Mitochondrion

We show that chronic impairment of mitochondrial respiration is associated with marked accumulation of cytochrome c (Cytc) protein. Using SCO2-deficient HCT116 cells lacking functional cytochrome c oxidase and wild-type cells exposed to sustained hypoxia, we found that substantial mitochondrial Cytc accumulation parallels reduced electron flux through Cytc. SCO2-deficient cells exhibited equally elevated Cytc levels under normoxia (19% O2) and hypoxia (0.1-3% O2). Wild-type cells under sustained hypoxia accumulated Cytc, reaching levels comparable to those in SCO2-deficient cells. This effect was reversible upon reoxygenation. Increased Cytc protein levels were also observed in other cell models, including primary cortical neurons cultured under chronic hypoxia and in cerebral cortex tissue from hypoxia-exposed mice. Cytc accumulation occurred independently of CYCS transcription, mRNA translation, HIF activation, ROS production and changes in mitochondrial network. Pharmacological inhibition of complex III was likewise accompanied by increased Cytc levels, whereas mitochondrial uncoupling had no effect, suggesting that impaired electron transfer rather than membrane depolarisation per se underlies this association. Raman spectroscopy revealed enrichment of reduced Cytc and an increased Cytc-to-cytochrome b ratio in respiration-deficient cells. Further supporting a stabilisation-based mechanism, the fraction of membrane-unbound ferro-Cytc was decreased in SCO2-deficient cells, consistent with moderate cardiolipin enrichment, which is known to enhance retention of Cytc at the inner mitochondrial membrane. Despite elevated mitochondrial Cytc content, SCO2-deficient cells were less susceptible to apoptosis induced by intermittent hypoxia or dichloroacetate. Together, these findings indicate that reduced electron flux through complex IV is associated with Cytc accumulation through increased protein stability and membrane retention without enhancing apoptotic sensitivity.

Codon usage bias is a fundamental genomic characteristic that prefers non-random preferential use of synonymous codons. It is a major determinant of translational efficiency, gene regulation, and molecular evolution. However, the evolutionary bias and functional relevance of codon usage bias across the plant lineage is poorly defined and yet to understand what are the major factors responsible for relative synonymous codon usage (RSCU) in genomes and how codon usage bias influences the gene regulation, molecular evolution genomes. A genome-wide codon usage bias study of coding DNA sequences of 262 plant genome was conducted. It encompassed more than 4.6 billion codons from > 11 million coding sequences. Relative synonymous codon usage, codon adaptation index, codon-anticodon mapping, effective number of codon (ENC)-GC3, GC1,2-GC3, parity rule 2 (PR2-bias), molecular economy, and machine learning approaches were used for the study. It was found that codon usage bias was strongly non-random and exhibited a clear phylogenetic structuring. The higher plants favoured A/T-ending, whereas early-diverging lineages were enriched in G/C-ending codons. Analysis of RSCU, codon adaptation index, and codon-anticodon pairing indicated that translational selection is mediated by tRNA availability, contributing sustainability to these molecular patterns. Machine-learning approaches identified a small subset of codons having outsized influence on genome-wide codon usage landscapes. Further studies revealed the presence of robust inverse relationships between the effective number of codons and GC content at synonymous third positions. Neutrality analysis revealed approximately 61% of variation was driven by mutational pressure, tempered by selective constraints. Phylogenetic reconstruction showed a progressive relaxation of codon bias from algae to angiosperms while maintaining a conserved molecular economy cost of ~ 30 ATP per codon across the lineages. The study revealed codon usage bias is lineage-specific evolutionary conserved trait governed by mutation, selection, and translational optimization.

Significance Missense mutations can lead to pathological effects in human cells. Predictive methods that account for structural context, such as AlphaMissense, can provide pathogenicity scores. The accumulation of pathogenicity hotspots can reveal important structural features within individual proteins of protein families, such as GLUT transporters. Mapping pathogenicity scores onto the structure can thus provide a mechanistic explanation of the protein function necessary for its role in the cell. Abstract Non-synonymous amino acid substitutions (missense mutations) are common in the general population; some are causative of serious disease. Depending on their structural context, they can disrupt protein function, folding, or dynamics. Computational predictive methods developed in recent years, such as AlphaMissense, provide new insights into how missense mutations affect protein structure by predicting and mapping their pathogenicity across each amino acid in the human proteome. In this study, we identify recurring patterns of pathogenicity prediction across the GLUT family membrane transporters encoded by genes slc2a1-14. Within the GLUT transporter family, we observe higher pathogenicity profiles in the transmembrane domains, particularly in pore-lining and binding-site residues. Predicted missense pathogenicity is elevated throughout residues assigned to the central cavity, suggesting sensitivity of the transport pathway. Another finding shows higher pathogenicity in specific transmembrane helices of the protein, with the same pattern across all proteins. On the other hand, we observed lower pathogenicity values in some representatives of the GLUT family. These findings show that the pathogenicity of glucose transport within the GLUT family may be shaped by functional redundancy and physiological essentiality across GLUT groups.

Ancient environmental DNA (aeDNA) from permafrost, lake, cave, and marine sediments provides a rich source of genetic data that captures broad perspectives of past biodiversity. Accurate dating is crucial for discovering ecologically relevant patterns from aeDNA, and molecular clock dating would allow for sample ages to be estimated from the recovered genetic material itself instead of the geological components. However, the fragmented and damaged nature of short-read ancient DNA (aDNA) from multiple taxonomic sources poses significant challenges and has limited this dating approach for aeDNA. Here we developed ratePlacer, a phylogeny-based method for analyzing aeDNA that can combine information from many short reads in a sample while accounting for DNA damage to provide maximum likelihood estimates of sample ages. Simulations demonstrate that ratePlacer accurately dates samples even under the fragmented, damaged conditions characteristic of aeDNA and outperforms Bayesian tip-dating approaches for taxonomically mixed samples commonly found in aeDNA. Yet age estimates from re-dating Kap Kobenhavn varied across taxa, highlighting the difficulty of molecular clock dating in aeDNA. This dating also revealed elevated G[->]T and C[->]A mismatches consistent with oxidative damage. These patterns reveal aDNA damage beyond deamination and that remains understudied, suggesting that aeDNA should be carefully evaluated in genomic and evolutionary analyses. The new dating method, ratePlacer, extends molecular clock dating of aDNA from single-specimen to pooled environmental DNA data, where traditional methods struggle.

Stem cell dysfunction and loss of renewal capacity are primary characteristics of tissue aging and decremental regeneration in response to injury. Alveolar type 2 cells (AT2) are key progenitor cells responsible for lung repair and are thought to be dysfunctional in diseases such as chronic obstructive pulmonary disease (COPD). AT2 cells are highly metabolic and rely on mitochondria, but how mitochondrial mechanisms influence their maintenance and cell fate is unclear. This gap is critical as no current therapies target lung repair or mitochondrial function in COPD. Here, we report that adenine nucleotide translocase 2 (ANT2), a key ATP/ADP transporter, is reduced in AT2 cells from COPD lungs, and that ANT2 loss impairs bioenergetics (ATP). We also identify, for the first time, ferroptotic susceptibility as a consequence of ANT2 loss in AT2 cells, leading to impaired self-renewal and progenitor capacity in alveolar organoids. Together, loss of ANT2 and the associated cellular dysfunction resulted in worsened lung damage or emphysema due to cigarette smoke in mice. Therapeutic restoration of ANT2 expression resulted in renewed AT2 stem cell function and prevention of emphysema by reducing oxidative stress and ferroptosis. These findings highlight the importance of ANT2 in metabolic regulation, plasticity, and cell resiliency of AT2 cells in the lung and that ANT2 is a potential target for lung repair. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=114 SRC="FIGDIR/small/737954v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@96d0caorg.highwire.dtl.DTLVardef@165b15dorg.highwire.dtl.DTLVardef@15f86baorg.highwire.dtl.DTLVardef@862745_HPS_FORMAT_FIGEXP M_FIG C_FIG

Premise: Genome skimming (GS) is a cost-effective approach for plant phylogenomics, but its ability to recover informative datasets from different genomic compartments, particularly genome-wide SNPs, remains poorly explored in Solanum. Methods: We evaluated shallow GS for phylogenetic inference in South American prickly Solanum lineages by recovering plastid, mitochondrial, and nuclear datasets, including coding regions and genome-wide SNPs. Phylogenies were inferred using maximum-likelihood and coalescent approaches under different SNP filtering strategies. Results: GS successfully recovered complete plastomes, organellar coding regions, and large SNP datasets, but failed to consistently assemble mitochondrial genomes or recover low-copy nuclear genes. SNP-based analyses, especially from the nuclear genome, produced stable, well-supported phylogenies that were largely congruent across inference methods. In contrast, coding-region datasets, particularly from the mitochondrial genome, showed greater topological discordance, revealing cytonuclear conflict. Discussion: Our results demonstrate that shallow GS is an effective strategy for generating informative SNP datasets for phylogenetic inference in Solanum, despite limitations in recovering complete mitochondrial genomes and low-copy nuclear loci. SNP-based analyses substantially expand the phylogenetic potential of GS, providing a practical and cost-effective alternative for systematic studies.

Background: Epigenetic clocks are markers of biological aging that may vary in their sensitivity to environmental stressors and lifestyle modifiers. To evaluate the utility of these biomarkers as sensors of the human exposome, we investigated how they respond to two powerful and opposing exposures: smoking, a source of oxidative stress, and the antioxidant-rich Mediterranean diet. Objectives: We assessed the sensitivity of eleven epigenetic clocks to diet and smoking and evaluated whether Mediterranean diet adherence modifies associations between smoking and epigenetic aging. Methods: We analysed 928 participants (mean age 41 years, 59% male) from the Airwave Health Monitoring Study. Linear regression models assessed associations between Mediterranean Diet Score (MDS) and epigenetic age acceleration (EAA), alongside smoking status and blood cotinine. Interaction terms between smoking status and MDS were included to detect dietary attenuation of smoking-related EAA. Models were adjusted for demographic, socioeconomic, lifestyle, and psychological covariates. Results: Higher MDS was associated with lower EAA for GrimAge ({beta} = -0.07 SD; 95% CI: -0.13, -0.01) and Bernabeu ({beta} = -0.08 SD; 95% CI: -0.14, -0.02) after false discovery rate correction. Smoking was strongly associated with increased EAA, particularly for GrimAge, Bernabeu, and DunedinPACE. Among current smokers, effect sizes were greater in those with lower dietary adherence (e.g. GrimAge: 1.79 SD, 95% CI: 1.54, 2.04) compared with those with higher adherence (1.35 SD, 95% CI: 1.01, 1.68; P_interaction < 0.001). Similar attenuation patterns were observed for Bernabeu. Higher intake of fruits, vegetables, and whole grains contributed most to the attenuation of smoking-related EAA. Conclusions: Our findings indicate that certain epigenetic clocks effectively capture the tension between harmful and protective exposures within the exposome. Rather than suggesting that diet neutralises the risks of tobacco, these results demonstrate that specific clocks are sensitive enough to monitor how lifestyle factors modify molecular responses to environmental toxins. This highlights the value of second-generation clocks in quantifying biological resilience.

Symmetry elements in nucleic acids are most strongly correlated with sites of biological function; however, their relevance to non-canonical structures remains underexplored. In this study, we demonstrate the presence and significance of trinucleotide symmetry elements within G-quadruplex (G4) motifs. Our central hypothesis is that the intra-strand mirror symmetry of trinucleotides has been evolutionarily selected to facilitate G4 formation builds on the established sequence-structure association of G-quadruplexes and the natural symmetry law governing nucleotide insertion during genome evolution. Using a conserved G4 motif in the first exon of the MTOR gene as a model, we showed remarkable trinucleotide symmetry preservation across primates and broader mammals, with functional G4 regions displaying locally elevated symmetry relative to the codon-biased exonic background. Analysis of experimentally validated oncogenic G4s, including c-MYC, BCL2, VEGF, and KRAS, revealed that mirror and reverse complement symmetries converge around biologically important G4s. To quantify this feature, we formulated two complementary descriptors: the mirror symmetry index (MSI) and its non-palindromic variant (nMSI). Across 14 oncogene-promoter wild-type G4s, the majority scored MSI [&ge;] 0.80 (mean 0.884), with only the loop-rich ATG7, BCR, and MDM2 motifs falling below this value, and the KRAS promoter G4 reached individual significance against its mononucleotide-preserving null distribution (p = 0.042). Most decisively, each wild-type G4 scored higher on MSI than its experimentally confirmed G4-abolished mutant in 12 of 14 paired comparisons (sign test, p = 0.0065; mean {Delta}MSI = +0.089, mean {Delta}nMSI = +0.192); the two reversals (BCL2 and HIF-1) are attributable to scrambled mutant controls that introduce more balanced trinucleotide compositions rather than to failure of the index. The directional trend was reproduced across three independently published datasets, with nMSI [&ge;] 0.50 separating G4-forming from non-G4 sequences at 77.8% sensitivity and 100% specificity, although the collective per-sequence signal from mononucleotide-preserving shuffles remained a non-significant trend (Stouffer combined Z = 1.197, p = 0.116). This first report of trinucleotide symmetry in G4 motifs posits that coordinated nucleotide insertion and quadruplet maintenance act as an evolutionary forcing mechanism that pre-organizes single strands for G4 folding.

Loss of chromosome Y (LOY) is the most common aneuploidy in aging men and is increasingly recognized as a marker of aging and genomic instability. Because LOY occurs in mosaic form, its degree reflects the fraction of cells lacking the Y chromosome. Existing SNP-array- and sequencing-based methods rely largely on single genomic features and indirect transformations to estimate this fraction. We developed BaySeq-Y, a Bayesian method that directly estimates LOY mosaicism from sequencing data using VCF files with read depth (DP) and allelic depth (AD). Within a rigorous Bayesian framework, BaySeq-Y integrates complementary LOY-associated genomic features, including decreased read depth and allelic imbalance, and can additionally leverage haplotype phasing to improve precision. In simulations and fluorescence in situ hybridization validation (FISH), BaySeq-Y provided accurate estimates and outperformed existing methods. Applications to ROSMAP and GTEx supported its biological relevance through transcriptomic validation, demonstrating its utility for quantifying LOY across diverse sequencing datasets.

Microalgal metabolism relies on their chloroplasts, and involves both nucleus and plastidial-encoded proteins of various evolutionary origins. The plastidial ATP synthase complex is a key player in photosynthesis, and has been extensively studied in plants. However, our knowledge in other photosynthetic eukaryotes remains limited, despite their importance in marine environments. Here, we report the characterisation of a novel homologue of the F-type ATP synthase alpha subunit, hereby named xATPA, widespread in microalgae but absent from other photosynthetic organisms. Comparisons of xATPA sequences and predicted structures revealed a specific feature, the bump domain, and highlighted the absence of an ATP-binding site. We assessed xATPA prevalence in microalgae in the global ocean using environmental data from Tara Oceans, with a particular focus on diatoms, and demonstrate that its expression is associated with polar summer conditions. Using a reverse genetic approach in the model diatom Phaeodactylum tricornutum, we show that xATPAP t has a plastidial localisation, and that xATPA KO mutants exhibit growth deficiencies in a combination of low temperature, low salinity and constant light, consistent with environmental analysis. Surprisingly, both RNAseq and physiological assays suggest that xATPA is not involved in ATP synthase functions. On the other hand, xATPA interacts with other F1 ATP synthase subunits in vitro, which we suggest forms transient unassembled complexes. This study hence represents a comprehensive analysis of a novel protein from the environment to the lab, and reveals a new player in the plastidial physiology of eukaryotic microalgae.

The genus Rhynchospora Vahl (beak-sedges) comprises approximately 381 accepted species with a worldwide distribution, all of which possess holocentric chromosomes, where centromeric activity is distributed almost along the entire chromosome. Despite the recent advances, the mechanisms governing the dynamics of meiotic recombination in holocentric plants remain poorly understood. Here, we developed haplotype-specific oligo-FISH probes for chromosomes 1, 2, and 3 based on a haplotype-phased genome assembly of Rhynchospora breviuscula (n = 5), enabling homolog-specific chromosome painting. Each probe set was labelled with a distinct fluorophore and hybridised in situ to metaphase chromosomes of the reference plant and seven F1 individuals derived from self-crossed reference plants. This approach allowed the unambiguous discrimination of homologous haplotypes and the indirect visualisation of crossover (CO) events in recombined chromosomes. We observed that recombination events were predominantly located in terminal chromosomal regions, consistent across individuals. These results corroborate previous findings from single-cell recombination mapping and provide independent cytological validation of the recombination landscape in this species. Our study establishes haplotype-specific chromosome painting as a robust tool for high-resolution mapping of meiotic recombination in holocentric plants across generations. Furthermore, these probes provided a foundation for future investigations into inverted meiosis, a mechanism characterized by an alternative pattern of chromosome segregation in holocentric species.

Abstract Background: The influence of taurine on biological ageing remains unclear, particularly whether it acts as a causal driver or a functional biomarker. We aimed to disentangle the distinct roles of plasma taurine relative abundance, dietary taurine supply, and genetic metabolic capacity on all-cause mortality and unhealthy ageing. Methods: This prospective study used data from the Korean Genome and Epidemiology Study (2001~2022). A subcohort of 2,321 participants (mean age 56.5 years; 51.4% female) with complete metabolomic, dietary, and genomic data was analyzed. Three independent pathways were evaluated: (1) plasma taurine/total amino acid (AA) ratio, (2) dietary taurine to protein ratio, and (3) a weighted genetic risk score (GRS) from 21 SNPs in taurine biosynthesis and transport genes. Primary outcomes were all-cause mortality and unhealthy ageing (Physiological Healthy Ageing Index [PHAI] score [&le;] 25th percentile). Results: A higher plasma taurine/total AA ratio was consistently associated with improved ageing outcomes. Participants in the highest quartile showed 29% lower all-cause mortality (Hazard Ratio [HR], 0.71; 95% Confidence Interval [CI], 0.52-0.98; P for trend = .04) and lower risk of PHAI-based unhealthy ageing (HR, 0.77; 95% CI, 0.59-1.00; P for trend = .04) versus the lowest quartile. Dietary taurine-to-protein ratio was not associated with mortality (P for trend = .70), nor was the GRS (P for trend = .74). Conclusions: The protective association of taurine was linked to its relative abundance within the systemic amino acid pool, rather than dietary intake or genetic predisposition, supporting taurine as a functional biomarker of metabolic efficiency rather than a deterministic causal driver of ageing.

Regeneration of skeletal muscle preserves muscle mass and function, which decline with age. Here, we sought to identify long noncoding (lnc)RNAs involved in skeletal muscle myogenesis and potentially relevant to muscle aging. Cross-sectional analysis of skeletal muscle transcriptomes from healthy 22-through 89-year-old individuals revealed lncRNA LANCL1-AS1 among the top declining transcripts. Conversely, LANCL1-AS1 increased robustly during skeletal myogenesis and promoted myogenic differentiation in culture. Affinity pulldown by ChIRP followed by mass spectrometry revealed that LANCL1-AS1 associated with the mitochondrial protein LRPPRC, enhancing the formation of the chaperone complex LRPPRC-SLIRP, which maintains longer poly(A) tails of mitochondrial (mt-)mRNAs and stabilizes mt-mRNAs. Importantly, while myoblasts from old rhesus monkey muscle expressed lower levels of LANCL1-AS1 and mt-mRNAs, and displayed lower mitochondrial activity than young monkey myoblasts, overexpressing LANCL1-AS1 in old myoblasts restored mitochondrial activity and myogenesis. We propose that the age-associated reduction in LANCL1-AS1 contributes to impaired mitochondrial function and reduced myogenic capacity in aging skeletal muscle.

Mitochondrial diseases (MDs) are clinically heterogeneous rendering ascertainment challenging. Estimates of pathogenic mitochondrial DNA (mtDNA) variants in the population range from 1 in 200 to 1 in 4,000 individuals. Inclusion of mtDNA sequencing in genomic databases facilitates comprehensive estimation of mtDNA variation. However, interpretation of low heteroplasmy variation is complex, due in part to misalignment of nuclear mitochondrial DNA transcripts (NUMTs), whilst conservative heteroplasmy thresholds likely omit relevant variation. Cumulative burden of mtDNA variation contributes to aging and neurodegeneration, and recent characterisation of mitochondrial genome constraint allows quantitation of this burden. We analysed whole genome sequencing of blood DNA from 3,500 healthy older individuals in the Medical Genome Reference Bank using mity, considering pathogenic mtDNA variants [&ge;]1% heteroplasmy. We identified 34 distinct pathogenic mtDNA variants in 62 individuals, giving a combined population allele frequency of 1.77% (95% CI 1.36-2.27) or 1 in 56 individuals. We evaluated inclusion of false positive (FP) calls due to two common NUMTs, which accounted for up to 16% of variants. Increasing heteroplasmy thresholding to eliminate all NUMT-FPs also eliminated much of the total variation, including pathogenic variants. We propose a sample-specific, scaled heteroplasmy threshold to maximise variant retention and mitigate NUMT-FPs. Finally, we characterised measures of mitochondrial constraint in this healthy older cohort, observing an association between variant burden and summed constraint, whilst mean constraint was higher in pathogenic variant carriers. These findings suggest pathogenic mtDNA variation is more common in the population than is currently appreciated. Findings are comparable to larger genomic databases when heteroplasmy thresholding is adjusted, and support earlier population-based estimates. Incorporation of low heteroplasmy variation is relevant, but interpretation is nuanced, and optimising variant retention requires consideration of NUMT-FP rates.

Many Vicia species contain vicine and convicine (VC), which limit the use of faba bean and some vetches in food and feed. The first step in VC biosynthesis in V. faba is shared with the riboflavin pathway and attributed to VC1, a member of the ribAB family. Since riboflavin is ubiquitous to life, we examined the distribution of VC production in genus Vicia. Three accessions of each of 33 Vicia species were grown in glasshouse conditions to provide fresh seeds for VC analysis and leaves for DNA analysis. PCR was used to amplify fragments of the VC1/ribAB gene for sequencing, and these sequences were used to create a phylogenetic tree. COX1 and ITS2 sequences were used for examining the nucleotide diversity in the subgenera. VC and DNA sequences consistent with VC1 were found only in members of subgenus Vicia. In V. lathyroides, VC1 was present but no VC was detected. There was less sequence diversity in VC1/ribAB sequences of subgenus Cracca than in those of subgenus Vicia, suggesting that ribAB remained under stricter purifying selection than VC1. VC1 is confirmed as a prerequisite for the presence of VC, and the gene and its products are restricted to subgenus Vicia. HighlightThe favism-causing factors of vetches and faba bean, vicine and convicine, depend on the presence of the VC1 variant of the ribAB gene, which is found in only one subgenus.

Autism development involves multiple genetic and early-life environmental factors. Studying the placenta's gene expression profile may reveal key mechanistic pathways in autism development. Here, using a nested case-cohort design within an Australian population-derived prebirth cohort study (n=1074), we identified 1,644 differentially expressed genes (DEGs; FDR<0.05) in the placenta of children with autism diagnosis (n=43), compared to those without (n=120). The top enriched pathways related to mitochondrial translation, oxidative stress, RNA processing and transcription regulation. CYP1A1, the most important xenobiotic-metabolising enzyme of the placenta, was the top downregulated DEG in the placenta of children with autism, while immuno-regulatory human leukocyte antigen (HLA)-related genes were among the top upregulated DEGs. A machine learning-based approach predicted autism from the transcriptomic data with a median sensitivity of 0.57 (2.5th-97.5th centiles: 0.29, 0.76) and median specificity of 0.92 (2.5th-97.5th centiles: 0.78, 0.98). Weighted Gene Correlation Network Analysis identified eight affected placental gene modules, with the largest five modules being enriched primarily for mitochondrial bioenergetics, oxidative phosphorylation and RNA processing pathways. This placental transcriptomic signature of impaired mitochondrial function and gene transcription regulation among infants subsequently diagnosed with autism has profound implications for understanding both risk factors and prediction, suggesting the possibility of identifying modifiable prenatal pathways to improve autism outcomes.

BackgroundAberrant NAD+ metabolism has been implicated in the pathogenesis of cancer cachexia, highlighting this pathway as a potential therapeutic target to mitigate skeletal muscle wasting. However, it remains unclear whether chemotherapeutic agents contribute to the onset of cachexia by disrupting NAD+ metabolism. Here, we investigated the effects of commonly used chemotherapy regimens on NAD+ metabolism in skeletal muscle and liver of healthy mice. MethodsHealthy mice were treated with either 2-week regimens of folfiri or cisplatin, or 5-week regimens of folfiri or folfox, with vehicle-treated mice serving as controls. Cachexia-related outcomes were assessed, while skeletal muscle and liver tissues were analyzed for NAD metabolites and markers of NAD+ metabolism. Given the consistent downregulation of the NAD+ biosynthetic enzyme Nrk2 in cachectic chemotherapy-treated mice, we examined skeletal muscle Nrk2/NRK2 expression across published murine and human cachexia datasets, and in additional models of muscle wasting and hypertrophy. ResultsNAD+ loss was observed in atrophic muscle following administration of cisplatin (2-week treatment; -14% vs controls, p=0.047) and folfiri (5-week treatment; -18%, p=0.069). In contrast, muscle NAD+ levels were preserved in non-atrophic groups (2-week folfiri and 5-week folfox). Muscle Nrk2 was the most responsive NAD+ biosynthetic enzyme, showing consistent downregulation across chemotherapy models with ongoing or developing muscle loss: cisplatin (-93%, p<0.001), folfiri (-84%, p<0.001) and folfox (-92%, p<0.001). In the liver, NAD+ levels declined after prolonged 5-week folfiri (-20%, p=0.013) and folfox (-15%, p=0.043) treatments. These changes were accompanied by distinct alterations in NAD+ biosynthesis pathways, indicating treatment-specific reorganization of hepatic NAD+ metabolism. Cross-study analyses revealed early and consistent skeletal muscle Nrk2 downregulation across multiple murine cachexia models and human inactivity studies, whereas cachexia-targeted interventions in rodents and resistance training in humans increased its expression. ConclusionsThese findings demonstrate that chemotherapy distrupts tissue NAD+ metabolism, with skeletal muscle NAD+ loss accompanying muscle atrophy and hepatic NAD+ levels declining after prolonged treatment. The early and robust responsiveness of muscle Nrk2 expression to changes in muscle mass underscores its potential as a dynamic indicator for predicting treatment-induced changes in muscle mass. Together, these results provide new molecular insight into the metabolic basis of chemotherapy-induced muscle wasting and support further investigation of NAD+-targeted strategies in this context.

Cell size is a critical morphological parameter determining cellular functional homeostasis, yet existing large-scale transcriptomic databases lack direct cell size measurement data. By integrating high-resolution immunofluorescence images with transcriptomics, we identified 457 genes significantly correlated with cell area. Based on these findings, we developed an algorithm, Cell Size Score (CSS), to predict cell size from gene expression profiles. Validation across multiple independent datasets, including human cell lines, mouse models, and single-cell spatial transcriptomics, confirmed that CSS accurately predicts cell size. Furthermore, we observed a significant positive correlation between CSS and broad-spectrum chemotherapy drug resistance, suggesting that increased cell volume confers survival advantages to cancer cells. Moreover, CSS analysis of aging revealed sex-dependent, tissue-specific patterns of change, wherein male adipose and cardiac tissues exhibited progressive hypertrophy with age, while female reproductive organs showed significant atrophy. Additionally, CSS significantly increased in skeletal muscle after exercise, indicating that this metric can capture dynamic physiological adaptation processes. This study establishes a bridge between transcriptomics and cell morphology, providing novel insights into retrospectively analyzing the role of cell size in pathological and physiological processes such as cancer and aging using existing omics data, as well as understanding the molecular mechanisms underlying cell size regulation.

Objectives: We tested how multi-level socioeconomic disadvantage relates to biological aging and systemic inflammation in women and men from the population-based Canadian Longitudinal Study on Aging (CLSA). Methods: We examined cross-sectional data from 8,516 CLSA participants with baseline measures on systemic inflammatory biomarkers (C-reactive protein, interleukin-6, and tumoral necrosis factor-) and biological aging (metabolomic and six DNA methylation [DNAm] age estimates). Plasma samples underwent metabolomic profiling by Metabolon, Inc. Metabolomic age was estimated separately in males and females using sex-stratified models based on age-correlated metabolite levels. DNAm data generated using the Illumina Infinium MethylationEPIC v1.0 array were used to estimate DNAm age across six established models, including Horvath, Hannum, PhenoAge, GrimAge, GrimAge2, and DunedinPACE. We used log-transformed metabolite levels to calculate metabolomic age by sex. We linked education, income, material and social deprivation to biomarkers of systemic inflammation and biological aging stratified by sex using generalized linear models. Multivariable models were adjusted by age, major behavioral risk factors, and chronic conditions. Results: Participants were aged on average of 62.6 years of age, and approximately 50% were females. In multivariable linear adjusted models, we found that in comparison to those earning [&ge;]$100K a year, women earning less <$20K were on average 1.14 (95%CI: 0.46, 1.82) year older with respect to metabolomic age; those earning [&ge;]$20K & <$50K were on average 0.90 (95%CI: 0.26, 1.53) years older; and those earning [&ge;]$50K & <$100K were on average 0.70 (95%CI: 0.05, 1.34) years older. We did not observe this dose response among men. A similar dose-response association was observed for interleukin-6 in both men and women. Discussion: These findings suggest that socioeconomic adversity influences not only inflammatory pathways but also distinct biological aging processes, including metabolomic aging.

The insertase Oxa1 is required for protein insertion into the inner mitochondrial membrane and for the biogenesis of oxidative phosphorylation complexes. While most eukaryotes encode one or two Oxa1 proteins, we identified three paralogs in Trypanosoma brucei: TbOxa1-1, TbOxa1-2, and TbOxa1-3. Knock-out of individual paralogs followed by phenotypic analyses and proteomic characterization of submitochondrial fractions revealed distinct functions. Respiratory chain complexes I and IV are primarily affected by loss of TbOxa1-1, whereas complex III and ATP synthase depend on TbOxa1-2; the ablation of TbOxa1-3 results in minor phenotypes in culture. In TbOxa1-2-depleted cells, ATP synthase biogenesis is compromised by the defective import or processing of the nuclear-encoded subunit-c, which also requires a rhomboid peptidase-like protein. Further, the ablation of TbOxa1-2 triggers accumulation of membrane proteins in the matrix, supporting its role in conservative sorting. Together, our results demonstrate that the trypanosomal Oxa1 machinery evolved a paralog-specific division of labor to manage a highly divergent mitochondrial membrane proteome.