Biology subjects

Mobley, J.

Publications and source records attributed to Mobley, J..

7 recordsLinked to original sources

bioRxiv2026

Vitamin B12 Improves Skeletal Muscle Mitochondrial Biology in Aged Mice

Age-related skeletal muscle deterioration is a commonly reported disability among older adults, attributed to several factors including mitochondrial dysfunction, a major hallmark of aging. Therapies to attenuate or reverse mitochondrial decline are limited. Despite identified positive relationships between vitamin B12 (B12) and mitochondrial biology, the impact of B12 supplementation on skeletal muscle mitochondria, in advanced aged, has not been examined. Thus, the impact of B12 supplementation on skeletal muscle mitochondrial biology was examined in (i) aged female mice, given 12 weeks of B12 supplementation (SUPP) or vehicle control, and (ii) in human primary myotubes. In the mouse model, mitochondrial DNA and content were measured with PCR and citrate synthase activity, respectively; mitochondrial morphology was examined using transmission electron microscopy; mitochondrial function was examined using extracellular metabolic flux analysis; and proteins and pathway enrichment was identified with proteomics. In the cell model, ROS and glutathione was measured using luminescent assays. The results demonstrated that SUPP in aged mice increased muscle mitochondrial content and improved morphology. Further, differentially expressed proteins were enriched in TCA cycle, OXPHOS, and oxidative stress pathways. In the cell model, B12 supplementation reduced ROS levels. This is the first study, to our knowledge, examining the impact of B12 supplementation on skeletal muscle mitochondrial biology in aged female mice. Results suggest that B12 supplementation improves mitochondrial biology in aged female mice.

molecular biology↗

bioRxiv2025

Glutamine deficiency enhances nuclear localization of TCA cycle enzymes and epigenetic modifications, impairing myogenesis

ObjectiveSkeletal muscle myogenesis, during development and regeneration, are primarily facilitated by the progeny of resident muscle stem cells, the muscle progenitor cells (MPCs). Intriguingly, extracellular glutamine (Gln) is required by MPCs. Disruptions in Gln availability or metabolism, impair MPC function and hinder skeletal muscle regeneration. Gln, a nutritionally non-essential amino acid (NEAA), which is most abundant in skeletal muscle, becomes conditionally essential during advancing age and catabolic states like sepsis, trauma, and strenuous situations. The intracellular mechanisms through which extracellular Gln availability affects MPC proliferation are poorly understood. Therefore, this study was aimed to examine how extracellular supply of Gln influences myogenesis through the spatial distribution of TCA-cycle enzymes. MethodsHuman primary muscle myoblasts (HSKM2) and murine MPCs (C2C12) were utilized for in-vitro experiments. To investigate the impact of Gln availability on MPC proliferation, proliferation assays, confocal imaging, succinyl-proteomics, and single-cell nuclei ATAC sequencing were used. ResultsGln (2 mM) significantly enhanced MPC viability and cell cycle progression, while Gln depletion (0 mM) reduced cell number and induced G0/G1 arrest. Gln deficiency downregulated MyoD expression, impairing myogenic potential. Inhibiting intracellular Gln metabolism also limited proliferation, indicating the necessity of Gln catabolism for cell proliferation. Gln deficiency increased nuclear localization of TCA cycle enzymes (DLST, OGDH), elevated histone succinylation, and restricted chromatin accessibility of the myogenic regulatory factor, MyoD1, highlighting a Gln dependent metabolic-epigenetic nexus in myogenic regulation. ConclusionThese findings demonstrate that extracellular Gln availability is a key regulator of myogenesis through its influence on metabolism, epigenetic modifications, and chromatin accessibility.

developmental biology↗

bioRxiv2024

Glucose-Sensing ChREBP Protein in the Pathogenesis of Diabetic Retinopathy

Glucose-sensing ChREBP and MondoA are transcriptional factors involved in lipogenic, inflammatory, and insulin signaling pathways implicated in metabolic disorders; however, limited ocular studies have been conducted on these proteins. We aimed to investigate the potential role of ChREBP in pathogenesis of diabetic retinopathy (DR). We used diabetic human and mouse retinal cryosections analyzed by immunohistochemistry. qRT-PCR was performed to quantify gene expression. To explore the role of ChREBP in rods, we generated caChREBPRP mice with constitutively active (ca) ChREBP. These mice underwent retinal function testing, followed by proteomic analysis using LC-MS. Furthermore, ARPE-19 cells were infected with lentiviral particles expressing human ChREBP (ARPE-19ChREBP) and subjected to global proteomics. Our results demonstrate that both pro-teins were expressed across the retina, although with distinct distribution patterns: MondoA was more prominently expressed in cones, while ChREBP was broadly expressed throughout the retina. Elevated expression of both proteins was observed in DR. This may have contributed to rod photo-receptor degeneration as we observed diminished scotopic ERG amplitudes detected in caChREB-PRP mice at P35. The retinal proteomic landscape indicated a decline in KEGG pathways associated with phototransduction, amino acid metabolism, and cell adhesion. Furthermore, rod-specific ca-ChREBP induced TXNIP expression. Consistent with altered retinal proteomics, ARPE-19ChREBP cells displayed a metabolic shift toward increased glyoxylate signaling, sugar metabolism, and lysosomal activation. Our study demonstrates that ChREBP overexpression causes significant metabolic reprograming triggering retinal functional loss in mice

cell biology↗

bioRxiv2024

Oxidative Injury to Lung Mitochondrial DNA is a Key Contributorfor the Development of Chemical Lung Injury

The mechanisms and extent to which inhalation of oxidant gases damage the mitochondrial genome contributing to the development of acute and chronic lung injury have not been investigated. C57BL/6 mice exposed to chlorine (Cl2) gas and returned to room air, developed progressive loss of lung DNA glycosylase OGG1, significant oxidative injury to mtDNA, decreased intact lung mitochondrial (mt) DNA, generation of inflammatory pathway by DAMPs causing airway and alveolar injury with significant mortality. Global proteomics identified over 1400 lung proteins with alteration of key mitochondrial proteins at 24 h post Cl2 exposure. Intranasal instillation of a recombinant protein containing mitochondrial targeted OGG1 (mitoOGG1) post exposure, decreased oxidative injury to mtDNA, lung mitochondrial proteome, severity of the acute and chronic lung injury and increased survival. These data show that injury to the mt-genome is a key contributor to the development of acute and chronic lung injury.

physiology↗

bioRxiv2024

Role of histone variants H2BC1 and H2AZ.2 in H2AK119ub nucleosome organization and Polycomb gene silencing

Ubiquitination of histone H2A at lysine 119 residue (H2AK119ub) plays critical roles in a wide range of physiological processes, including Polycomb gene silencing 1,2, replication 3-5, DNA damage repair 6-10, X inactivation 11,12, and heterochromatin organization 13,14. However, the underlying mechanism and structural basis of H2AK119ub remains largely elusive. In this study, we report that H2AK119ub nucleosomes have a unique composition, containing histone variants H2BC1 and H2AZ.2, and importantly, this composition is required for H2AK119ub and Polycomb gene silencing. Using the UAB domain of RSF1, we purified H2AK119ub nucleosomes to a sufficient amount and purity. Mass spectrometry analyses revealed that H2AK119ub nucleosomes contain the histone variants H2BC1 and H2AZ.2. A cryo-EM study resolved the structure of native H2AK119ub nucleosomes to a 2.6A resolution, confirming H2BC1 in one subgroup of H2AK119ub nucleosomes. Tandem GST-UAB pulldown, Flag-H2AZ.2, and HA-H2BC1 immunoprecipitation revealed that H2AK119ub nucleosomes could be separated into distinct subgroups, suggesting their composition heterogeneity and potential dynamic organization. Knockout or knockdown of H2BC1 or H2AZ.2 reduced cellular H2AK119ub levels, establishing H2BC1 and H2AZ.2 as critical determinants of H2AK119ub. Furthermore, genomic binding profiles of H2BC1 and H2AZ.2 overlapped significantly with H2AK119ub binding, with the most significant overlapping in the gene body and intergenic regions. Finally, assays in developing embryos reveal an interaction of H2AZ.2, H2BC1, and RING1A in vivo. Thus, this study revealed, for the first time, that the H2AK119ub nucleosome has a unique composition, and this composition is required for H2AK119ub and Polycomb gene silencing.

molecular biology↗

bioRxiv2023

Alloreactivity and autoreactivity converge to support B cell epitope targeting in transplant rejection

Donor-specific antibody (DSA) responses against human leukocyte antigen (HLA) proteins mismatched between kidney transplant donors and recipients cause allograft loss. The rules governing the immunogenicity of non-self donor HLA are poorly understood. Using single-cell, molecular, structural, and proteomic techniques, we profiled the HLA-specific B cell response in the kidney and blood of a transplant recipient with antibody-mediated rejection (AMR). We observed an immunodominant B cell antibody response focused on topographically exposed, solvent-accessible mismatched HLA residues along the peptide-binding groove - a subregion comprising only 20% of the HLA molecule. We further demonstrated that, even within a diverse cohort of transplant recipients, the B cell alloresponse consistently converges on this same immunodominant subregion on the crown of the HLA molecule. Based on these findings, we propose that B cell immunodominance in transplant rejection relies on antigenic topography, and we suggest that this link could be exploited for organ matching and therapeutics.

immunology↗

bioRxiv2020

Heme Impairs Alveolar Epithelial Sodium Channels Post Toxic Gas Inhalation

We previously reported that cell-free heme (CFH) is increased in the plasma of patients with acute and chronic lung injury and causes pulmonary edema in animal model of acute respiratory distress syndrome (ARDS) post inhalation of halogen gas. However, the mechanisms by which CFH causes pulmonary edema are unclear. Herein we report for the first time the presence of CFH and chlorinated lipids (formed by the interaction of halogen gas, Cl2, with plasmalogens) in the plasma of patients and mice exposed to Cl2 gas. Ex vivo incubation of red blood cells (RBC) with halogenated lipids caused oxidative damage to RBC cytoskeletal protein spectrin, resulting in hemolysis and release of CFH. A single intramuscular injection of the heme-scavenging protein hemopexin (4 {micro}g/kg body weight) in mice, one hour post halogen exposure, reversed RBC fragility and decreased CFH levels to those of air controls. Patch clamp and short circuit current measurements revealed that CFH inhibited the activity of amiloride-sensitive (ENaC) and cation sodium (Na+) channels in mouse alveolar cells and trans-epithelial Na+ transport across human airway cells with EC50 of 125 nM and 500 nM, respectively. Molecular modeling identified 22 putative heme-docking sites on ENaC (energy of binding range: 86-1563 kJ/mol) with at least 2 sites within its narrow transmembrane pore, potentially capable of blocking Na+ transport across the channel. In conclusion, results suggested that CFH mediated inhibition of ENaC activity may be responsible for pulmonary edema post inhalation injury.

physiology↗