Biology subjects

Seaborne, R. A. E.

Publications and source records attributed to Seaborne, R. A. E..

5 recordsLinked to original sources

bioRxiv2026

The ribosomal DNA landscape of mammalian muscle during acute and chronic physiological stress

Ribosomal DNA (rDNA) is a highly repetitive and complex locus within the mammalian genome that exhibits substantial inter-individual variation in number of rDNA copies and epigenetic regulation. Nonetheless, our understanding of rDNA biology in skeletal muscle during periods of physiological stress is limited. Using publicly available whole genome and reduced representative bisulfite sequencing data sets, we identify a concurrent reduction in both the number of rDNA copies and the methylation profile of the rDNA in aged vs young mice, supported by large effect sizes and permutation testing, with significant reductions in methylation of the 18S coding unit in aged, compared to young controls (p = 0.024). We found a strong positive correlation between rDNA copy number and 18S methylation across both young and aged mice (p = 0.004; Spearman rho = 0.842). After analysing publicly available muscle (skeletal and cardiac) data sets following acute insult (endurance exercise, cancer cachexia, spinal cord injury), we do not observe a similarly coordinated epi-genetic modification in rDNA biology but uncover tissue and sex-specific differences in rDNA copy number or methylation status, in isolation. These findings suggest ageing as a unique physiological insult in which coordinated epi-genomic remodelling of the rDNA region appears, representing a previously underappreciated feature of the muscle ageing trajectory.

physiology↗

bioRxiv2026

Integrated Single-Fiber Multi-Omics Links an Inflammatory-Associated Myofiber State to Altered Myosin Dynamics in Patients with ICU-acquired weakness

Skeletal muscle dysfunction is a pervasive complication of critical illness that worsens survival and recovery, yet remains poorly explained by current clinical or molecular markers. To directly connect disease-associated molecular states to the contractile machinery, this study combined sequential functional, transcriptomic, and proteomic profiling of the same single human skeletal myofibers from critically ill patients in the intensive care unit with acquired weakness (ICU-AW) and controls. Despite marked donor-level heterogeneity, integrated analysis revealed a subtle yet conserved myofiber state enriched in ICU-AW, characterized by inflammatory and chemotactic gene programs, intracellular structural remodeling, and bioenergetic adaptation. Nineteen features were significantly altered at both RNA and protein levels from the same myofiber, linking an inflammatory transcriptional landscape to a proteomic shift toward mitochondrial and translational machinery and away from membrane-associated signaling. Functionally, fibers in this state displayed selectively disrupted myosin dynamics, evidenced by prolonged ATP turnover time of myosin heads in their super-relaxed conformation, implicating altered myosin energetics as a contributor to muscle dysfunction. These findings define a discrete, disease-associated myofiber state and establish an integrative single-fiber framework for connecting multi-omic heterogeneity to molecular motor function in complex human disease. Graphical AbstractSingle-fiber multi-omic and functional analysis reveals a stress-adapted myofiber state in ICU-AW. Specifically, for the present study, myofibers from ICU-AW donors and control donors were isolated and functionally profiled for myosin dynamics before being split for simultaneous transcriptomic and proteomic analysis. Integrated analysis then identified a reproducible fiber phenotype enriched in ICU-AW, characterized by inflammatory transcriptional signatures coordinated with mitochondrial proteomic remodeling and altered myosin super-relaxed state energetics. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/706099v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@95715forg.highwire.dtl.DTLVardef@1462ff8org.highwire.dtl.DTLVardef@f74996org.highwire.dtl.DTLVardef@10055ec_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2024

Integrated single cell functional-proteomic profiling of human skeletal muscle reveals a shift in cellular specificity in nemaline myopathy

Skeletal muscle is a complex syncytial arrangement of an array of cell types and, in the case of muscle specific cells (myofibers), sub-types. There exists extensive heterogeneity in skeletal muscle functional behaviour and molecular landscape, at the cell composition, myofiber sub-type and intra-myofiber sub-type level. This heterogeneity highlights limitations in currently applied methodological approaches, which has stagnated our understanding of fundamental skeletal muscle biology in both healthy and myopathic contexts. Here, we developed a novel approach that combines a fluorescence based assay for the biophysical examination of the sarcomeric protein, myosin, coupled with same-myofiber high sensitivity proteome profiling, termed Single Myofiber Protein Function-Omics (SMPFO). Successfully applying this approach to healthy human skeletal muscle tissue, we identify the integrate relationship between myofiber functionality and the underlying proteomic landscape that guides divergent, but physiologically important, behaviour in myofiber sub-types. By applying SMPFO to two forms of human nemaline myopathy (ACTA1 and TNNT1 mutations), we reveal significant reduction in the divergence of myofiber sub-types, across both biophysical and proteomic behaviour. Collectively, we develop SMPFO as a novel approach to study skeletal muscle with greater specificity, accuracy and resolution then currently applied methods, facilitating that advancement in understanding of SkM tissue in both healthy and diseased states.

physiology↗

bioRxiv2024

Dysregulated Skeletal Muscle Myosin Super-relaxation in Type II, but Not Type I, Diabetes Mellitus

Disrupted energy balance is critical for the onset and development of Type II diabetes. The exact underlying metabolic mechanisms remain incomplete but skeletal muscle is thought to play an important pathogenic role. As the super-relaxed state of its most abundant protein, myosin, regulates cellular energetics, here, we aimed to investigate whether it is altered in patients with type II diabetes. For that, we used vastus lateralis biopsy specimens (obtained from patients with type II diabetes and matched controls) and run a combination of structural and functional assays consisting of loaded Mant-ATP chase experiments, X-ray diffraction and LC-MS/MS proteomics in isolated muscle fibres. Our studies revealed a greater muscle myosin super-relaxation and decreased cellular ATP demand in patients than controls. Subsequent proteomic analyses indicated that these (mal)adaptations likely originated from remodeled sarcomeric proteins and greater myosin glycation levels in patients than controls. Overall, our findings emphasize a complex molecular dysregulation of myosin super-relaxed state and energy consumption in type II diabetes. Ultimately, pharmacological targeting of myosin could benefit skeletal muscle and whole-body metabolic health through the enhancement of ATP consumption. Significance StatementMyosin super-relaxation, essential for the regulation of skeletal muscle metabolic rate, is disrupted in type II diabetes due to protein hyper-glycation. As a consequence, myosin ATP demand is significantly lowered. Overall, our findings provide a strong rationale for the use of activators of myosin ATPase to enhance basal energy expenditure in type II diabetes.

cell biology↗

bioRxiv2023

Remodelling of Skeletal Muscle Myosin Metabolic States in Hibernating Mammals

Hibernation is a period of metabolic suppression utilized by many small and large mammal species to survive during winter periods. As the underlying cellular and molecular mechanisms remain incompletely understood, our study aimed to determine whether skeletal muscle myosin and its metabolic efficiency undergo alterations during hibernation to optimize energy utilization. We isolated muscle fibers from small hibernators, Ictidomys tridecemlineatus and Eliomys quercinus and larger hibernators, Ursus arctos and Ursus americanus. We then conducted loaded Mant-ATP chase experiments alongside X-ray diffraction to measure resting myosin dynamics and its ATP demand. In parallel, we performed multiple proteomics analyses. Our results showed a preservation of myosin structure in U. arctos and U. americanus during hibernation, whilst in I. tridecemlineatus and E. quercinus, changes in myosin metabolic states during torpor unexpectedly led to higher levels in energy expenditure of type II, fast-twitch muscle fibers at ambient lab temperatures (20{degrees}C). Upon repeating loaded Mant-ATP chase experiments at 8{degrees}C (near the body temperature of torpid animals), we found that myosin ATP consumption in type II muscle fibers was reduced by 77-107% during torpor compared to active periods. Additionally, we observed Myh2 hyper-phosphorylation during torpor in I. tridecemilineatus, which was predicted to stabilize the myosin molecule. This may act as a potential molecular mechanism mitigating myosin-associated increases in skeletal muscle energy expenditure during periods of torpor in response to cold exposure. Altogether, we demonstrate that resting myosin is altered in hibernating mammals, contributing to significant changes to the ATP consumption of skeletal muscle. Additionally, we observe that it is further altered in response to cold exposure and highlight myosin as a potentially contributor to skeletal muscle non-shivering thermogenesis.

physiology↗