Regulation of food intake in mammals is complex and controlled by an interplay between hedonic and homeostatic signals, including hormones like leptin, which senses fat storage and suppresses food intake. Caenorhabditis elegans lack leptin and leptin receptors but still exhibit controlled eating. Here, we show that in C. elegans eating can be regulated by a balance between saturated and monounsaturated fatty acids interacting with transcriptional pathways regulating lipid synthesis, c-AMP response element binding protein and AMP kinase. This effect is mediated at the endoplasmic reticulum through formation of phospholipids and activation of the IRE-1 sensor in the nervous system, which controls behavior through neuronal serotonin and the G-protein-coupled ligand/ receptor pair PDF-1/PDFR-1. We show that this peptide/receptor pair may be an ancestral precursor of the whole family of GLP-1/GIP-related peptides and their receptors. Indeed, administration of a 37 amino acid peptide derived from PDF-1 resulted in a reduction in body weight and improved insulin sensitivity in mice. In worms, signaling through this pathway induced food-leaving behavior on concentrated food and roaming behavior on dispersed food, a state we have termed "food-apathy," paralleling pharmacologic effects of GLP-1/GIP-related peptides in humans. These findings highlight the potential evolutionary origin of this family of hormones and their receptors, and its link to metabolic and neuronal responses in control of feeding behavior.

Leber congenital amaurosis (LCA) and Early-onset severe retinal dystrophy (EOSRD) manifest within the first months and the first years of life, respectively. They are the leading cause of severe vision impairment in childhood. Using next generation sequencing, we identified eight families of patients with LCA/EOSRD carrying biallelic combination of six germline variants in DDX41 , encoding a DEAD-box ATPase RNA helicase involved in RNA splicing, innate immunity and hematopoiesis. In fibroblasts from a patient carrying the homozygous missense variant c.1187T>C (p. Ile396Thr) and in the retina of Ddx41 I396T/I396T mice, DDX41 protein expression was decreased. Electroretinogram recordings in these animals also revealed significant visual dysfunction since the first month of age, supporting a pathogenic role of DDX41 in retinal physiology. Immunohistochemical staining showed that the protein localized to nuclei in all major retinal cell types and to photoreceptor synapses, while biochemical assays showed that LCA/EOSRD variants disrupt DDX41 interactions with RNA through misfolding or the formation of non-productive aggregates, resulting in loss-of-function. Transcriptomic profiling of mutant mouse retinas revealed dysregulation of gene networks associated with Müller cells (MCs), glial cells essential for maintaining retinal structure, metabolic balance, and immune surveillance. The dysregulated pathways chiefly involved cell morphogenesis and junction formation, consistent with immunohistological analyses of widespread architectural disruption and nuclear disorganization, identifying MCs as a site of dysfunction. Together, these findings establish for the first time the involvement of DDX41 in LCA/EOSRD and provide new insights into the role of helicases in retinal homeostasis.

The actin cytoskeleton is a fundamental and highly conserved structure that functions in diverse cellular processes, yet its direct contribution to organismal aging remains unclear. Here, we systematically interrogated how genetic and pharmacologic perturbations of actin structure and function influence lifespan and various hallmarks of aging in Caenorhabditis elegans. Whole-animal and tissue-specific knockdown of actin and key actin-binding proteins (ABPs)—arx-2 (Arp2/3), unc-60 (cofilin), and lev-11 (tropomyosin)—led to premature disruption of filament organization, reduced lifespan, and tissue-specific physiological defects. Actin dysfunction also displayed a more “aged” transcriptome using previously validated transcriptomics clocks, and broadly exacerbated many age-associated phenotypes, including mitochondrial dysfunction, lipid dysregulation, loss of proteostasis, impaired autophagy, and intestinal barrier failure. Pharmacological destabilization with Latrunculin A mirrored genetic knockdowns, while mild stabilization with Jasplakinolide modestly extended lifespan, emphasizing that optimal and finely tuned actin function is critical for healthy aging. Finally, analysis of human genome-wide association data revealed that common ACTB polymorphisms correlate with differences in age-related decline in gait speed, suggesting some links between aging and actin across organisms. Taken together, our results provide a comprehensive and publicly accessible resource that maps, for the first time, how changes in actin integrity correlate with diverse aging phenotypes across tissues. This descriptive framework is intended to enable future mechanistic discovery by offering a deep, unbiased dataset that can be integrated with emerging studies to define how actin dynamics can potentially influence aging.

Inherited retinal diseases (IRDs) comprise a diverse group of disorders that frequently lead to progressive vision impairment and blindness. Despite advances in genetic testing, a significant number of IRD cases remain genetically unsolved, often due to unidentified disease-associated genes or variants. This study reports additional cases for the newly discovered IRD genes of the AP-5 complex. A comprehensive ophthalmological evaluation was performed for all patients, including retinal imaging (multimodal imaging), visual field testing, and electroretinogram (ERG) testing. Whole-genome and -exome sequencing (WGS and WES) were performed for clinically unsolved IRD patients, and data were analyzed to identify underlying causal variants. The identified variants were subsequently validated using Sanger sequencing. Five unrelated patients from Europe and Iran were identified with a distinctive macular degeneration associated with bi-allelic variants in AP5Z1 (HGNC: 22197) and AP5B1 (HGNC: 25104), subunits of the vesicular fifth adaptor protein (AP-5) complex. The AP-5 complex is the part of the intracellular trafficking machinery thought to be involved in cellular homeostasis and lysosomal functioning in the retinal pigment epithelium (RPE). The identification of bi-allelic variants in two proteins of the AP-5 complex expand the characterization of AP-5 genes in sustaining and preserving normal macular function.

Retinitis pigmentosa (RP) is an inherited retinal disease (IRD) characterized usually by progressive photoreceptor degeneration, leading to night blindness, peripheral visual field loss, and can progress to central vision impairment in some individuals. Despite advances in genomic diagnostics, many individuals with RP remain without a molecular diagnosis. We identified bi-allelic ultra-rare variants in fibronectin type II and Spry domain-containing protein 1-like (FSD1L) in six individuals with RP with or without neurological features from four unrelated families. FSD1L encodes a cytoplasmic protein, variants of which have not previously been associated with Mendelian disease. The gene is expressed in both human and mouse retinas that are enriched in cone and rod photoreceptors. Immunofluorescence and ultrastructure expansion microscopy show that FSD1L localizes along the photoreceptor microtubule axoneme, including the connecting cilium and outer segment, supporting a possible role in intracellular trafficking. A retina-enriched isoform of FSD1L includes an alternatively spliced exon (exon 10b), which we characterize as absent in minigene assays and affected individual-derived lymphocytes due to a deep intronic 26 nt deletion. Together, these findings support the association between bi-allelic disruption of FSD1L and IRD.

Metabolism unfolds within specific organelles in eukaryotic cells. Lysosomes are highly metabolically active organelles, and their metabolic states dynamically influence signal transduction, cellular homeostasis and organismal physiopathology. Despite the importance of lysosomal metabolism, a method for its in vivo measurement is currently lacking. Here we report a fluorescence-detected mid-infrared photothermal microscope (FILM) implemented with optical boxcar demodulation, artificial intelligence-assisted data denoising and spectral deconvolution, to map metabolic activity and composition of individual lysosomes in living cells and organisms. Using this method, we uncovered lipolysis and proteolysis heterogeneity across lysosomes within the same cell, as well as early-onset lysosomal dysfunction during organismal aging. In addition, we discovered organelle-level metabolic changes associated with diverse lysosomal storage diseases. This method holds the broad potential to profile metabolic fingerprints of individual organelles within their native context and quantitatively assess their dynamic changes under different physiological and pathological conditions, providing a high-resolution chemical cellular atlas.

The standard for in silico pathogenicity prediction of in-frame insertions and deletions (indels) is less established compared to other types of variations. We aimed to systematically assess the performance of in silico machine learning (ML) tools on a patient cohort with inherited retinal diseases (IRDs). The performance of four ML tools (CADD, FATHMM-indel, VEST4, and MetaRNN-indel) was compared. Among them, MetaRNN-indel showed the best overall results. MetaRNN-indel was then applied to 1013 unsolved IRD patients, identifying two likely pathogenic causal variants in two unrelated IRD patients by confirming clinical phenotypes. Hence, our findings indicate that reliable prediction of the pathogenicity of in-frame indels can be achieved using existing ML tools with proper evaluation and tuning.

Microbiota-derived metabolites have emerged as key regulators of longevity. The metabolic activity of the gut microbiota, influenced by dietary components and ingested chemical compounds, profoundly impacts host fitness. While the benefits of dietary prebiotics are well-known, chemically targeting the gut microbiota to enhance host fitness remains largely unexplored. Here, we report a novel chemical approach to induce a pro-longevity bacterial metabolite in the host gut. We discovered that wild-type Escherichia coli strains overproduce colanic acids (CAs) when exposed to a low dose of cephaloridine, leading to an increased life span in the host organism Caenorhabditis elegans. In the mouse gut, oral administration of low-dose cephaloridine induced transcription of the capsular polysaccharide synthesis (cps) operon responsible for CA biosynthesis in commensal E. coli at 37 °C, and attenuated age-related metabolic changes. We also found that low-dose cephaloridine overcomes the temperature-dependent inhibition of CA biosynthesis and promotes its induction through a mechanism mediated by the membrane-bound histidine kinase ZraS, independently of cephaloridine's known antibiotic properties. Our work lays a foundation for microbiota-based therapeutics through chemical modulation of bacterial metabolism and highlights the promising potential of leveraging bacteria-targeting drugs in promoting host longevity.

The spatial distribution of diverse biomolecules in multicellular organisms is essential for their physiological functions. High-throughput in situ mapping of biomolecules is crucial for both basic and medical research, and requires high scanning speed, spatial resolution, and chemical sensitivity. Here, we developed a Tissue Expansion method compatible with matrix-assisted laser desorption/ionization Mass spectrometry Imaging (TEMI). TEMI reaches single-cell spatial resolution without sacrificing voxel throughput and enables the profiling of hundreds of biomolecules, including lipids, metabolites, peptides (proteins), and N-glycans. Using TEMI, we mapped the spatial distribution of biomolecules across various mammalian tissues and uncovered metabolic heterogeneity in tumors. TEMI can be easily adapted and broadly applied in biological and medical research, to advance spatial multi-omics profiling.

Intracellular lipid binding proteins (iLBPs) play crucial roles in lipid transport and cellular metabolism across the animal kingdom. Recently, a fat-to-neuron axis was described in Caenorhabditis elegans , in which lysosomal activity in the fat liberates polyunsaturated fatty acids (PUFAs) that signal to neurons and extend lifespan with durable fecundity. In this study, we investigate the structure and binding mechanisms of a lifespan-extending lipid chaperone, lipid binding protein-3 (LBP-3), which shuttles dihomo-γ-linolenic (DGLA) acid from intestinal fat to neurons. We present the first high-resolution crystal structure of LBP-3, which reveals a classic iLBP fold with an unexpected and unique homodimeric arrangement via interstrand interactions that is incompatible with ligand binding. We identify key ionic interactions that mediate DGLA binding within the lipid binding pocket. Molecular dynamics simulations further elucidate LBP-3's preferential binding to DGLA due to its rotational freedom and access to favorable binding conformations compared to other 20-carbon PUFAs. We also propose that LBP-3 dimerization may be a unique regulatory mechanism for lipid chaperones.

The endolysosomal system plays a crucial role in maintaining cellular homeostasis and promoting organism fitness. The pH of its acidic compartments is a crucial parameter for proper function, and it is dynamically influenced by both intracellular and environmental factors. Here, we present a method based on fluorescence lifetime imaging microscopy (FLIM) for quantitatively analyzing the pH profiles of acidic endolysosomal compartments in diverse types of primary mammalian cells and in live organism Caenorhabditis elegans. This FLIM-based method exhibits high sensitivity in resolving subtle pH differences, thereby revealing heterogeneity within a cell and across cell types. This method enables rapid measurement of pH changes in the acidic endolysosomal system in response to various environmental stimuli. Furthermore, the fast FLIM measurement of pH-sensitive dyes circumvents the need for transgenic reporters and mitigates potential confounding factors associated with varying dye concentrations or excitation light intensity. This FLIM approach offers absolute pH quantification and highlights the significance of pH heterogeneity and dynamics, offering a valuable tool for investigating lysosomal functions and their regulation in various physiological and pathological contexts.

The epigenome is sensitive to metabolic inputs and is crucial for aging. Lysosomes act as a signaling hub to sense metabolic cues and regulate longevity. We found that lysosomal metabolic pathways signal through the epigenome to regulate transgenerational longevity in Caenorhabditis elegans. Activation of lysosomal lipid signaling and lysosomal adenosine monophosphate-activated protein kinase (AMPK) or reduction of lysosomal mechanistic target of rapamycin (mTOR) signaling increased the expression of a histone H3.3 variant and increased its methylation on K79, leading to life-span extension across multiple generations. This transgenerational prolongevity effect required intestine-to-germline transportation of histone H3.3 and a germline-specific H3K79 methyltransferase and was recapitulated by overexpressing H3.3 or the H3K79 methyltransferase. Thus, signals from a lysosome affect the epigenome and link the soma and germ line to mediate transgenerational inheritance of longevity.

Advances in cellular physiology research have created the need to visualize metabolic processes in both time and space. Vibrational microscopy has emerged as a promising method at the forefront of this field. Here, we summarize recent progress and offer perspectives on the application of vibrational microscopy for uncovering metabolic actions in vivo.

Differential and even opposing functions of two major antioxidant transcription factors Nrf1 and Nrf2 (encoded by Nfe2l1 and Nfe2l2, respectively) are determined by distinctions in their tempospatial positioning, topological repartitioning, proteolytic processing, and biochemical modification, as well as in their shared evolutionary origin. As a matter of fact, the allelopathic potentials of Nrf1 and Nrf2 (both resembling two entangled ‘Yin-Yang’ quanta that comply with a dialectic law of the unity of opposites) are fulfilled to coordinately control redox physiological homeostasis so as to be maintained within the presetting thresholds. By putative exponential curves of redox stress and intrinsic anti-redox capability, there is inferable to exist a set point at approaching zero with the ‘Golden Mean’ for the healthy survival (i.e., dubbed the ‘zero theory’). A bulk of the hitherto accumulating evidence demonstrates that the set point of redox homeostasis is dictated selectively by multi-hierarchical threshold settings, in which the living fossil-like Nrf1 acts as a robust indispensable determinon, whereas Nrf2 serves as a versatile chameleon-like master regulon, in governing the redox homeodynamic ranges. This is attributable to the facts that Nrf2 has exerted certain ‘double-edged sword’ effects on life process, whereas Nrf1 executes its essential physiobiological functions, along with unique pathophysiological phenotypes, by integrating its ‘three-in-one’ roles elicited as a specific triplet of direct sensor, transducer and effector within multi-hierarchical stress responsive signaling to redox metabolism and target gene reprogramming. Here, we also critically reviewed redox regulation of physio-pathological functions from the eco-evo-devo perspectives, through those coding rules (redox code, stress-coping code, and topogenetic code). The evolving concepts on stress and redox stress were also further revisited by scientific principles of physics and chemistry. Besides, several novel concepts such as oncoprotists, Reverse Central Dogma, and Grand Redox-Unifying Theory’ (GRUT) of life, together with diffusive reactive species (DRS)-based murburn concept integrating all stochastic electron-, proton- and/or moiety-transfer reactive and interactive processes (e.g., PCHEMS), are introduced in this interdisciplinary and synthetic review.

The actin cytoskeleton is a fundamental and highly conserved structure that functions in diverse cellular processes, yet its direct contribution to organismal aging remains unclear. Here, we systematically interrogated how genetic and pharmacologic perturbations of actin structure and function influence lifespan and various hallmarks of aging in Caenorhabditis elegans . Whole-animal and tissue-specific knockdown of actin and key actin-binding proteins (ABPs) - arx-2 (Arp2/3), unc-60 (cofilin), and lev-11 (tropomyosin) - led to premature disruption of filament organization, reduced lifespan, and tissue-specific physiological defects. Bulk and single-nucleus RNA-sequencing revealed that ABP knockdowns elicited a strongly “aged” transcriptome. Actin dysfunction broadly exacerbated many age-associated phenotypes, including mitochondrial dysfunction, lipid dysregulation, loss of proteostasis, impaired autophagy, and intestinal barrier failure. Pharmacological destabilization with Latrunculin A mirrored genetic knockdowns, while mild stabilization with Jasplakinolide modestly extended lifespan, emphasizing that optimal and finely-tuned actin function is critical for healthy aging. Finally, analysis of human genome-wide association data revealed that common ACTB polymorphisms correlate with differences in age-related decline in gait speed, suggesting evolutionary conservation of actin’s role in healthy aging. Taken together, our results provide a comprehensive and publicly accessible resource that maps, for the first time, how actin integrity intersects with diverse aging pathways across tissues and scales. This descriptive framework is intended to enable future mechanistic discovery by offering a deep, unbiased dataset that can be integrated with emerging studies to define how actin dynamics contribute to aging.

Metabolism is fundamental to organism physiology and pathology. From the intricate network of metabolic reactions, diverse chemical molecules, collectively termed metabolites, are produced. In multicellular organisms, metabolite communication between different tissues is vital for maintaining homeostasis and adaptation. However, the molecular mechanisms mediating these metabolite communications remain poorly understood. Here, we focus on nucleosides and nucleotides, essential metabolites involved in multiple cellular processes, and report the pivotal role of the SLC29A family of transporters in mediating nucleoside coordination between the soma and the germline. Through genetic analysis, we discovered that two Caenorhabditis elegans homologs of SLC29A transporters, Equilibrative Nucleoside Transporter ENT-1 and ENT-2, act in the germline and the intestine, respectively, to regulate reproduction. Their knockdown synergistically results in sterility. Further single-cell transcriptomic and targeted metabolomic profiling revealed that the ENT double knockdown specifically affects genes in the purine biosynthesis pathway and reduces the ratio of guanosine to adenosine levels. Importantly, guanosine supplementation into the body cavity/pseudocoelom through microinjection rescued the sterility caused by the ENT double knockdown, whereas adenosine microinjection had no effect. Together, these studies support guanosine as a rate-limiting factor in the control of reproduction, uncover the previously unknown nucleoside/nucleotide communication between the soma and the germline essential for reproductive success, and highlight the significance of SLC-mediated cell-nonautonomous metabolite coordination in regulating organism physiology.

Geroscience aims to target the aging process to extend healthspan. However, even isogenic individuals show heterogeneity in natural aging rate and responsiveness to pro-longevity interventions, limiting translational potential. Using RNAseq analysis of young, isogenic, subpopulations of Caenorhabditis elegans selected solely on the basis of the splicing pattern of an in vivo minigene reporter that is predictive of future life expectancy, we find a strong correlation in young animals between predicted life span and alternative splicing of mRNAs related to lipid metabolism. The activity of two RNA splicing factors, Reversed Polarity-1 (REPO-1) and Splicing Factor 1 (SFA-1), early in life is necessary for C. elegans response to specific longevity interventions and leads to context-specific changes to fat content that is mirrored by knockdown of their direct target POD-2/ACC1. Moreover, POD-2/ACC1 is required for the same longevity interventions as REPO-1/SFA-1. In addition, early inhibition of REPO-1 renders animals refractory to late onset suppression of the TORC1 pathway. Together, we propose that splicing factor activity establishes a cellular landscape early in life that enables responsiveness to specific longevity interventions and may explain variance in efficacy between individuals.