Drugs ◽  
High Content Screening ◽  
Screening Assay ◽  
Detection Assay ◽  
Phosphohistone H3 ◽  
Small Molecule Modulators

High-content screening has emerged as a newand powerful technique for identifying small-molecule modulators ofmammalian cell biology. The authors describe the development and execution of a high-content screen to identify smallmolecules that induce mitotic arrest in mammalian cancer cells. Many widely used chemotherapeutics, such as Taxol ®and vinblastine, induce mitotic arrest, and the creation of new drugs that also induce mitotic arrest may have tremendous therapeutic value. In their screen, the authors employed a simple DNA stain (DAPI) and a sensitive nonparametric statistical test to identify compounds from an internal collection of 13,000 high-quality lead-like small molecules. Subsequent analysis of 1 active compound indicated that it induces mitotic arrest, assessed using a high-content phosphohistone H3 detection assay, and caused cell proliferation defects inmultiple cancer cell lines. The active compound, a quinazolinone originating from a natural product-like subset of the screened compounds, is active in cells at 500nMand appears to act by inhibiting the polymerization of tubulin.

scholarly journals Turning the tide on Alzheimer’s disease: modulation of γ-secretase

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Joanna E. Luo ◽  
Yue-Ming Li
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Small Molecule ◽  
Molecular Mechanisms ◽  
Neurodegenerative Disorder ◽  
Complex Structure ◽  
Aβ Peptides ◽  
Small Molecule Probes ◽  
Small Molecule Modulators ◽  
And Function

AbstractAlzheimer’s disease (AD) is the most common type of neurodegenerative disorder. Amyloid-beta (Aβ) plaques are integral to the “amyloid hypothesis,” which states that the accumulation of Aβ peptides triggers a cascade of pathological events leading to neurodegeneration and ultimately AD. While the FDA approved aducanumab, the first Aβ-targeted therapy, multiple safe and effective treatments will be needed to target the complex pathologies of AD. γ-Secretase is an intramembrane aspartyl protease that is critical for the generation of Aβ peptides. Activity and specificity of γ-secretase are regulated by both obligatory subunits and modulatory proteins. Due to its complex structure and function and early clinical failures with pan inhibitors, γ-secretase has been a challenging drug target for AD. γ-secretase modulators, however, have dramatically shifted the approach to targeting γ-secretase. Here we review γ-secretase and small molecule modulators, from the initial characterization of a subset of NSAIDs to the most recent clinical candidates. We also discuss the chemical biology of γ-secretase, in which small molecule probes enabled structural and functional insights into γ-secretase before the emergence of high-resolution structural studies. Finally, we discuss the recent crystal structures of γ-secretase, which have provided valuable perspectives on substrate recognition and molecular mechanisms of small molecules. We conclude that modulation of γ-secretase will be part of a new wave of AD therapeutics.

scholarly journals Identification of a small molecule inhibitor that stalls splicing at an early step of spliceosome activation

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Anzhalika Sidarovich ◽  
Cindy L Will ◽  
Maria M Anokhina ◽  
Javier Ceballos ◽  
Sonja Sievers ◽  
...  
Keyword(s):  
Small Molecule ◽  
Intermediate Stage ◽  
Mrna Splicing ◽  
Molecule Inhibitor ◽  
Specific Proteins ◽  
Extensive Exchange ◽  
Dynamics And Function ◽  
Spliceosome Activation ◽  
And Function ◽  
Related Proteins

Small molecule inhibitors of pre-mRNA splicing are important tools for identifying new spliceosome assembly intermediates, allowing a finer dissection of spliceosome dynamics and function. Here, we identified a small molecule that inhibits human pre-mRNA splicing at an intermediate stage during conversion of pre-catalytic spliceosomal B complexes into activated Bact complexes. Characterization of the stalled complexes (designated B028) revealed that U4/U6 snRNP proteins are released during activation before the U6 Lsm and B-specific proteins, and before recruitment and/or stable incorporation of Prp19/CDC5L complex and other Bact complex proteins. The U2/U6 RNA network in B028 complexes differs from that of the Bact complex, consistent with the idea that the catalytic RNA core forms stepwise during the B to Bact transition and is likely stabilized by the Prp19/CDC5L complex and related proteins. Taken together, our data provide new insights into the RNP rearrangements and extensive exchange of proteins that occurs during spliceosome activation.

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