Biology subjects

Thalacker-Mercer, A. E.

Publications and source records attributed to Thalacker-Mercer, A. E..

10 recordsLinked to original sources

bioRxiv2026

Integrative Multi-omics Analysis of the Human Skeletal Muscle Response to Endurance or Resistance Exercise: Findings from the Molecular Transducers of Physical Activity Consortium (MoTrPAC)

The Molecular Transducers of Physical Activity Consortium (MoTrPAC) was established to systematically characterize the molecular basis of the health benefits of exercise. Here, we present the integrative, multi-omics response of human skeletal muscle to acute endurance (EE) and resistance (RE) exercise. Distinct temporal responses were observed, with changes in ATAC-seq, phosphoproteome, and metabolome occurring before changes in the transcriptome and proteome. These distinct temporal multi-omic dynamics were used to identify transcriptional regulatory hubs converging around MEF2A and NFIC regulation of autophagy, angiogenesis and metabolism. Further, early RE-specific phosphoproteome signatures counteracted epigenetic modifications and downregulated transcripts involved in protein turnover. Additional findings include suppression of HIPK2/3 kinase signatures linked to the acute exercise regulation of sarcomeric proteins TTN, NEB, ANKRD2 and LMOD2. Our data demonstrate distinct temporal regulation across the multi-omic landscape of human skeletal muscle, with EE and RE eliciting common and unique molecular signatures.

systems biology↗

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

DOCK7 regulates WNT signaling and NMJ clustering in normal and DMD muscle

Dedicator of cytokinesis (DOCK) proteins have diverse and critical roles in myoblast fusion, glucose metabolism, skeletal muscle regeneration, and neuronal polarity. DOCK7 is part of this 11-member family of atypical guanine exchange factors. While human DOCK7 pathogenic variants are rare, biallelic variants are known to result in hypotonia and ataxia in children. More recently, DOCK variants have been shown to impact disease pathogenesis as secondary modifiers. We used tissue-specific Dock7 conditional knockout mice to evaluate the roles of DOCK7 in the skeletal muscles and motor neurons. Both Dock7 knockout models developed significant nerve and muscle function deficits; however, the Dock7 motor neuron knockout mice showed more severe locomotor pathologies. RNA sequencing of the Dock7 knockout skeletal muscles revealed dysregulation of WNT signaling factors, and skeletal muscle repair and regeneration pathways. Molecular analysis revealed impaired DOCK7-mediated RAC1 activation that impacted acetylcholine receptor (AChR) clustering in both models. Interestingly, haploinsufficiency of Dock7 in mdx mice improved overall dystrophic muscle pathologies. We propose that DOCK7-RAC1 interactions result in activation of the WNT signaling pathway via the LRP4 receptor in the skeletal muscle and facilitate post-synaptic AChR clustering. Graphical AbstractDOCK7 is a regulator of normal and DMD muscle via the RAC1/WNT pathway. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=124 SRC="FIGDIR/small/696254v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@1ce84eaorg.highwire.dtl.DTLVardef@17855f1org.highwire.dtl.DTLVardef@15263d2org.highwire.dtl.DTLVardef@1778e0_HPS_FORMAT_FIGEXP M_FIG C_FIG

genetics↗

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↗

bioRxiv2025

Vitamin B12 supports skeletal muscle oxidative phosphorylation capacity in male mice

ObjectivesVitamin B12 plays a vital role in folate-mediated one-carbon metabolism (FOCM), a series of one-carbon transfer reactions that generate nucleotides (thymidylate (dTMP) and purines) and methionine. Inadequate levels of B12 impair FOCM, depressing de novo thymidylate (dTMP) synthesis, which in turn leads to uracil accumulation in DNA. This phenomenon has been well documented in nuclear DNA. Our previous work in liver tissue has shown that mitochondrial DNA (mtDNA) is more sensitive to FOCM impairments in that mtDNA exhibits elevated uracil levels before uracil concentrations in nuclear DNA change. However, the functional consequences of uracil accumulation in mtDNA are largely unknown. The purpose of this study was to determine how a functional B12 deficiency (induced by reduced levels of the B12-dependent enzyme methionine synthase (MTR)) and dietary B12 deficiency affects mtDNA integrity and mitochondrial function in energetic and mitochondria-rich tissues such as skeletal muscle. MethodsMale Mtr+/+ and Mtr+/- mice were weaned to either an AIN93G-based control (C) diet containing 25 {micro}g/kg vitamin B12 or a B12-deficient (-B12) diet containing 0 {micro}g/kg vitamin B12 to explore the effects of functional (Mtr+/-) and dietary B12 deficiency on muscle weight, uracil content in mtDNA, mtDNA content, and oxidative phosphorylation complex capacity in skeletal muscle. Aged (20-22mo) male C57BL6/N mice were acclimated to an AIN93G control diet four weeks, then received either weekly injections of saline (vehicle control [30 uL 0.9% NaCl]) or B12 (0.65mg per 30uL 0.9% NaCl) in each of two hindleg muscles [1.25 mg B12 total]) for 8 weeks. ResultsThe tibialis anterior (TA) muscle from Mtr+/-mice exhibited lowered maximal respiratory capacity of complex I, II, and IV of the electron transport chain than did TA from Mtr+/+ mice. Exposure to the -B12 diet lowered maximal capacity of complex I in red, mitochondrially rich muscle (soleus and mitochondria-rich portions of quadriceps and gastrocnemius) (p=0.02). Levels of uracil accumulation in mtDNA in red muscle and gastrocnemius were elevated [~]10 fold with exposure to -B12 diet (p=0.04 and p<0.001, respectively). In aged mice gastrocnemius complex IV activity increased with intramuscular B12 supplementation (p=0.04) ConclusionsExposure to a B12-deficient diet led to uracil accumulation in mtDNA and impaired maximal oxidative capacity in two different types of skeletal muscle. B12 supplementation improved complex IV maximal capacity in gastrocnemius from aged mice.

biochemistry↗

bioRxiv2022

Impairments in SHMT2 expression or cellular folate availability reduce oxidative phosphorylation and pyruvate kinase activity

BackgroundSerine hydroxymethyltransferase 2 (SHMT2) catalyzes the reversible conversion of tetrahydrofolate (THF) and serine producing THF-conjugated one-carbon units and glycine in the mitochondria. Biallelic SHMT2 variants were identified in humans and suggested to alter the proteins active site, potentially disrupting enzymatic function. SHMT2 expression has also been shown to decrease with aging in human fibroblasts. Immortalized cells models of total SHMT2 loss or folate deficiency exhibit decreased oxidative capacity and impaired mitochondrial complex I assembly and protein levels, suggesting folate-mediated one-carbon metabolism (FOCM) and the oxidative phosphorylation system are functionally coordinated. This study examined the role of SHMT2 and folate availability in regulating mitochondrial function, energy metabolism, and cellular proliferative capacity in both heterozygous and homozygous cell models of reduced SHMT2 expression. MethodsPrimary mouse embryonic fibroblasts (MEF) were isolated from a C57Bl/6 dam crossed with a heterozygous Shmt2+/- male to generate Shmt2+/+ (wild-type) or Shmt2+/- (HET) MEF cells. In addition, haploid chronic myeloid leukemia cells (HAP1, wild-type) or HAP1 cells lacking SHMT2 expression ({Delta}SHMT2) were cultured for 4 doublings in either low-folate or folate-sufficient culture media. Cells were examined for proliferation, total folate levels, mtDNA content, protein levels of pyruvate kinase and PGC1, pyruvate kinase enzyme activity, mitochondrial membrane potential, and mitochondrial function. ResultsHomozygous loss of SHMT2 in HAP1 cells impaired cellular folate accumulation and altered mitochondrial DNA content, membrane potential, and basal respiration. Formate rescued proliferation in HAP1, but not {Delta}SHMT2, cells cultured in low-folate medium. Pyruvate kinase activity and protein levels were impaired in {Delta}SHMT2 cells and in MEF cells exposed to low-folate medium. Mitochondrial biogenesis protein levels were elevated in Shmt2+/- MEF cells, while mitochondrial mass was increased in both homozygous and heterozygous models of SHMT2 loss. ConclusionsThe results from this study indicate disrupted mitochondrial FOCM impairs mitochondrial folate accumulation and respiration, glycolytic activity, and cellular proliferation. These changes persist even after a potentially compensatory increase in mitochondrial biogenesis as a result of decreased SHMT2 levels.

biochemistry↗

bioRxiv2021

Reduced Shmt2 expression impairs mitochondrial folate accumulation and respiration, and leads to uracil accumulation in mouse mitochondrial DNA

BackgroundAdequate cellular thymidylate (dTMP) pools are essential for preservation of nuclear and mitochondrial genome stability. Previous studies have indicated that disruption in dTMP synthesis in the nucleus leads to increased uracil misincorporation into DNA affecting genome stability. To date, the effects of impaired mitochondrial dTMP synthesis in non- transformed tissues have been understudied. ObjectiveThis study aimed to determine the effects of decreased serine hydroxymethyltransferase 2 (Shmt2) expression and dietary folate deficiency on mitochondrial DNA integrity and mitochondrial function in mouse tissues. MethodsLiver mitochondrial DNA (mtDNA) content, and uracil content in liver mtDNA was measured in Shmt2+/- and Shmt2+/+ mice weaned onto either a folate-sufficient control diet (2 mg/kg folic acid, C) or a modified diet lacking folic acid (0 mg/kg folic acid, FD) for 7 wks. Shmt2+/- and Shmt2+/+ mouse embryonic fibroblasts (MEF cells) were cultured in defined culture medium containing either 0 or 25 nM folate to assess proliferative capacity and mitochondrial function. ResultsShmt2+/- mice exhibited 48-67% reduction in SHMT2 protein levels in tissues. Interestingly, Shmt2+/- mice consuming the folate-sufficient C diet exhibited a 25% reduction in total folate in liver mitochondria. There was also a >20-fold increase in uracil in liver mtDNA in Shmt2+/- mice consuming the C diet, and dietary folate deficiency also increased uracil content in mouse liver mtDNA from both Shmt2+/+ and Shmt2+/- mice. Furthermore, decreased Shmt2 expression in MEF cells reduced cell proliferation, mitochondrial membrane potential, and oxygen consumption rate. ConclusionsThis study demonstrates that Shmt2 heterozygosity and dietary folate deficiency impair mitochondrial dTMP synthesis, as evidenced by the increased uracil in mtDNA. In addition, Shmt2 heterozygosity impairs mitochondrial function in MEF cells. These findings suggest that elevated uracil in mtDNA may impair mitochondrial function.

biochemistry↗

bioRxiv2021

NaCT (SLC13A5) facilitates citrate import and metabolism under nutrient-limited conditions

Citrate lies at a critical node of metabolism linking tricarboxylic acid metabolism and fatty acid synthesis via acetyl-coenzyme A. Recent studies have linked the sodium citrate transporter (NaCT), encoded by SLC13A5, to dysregulated hepatic metabolism and pediatric epilepsy. To examine how NaCT-mediated citrate metabolism contributes to the pathophysiology of these diseases we applied 13C isotope tracing to SLC13A5-deficient hepatocellular carcinoma (HCC) cell lines and primary rat cortical neurons. Exogenous citrate contributed to intermediary metabolism at appreciable levels only under hypoxic conditions. In the absence of glutamine, citrate supplementation increased de novo lipogenesis and growth of HCC cells. Knockout of SLC13A5 in Huh7 cells compromised citrate uptake and catabolism. Citrate supplementation rescued Huh7 cell viability in response to glutamine deprivation and Zn2+ treatment, and these effects were mitigated by NaCT deficiency. Collectively, these findings demonstrate that NaCT-mediated citrate uptake is metabolically important under nutrient limited conditions and may facilitate resistance to metal toxicity.

cell biology↗

bioRxiv2020

Lifelong Ulk1-mediated autophagy deficiency in muscle induces mitochondrial dysfunction and contractile weakness

The accumulation of damaged mitochondria due to insufficient autophagy has been implicated in the pathophysiology of sarcopenia resulting in reduced contractile and metabolic function. Ulk1 is an autophagy-related kinase that initiates autophagosome assembly and may also play a role in autophagosome degradation (i.e., autophagy flux), but the contribution of Ulk1 to healthy muscle aging is unclear. We found that Ulk1 phosphorylation declines in both human and mouse muscle tissue with age, therefore the purpose of this study was to investigate the role of Ulk1-mediated autophagy in skeletal muscle aging. At age 22 months (80% survival rate), muscle contractile and metabolic function were assessed using electrophysiology in muscle specific Ulk1 knockout mice (MKO) and their littermate controls (LM). Specific peak-isometric torque of the ankle dorsiflexors (normalized by tibialis anterior muscle cross-sectional area) and specific force of the fast-twitch extensor digitorum longus muscles were reduced in MKO mice compared to LM mice (p<0.03). Permeabilized muscle fibers from MKO mice had greater mitochondrial content, yet lower mitochondrial oxygen consumption and greater reactive oxygen species production compared to fibers from LM mice (p[≤] 0.04). Altered neuromuscular junction innervation patterns and changes in autophagosome numbers and/or flux in muscles from MKO may have contributed to decrements in contractile and metabolic function. Results from this study support an important role of Ulk1-mediated autophagy in skeletal muscle with age, reflecting Ulk1s dual role in maintaining mitochondrial integrity through autophagosome assembly and degradation. A lifetime of insufficient Ulk-1-mediated autophagy in skeletal muscle exacerbates age-related contractile and metabolic dysfunction.

physiology↗

bioRxiv2019

Serine and glycine are essential for human muscle progenitor cell population expansion

Skeletal muscle regeneration is reliant on a population of muscle specific adult stem cells (muscle progenitor cells; MPCs). During regeneration, the MPC population undergoes a transient and rapid period of population expansion, which is necessary to repair damaged myofibers and restore muscle homeostasis. Much research has focused on the age-related accumulation of negative regulators of regeneration, while the age-related decline of nutrient and metabolic determinants of the regenerative process needs examination. We hypothesized that older individuals, a population that is at risk for protein malnutrition, have diminished availability of amino acids that are necessary for MPC function. Here, we identified that levels of the non-essential amino acid serine are reduced in the skeletal muscle of healthy, older individuals. Furthermore, using stable-isotope tracing studies, we demonstrate that primary, human MPCs (hMPCs) exhibit a limited capacity for de novo biosynthesis of serine and the closely related amino acid glycine. We identified that serine and glycine are essential for hMPC proliferation and, therefore, population expansion. Serine and glycine were necessary to support synthesis of the intracellular antioxidant glutathione, and restriction of serine and glycine was sensed in an EIF2-dependent manner resulting in cell cycle arrest in G0/G1. In conclusion, we elucidate that, despite an absolute requirement of serine/glycine for hMPC proliferation, availability of serine in the skeletal muscle microenvironment is limited to the hMPCs of healthy older adults and is a likely underlying mechanism for impaired skeletal muscle regeneration with advancing age.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=176 SRC=\"FIGDIR/small/833798v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (32K):\norg.highwire.dtl.DTLVardef@438298org.highwire.dtl.DTLVardef@32f66dorg.highwire.dtl.DTLVardef@1d08aeeorg.highwire.dtl.DTLVardef@511c7f_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

cell biology↗