Biology subjects

Charli, A.

Publications and source records attributed to Charli, A..

3 recordsLinked to original sources

bioRxiv2024

Mitochondrial Stress Disassembles Nuclear Architecture through Proteolytic Activation of PKCδ and Lamin B1 Phosphorylation in Neuronal Cells: Implications for Pathogenesis of Age-related Neurodegenerative Diseases

Mitochondrial dysfunction and oxidative stress are hallmarks of pathophysiological processes in age-related neurodegenerative diseases including Parkinsons, Alzheimers and Huntingtons diseases. Neuronal cells are highly vulnerable to mitochondrial stress, however, the cellular and molecular mechanisms underlying the enhanced vulnerability are not well understood. Previously, we demonstrated that the novel PKC isoform PKC{delta} is highly expressed in dopamin(DA)ergic neurons and plays a key role in inducing apoptotic cell death during neurotoxic stress via caspase-3-mediated proteolytic activation. Herein, we further uncovered a key downstream molecular event of PKC{delta} signaling following mitochondrial dysfunction that governs neuronal cell death by dissembling nuclear architecture. Exposing N27 DAergic cell line to the mitochondrial complex-1 inhibitor tebufenpyrad induced PKC{delta} phosphorylation at the T505 activation loop accompanied by caspase-3-dependent proteolytic activation of the kinase. Subcellular analysis using high-resolution 3D confocal microscopy revealed that proteolytically activated cleaved PKC{delta} translocates to the nuclear compartment, colocalizing with Lamin B1. Electron microscopy also enabled the visualization of nuclear membrane damage triggered by subjecting the DAergic neuronal cells by Tebufenpyrad (Tebu) toxicity. In silico analyses identified that the threonine site on Lamin B1 (T575) is likely phosphorylated by PKC{delta}, suggesting that Lamin B1 serves as a key downstream target of the kinase. Interestingly, N27 DAergic cells stably expressing the PKC{delta} proteolytic cleavage site-resistant mutant failed to induce nuclear damage, PKC{delta} activation, and Lamin B1 phosphorylation. Furthermore, CRISPR/Cas9-based stable knockdown of PKC{delta} greatly attenuated Tebu-induced Lamin B1 phosphorylation. Also, studies using Lamin B1T575G mutated at phosphorylation and PKC{delta}-{Delta}NLS-overexpressing N27 cells showed that PKC{delta} activation and translocation to the nuclear membrane are critically required for phosphorylating Lamin B1 at T575 to induce nuclear membrane damage during Tebu insult. Additionally, Tebu failed to induce Lamin B1 damage and Lamin B1 phosphorylation in organotypic midbrain slices cultured from PKC{delta}-/- mouse pups. More importantly, we observed higher PKC{delta} activation, Lamin B1 phosphorylation and Lamin B1 loss in nigral DAergic neurons from the postmortem brains of PD patients as compared to age-matched healthy control brains, thus providing translational relevance of our finding. Collectively, our data reveal that PKC{delta} functions as a Lamin B1 kinase to disassemble the nuclear membrane during the neuronal cell death process triggered by mitochondrial stress. This mechanistic insight may have important implications for the etiology of age-related neurodegenerative diseases resulting from mitochondrial dysfunction as well as for the development of novel treatment strategies.

neuroscience↗

bioRxiv2020

Organic dust induced mitochondrial dysfunction could be targeted via cGAS-STING or mitochondrial NOX-2 inhibition

ABSTRACTOrganic dust (OD) exposure in animal production industries poses serious respiratory and other health risks. OD consists of microbial products and particulate matter and OD exposure induced respiratory inflammation is under intense investigation. However, the effect of OD exposure on brain largely remains unknown. Recently, we have shown that OD exposure of brain microglial cells induces an inflammatory phenotype with the release of mitochondrial DNA (mt-DNA). Therefore, we tested a hypothesis that OD-exposure induced secreted mt-DNA signaling drives the inflammation. OD samples were collected from commercial swine operations and a filter sterilized OD extract (ODE) was prepared. Mouse (C57BL/6) microglial cell line was treated with medium or ODE (5%) for 48 hours along with either PBS or mitoapocynin (MA, 10 μM, NOX-2 inhibitor). Microglia treated with control or anti-STING siRNA were exposed to medium or ODE. Next, mouse (C57BL/6) pups were euthanized under an approved protocol, organotypic brain slice cultures (BSCs) were prepared and exposed to medium or ODE with or without MA treatment daily for five days. Culture supernatant, cell pellets and mt-free cytosolic fractions were processed to quantify mt-superoxide, mt-DNA, cytochrome C, TFAM, mitochondrial stress markers and mt-DNA induced signaling via cGAS-STING and TLR9. Data were analyzed using one-way ANOVA and post-hoc tests. A p value of ≤ 0.05 was considered significant. ODE exposure increased the mt-superoxide formation andMA treatment decreased the ODE-induced mt-DNA release into cytosol. ODE exposure increased the cytochrome C and TFAM levels. ODE increased MFN1/2 and PINK1 but not DRP1 and MA treatment decreased the MFN2 expression. MA treatment decreased the ODE-exposure induced mt-DNA signaling via cGAS-STING and TLR9. Anti-STING siRNA decreased the ODE-induced increase in IRF3, IFN-β and Iba1 expression. In BSCs, MA-treatment decreased the ODE induced TNF-α, IL-6 and MFN1. Taken together, OD exposure induced mt-DNA signaling could be curtailed through mitochondrial NOX-2 inhibition or STING suppression to reduce neuroinflammation.Competing Interest StatementAGK has an equity interest in PK Biosciences Corporation located in Ames, IA. The terms of this arrangement have been reviewed and approved by Iowa State University per its conflict of interest policies. All other authors have declared no potential conflicts of interest.View Full Text

cell biology↗

bioRxiv2019

Mitochondrial Dysfunction Induces Epigenetic Dysregulation by H3K27 Hyperacetylation to Perturb Active Enhancers in Parkinson’s Disease Models

Genetic mutations explain only 10-15% of cases of Parkinsons disease (PD), while an overriding environmental component has been implicated in the etiopathogenesis of PD. But regardless of where the underlying triggers for the onset of familial and sporadic PD fall on the gene-environment axis, mitochondrial dysfunction emerges as a common mediator of dopaminergic neuronal degeneration. Herein, we employ a multidisciplinary approach to convincingly demonstrate that neurotoxicant exposure- and genetic mutation-driven mitochondrial dysfunction share a common mechanism of epigenetic dysregulation. Under both scenarios, lysine 27 acetylation of likely variant H3.2 (H3.2K27ac) increased in dopaminergic neuronal models of PD, thereby opening that region to active enhancer activity via H3K27 hyperacetylation. These vulnerable epigenomic loci represent potential transcription factor motifs for PD pathogenesis. We further confirmed the mitochondrial dysfunction induced H3K27ac during neurodegeneration in ex vivo models of PD. Our results reveal an exciting axis of exposure/mutation-mitochondrial dysfunction-metabolism-H3K27ac-transcriptome for PD pathogenesis. Collectively, the novel mechanistic insights presented here interlinks mitochondrial dysfunction to epigenetic transcriptional regulation in dopaminergic degeneration as well as offer potential new epigenetic intervention strategies for PD.

neuroscience↗