Biology subjects

Prasad, P.

Publications and source records attributed to Prasad, P..

At least 19 recordsLinked to original sources

bioRxiv2026

Hyperactivation of β-catenin rewires mitochondrial fuel selection in liver cancer cells

Metabolic state is tightly linked to cell fate, and expression of the mitochondrial pyruvate carrier (MPC), which imports pyruvate into mitochondria, is a robust marker of it. The transcriptional regulators that set metabolic state, however, remain poorly defined. In the liver, {beta}-catenin is one of the determinants of metabolic zonation, and activating CTNNB1 mutations are frequent in hepatocellular carcinoma. Here we show that {beta}-catenin regulates metabolic state in liver cancer cells. An unbiased CRISPR screen that uses endogenous MPC1 transcript abundance as the marker identifies CTNNB1. Activating {beta}-catenin pharmacologically or genetically reduces MPC1 and MPC2 transcript and protein levels, and MPC expression is also low in {beta}-catenin-driven liver tumors in vivo. Beyond the MPC, {beta}-catenin activation impairs assembly of respiratory Complexes III and IV, reduces mitochondrial DNA content and respiratory capacity, elongates mitochondrial networks and remodels the lipidome. Stable isotope tracing shows reduced entry of glucose- and lactate-derived carbon into the TCA cycle alongside increased oxidation of palmitate and oleate. These findings identify {beta}-catenin as a regulator of metabolic state that drives a fuel switch from carbohydrate to fatty acid oxidation in liver cancer cells.

biochemistry↗

bioRxiv2026

A genetic screen to identify novel interactions of the phosphatidylinositol phosphate signaling pathway in Drosophila spermatogenesis

Phosphatidylinositol (PI) phosphates (PIPs) are membrane lipids with crucial roles in intracellular signaling, membrane trafficking and cytoskeletal organization. To perturb PIP function in Drosophila melanogaster sperm development, we previously expressed the Salmonella PIP phosphatase SigD (also known as SopB) under control of a spermatocyte- specific promoter. SigD expression results in reduced levels of plasma membrane PI 4,5-bisphosphate [PI(4,5)P2; henceforth PIP2] and male sterility. Transgenic lines expressing high levels of SigD (SigD-high) exhibit defects in spermatocyte cytokinesis, ciliary transition zone maturation and spermatid elongation, whereas lines expressing low levels of SigD (SigD-low) have defects in nuclear shaping and spermatid cell polarization. To further investigate the role of PIPs during Drosophila sperm development, we conducted a sensitized genetic screen to identify suppressors and enhancers of SigD-low phenotypes. From a screen of 760 chromosomal deficiencies spanning 83.6% of chromosomes 2 and 3, we identified 80 interacting regions that were further narrowed down to 40 individual suppressors. Among the strongest interactors were genes involved in microtubule dynamics, nucleocytoplasmic transport and membrane trafficking. The novel genetic interactions identified by our screen pave the way for further investigation into molecular mechanisms underlying the affected aspects of sperm development.

genetics↗

bioRxiv2026

Derlin-mediated ERAD of lipid regulator ORMDL3 safeguards mitochondrial function

Mammalian Derlin proteins (Derlin-1, Derlin-2, and Derlin-3) are conserved components of the endoplasmic reticulum-associated degradation (ERAD) machinery that mediate the retrotranslocation and proteasomal degradation of misfolded ER-resident proteins. However, their paralog-specific contributions to cellular homeostasis remain poorly understood. Here, we show that Derlin deficiency disrupts mitochondrial architecture and results in mitochondrial fragmentation and tightening of ER-mitochondria contact sites (MERCs) in HEK293 cells. Mechanistically, we identify ORMDL proteins, evolutionarily conserved negative regulators of sphingolipid biosynthesis, as substrates of Derlin-2- and Derlin-3-dependent ERAD. Derlin deficiency leads to selective accumulation of ORMDL3 and its dose-dependent enrichment at MERCs, where it drives mitochondrial dysfunction in respiration and calcium handling. Reducing ORMDL3 levels restores mitochondrial function, establishing ORMDL3 as a key effector downstream of Derlin loss. Our work establishes ERAD as a critical mechanism of protein quantity control that safeguards organelle homeostasis by preventing aberrant accumulation and mislocalization of ER clients at inter-organelle contact sites. Given that ORMDL family members are central regulators of sphingolipid metabolism and are genetically linked to inflammation, cancer, asthma, inflammatory bowel disease, type 1 and type 2 diabetes, multiple sclerosis, obesity, and nonalcoholic fatty liver disease, these findings connect ERAD-dependent spatial control to sphingolipid homeostasis and a broad spectrum of human pathologies.

cell biology↗

bioRxiv2026

Circular Smad1-Encoded Polypeptide Regulates Myogenesis

The majority of RNAs transcribed from the genome are non-coding RNAs (ncRNAs) that are involved in regulating the expression of protein-coding genes. However, a growing body of research highlights several novel microproteins encoded by unconventional ncRNAs such as long non-coding RNAs and circular RNAs (circRNAs) as important regulators of disease and development. Although several circRNAs have recently been reported to translate into functional peptides in diverse tissues, their roles in skeletal muscle remain largely unexplored. In this study, polyribosome-associated RNA sequencing and publicly available translatable circRNAs from the riboCIRC database were curated to discover potential protein-coding circRNAs in mouse C2C12 skeletal muscle cells. We validated a few circRNAs with high potential of translating into proteins in mouse C2C12 cells, including circular Smad1 (circSmad1) that encodes a 194 amino acid peptide called circSmad1-194aa. Interestingly, silencing of circSmad1 in C2C12 cells resulted in loss of myotube fusion and maturation. CircSmad1-194aa was found to contain the DNA-binding SMAD1-MH1 domain that localized into the nucleus during myogenesis. Moreover, CircSmad1-194aa associates with the BMP-responsive element (BRE) in the Id1 promoter that is known to inhibit Myod1-driven myoblast differentiation. We propose that circSmad1-194aa promotes myogenesis by masking Id1-BRE from SMAD complex interaction, leading to suppression of ID1 expression and upregulation of MYOD1. Together, our findings identify circSmad1-194aa as a novel regulator of skeletal muscle differentiation and highlight the potential for the discovery of other functional circRNA-derived peptides in muscle pathophysiology. HIGHLIGHTSO_LIRNA-seq discovers hundreds of polysome-associated circRNAs in mouse C2C12 myoblasts C_LIO_LIProtein-coding circRNAs exhibit myogenesis-specific expression C_LIO_LICircSmad1 is abundant, methylated, and translated into a 194aa polypeptide C_LIO_LICircSmad1-encoded polypeptide promotes myogenesis C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=172 SRC="FIGDIR/small/708385v1_ufig1.gif" ALT="Figure 1"> View larger version (38K): org.highwire.dtl.DTLVardef@2074cforg.highwire.dtl.DTLVardef@d4fd98org.highwire.dtl.DTLVardef@1554735org.highwire.dtl.DTLVardef@1b6124a_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

Metabolomic, Lipidomic, and Enterohormone Changes in the Progression from MASLD to MASH

Background & AimsMetabolic Dysfunction-Associated Steatotic Liver Disease (MASLD) and Metabolic Dysfunction-Associated Steatohepatitis (MASH) represent progressive stages of liver disease, with distinct metabolic and cellular alterations. This study investigates the progression from MASLD to MASH through metabolomics, lipidomics, and assessment of hormones. MethodsMale C57BL/6NTac mice were fed a high-fat diet for 16 weeks to induce MASLD and for 29 weeks to develop MASH. Aged-matched controls on a normal diet were used for comparison. Histology confirmed the progression of MASLD to MASH. We performed metabolomic and lipidomic profiling of liver, colon, and stool samples to identify metabolic and lipid alterations. Plasma enteroendocrine hormones and cytokines were quantified. Immunofluorescence was performed to assess enteroendocrine cells changes in the colon and the association of serotonin (5-HT) with fibronectin in the liver. ResultsMetabolomic and lipidomic analysis revealed significant alterations at different stages of the disease. Specifically, cholic acid was increased across the liver, colon, and stool in both MASLD and MASH mice compared to controls. Compared to the control group, MASLD mice exhibited an increase in enteroendocrine hormones, GLP-1, GIP, and PYY, whereas no changes were observed in MASH mice. Comparing MASLD to MASH livers, we found hepatic 5-HT levels were increased in MASH mice compared to MASLD mice. The MASH liver also exhibited a colocalization between fibronectin and 5-HT, suggesting a potential role of 5-HT in liver fibrosis. ConclusionsOur study provides novel insights into the progressive metabolic and hormonal changes from MASLD to MASH. The increase in cholic acid and differential enteroendocrine hormone responses highlight the complex interactions between the gut and liver in metabolic liver diseases. These findings suggest that enteroendocrine hormones may play a role in the progression of MASLD to MASH as well as liver fibrosis, offering potential therapeutic avenues for targeting the gut-liver axis in metabolic liver diseases.

cell biology↗

bioRxiv2025

Bone Marrow Adipokine Mediates Hematopoietic Regeneration and Stem Cell Fitness

Bone marrow (BM) hematopoietic stem cells (HSCs) are exquisitely sensitive to cues from the BM microenvironment (ME), which is critical for their engraftment and regeneration following myeloablative stress. Retinoic acid signaling, acting on both HSCs and niche cells, has emerged as a central regulator of this process. Among ME components, BM adipocytes (BMAs), which can comprise up to 45% of BM volume and expand dramatically during the pancytopenic phase after myeloablation, play a previously underappreciated role in hematopoietic recovery. Here, we identify retinoid X receptor (RXR) signaling in BMAs as a key regulator of the adipokine Resistin, which promotes HSC self-renewal and functional fitness by activating NF-{kappa}B signaling. Conditional loss of RXR in adiponectin-expressing cells suppressed Resistin production, resulting in reduced NF-{kappa}B activity in HSCs, impaired self-renewal, and defective multilineage hematopoietic regeneration. Functionally, in vivo Resistin neutralization impaired hematopoietic reconstitution, whereas supplementation with either monomeric or dimeric Resistin enhanced HSC self-renewal and long-term lympho-hematopoietic reconstitution in an NF-{kappa}B-dependent manner. Together, these findings establish BMA-derived Resistin as an RXR-dependent, critical extrinsic regulator of HSC self-renewal and regenerative hematopoiesis, underscoring its essential role in lympho-myeloid reconstitution after myeloablation. Disclosures: The authors declare no relevant conflicts of interest.

physiology↗

bioRxiv2025

Dynamics of mound repair behavior in termites

Termites of the species Odontotermes obesus are found widely across the Indian peninsula. Their presence is visible due to the large over-ground mounds that they build across the landscape. These mounds house their entire colony, including major and minor workers, soldiers, the queen and king, and their brood. In addition, the termites also farm a specific variety of fungus within the mounds, which helps them break down the cellulose-rich food. The precise function of the mound architecture is the subject of some debate, and it has been argued that its structure serves physiological functions including thermoregulation or gas exchange for the entire colony. We hypothesized that if the mound structure indeed serves an important function, termites must be able to sense and repair any injury to their mound structure. To test this hypothesis, we created a hole in the mound wall and observed the dynamics of ensuing breach repair. The breach is quickly detected and repaired by the termites along a consistently sigmodial pattern involving the exponential recruitment of termites during the initial phases of repair, and a derecruitment of termites in the final phases. This behavior provides us with an assay to investigate various phenomena pertaining to mound-building, and to examine the sensory cues involved in detection and repair of the mound. From the basic considerations about the stimuli involved, we developed a logistic model to describe breach repair behavior, highlighting the critical role of a systemic cue in initiating this response. Through a series of experiments utilizing various behavioral assays, we identified the presence of light as this systemic cue that triggers mound repair. When light enters the mound, it triggers construction activity in termites, even though the major and minor workers responsible for repairs do not have image-forming eyes. This suggests that termites detect light through an extraocular mechanism

animal behavior and cognition↗

bioRxiv2025

ER-Localized Ceramide Accumulation Due to Disrupted CERT Transport Contributes to Replicative Senescence

Ceramides regulate diverse cellular processes through compartment-specific accumulation. While mitochondrial ceramide accumulation promotes apoptosis, its regulation and function during senescence remain incompletely understood. Here, we integrate lipidomics, transcriptomics, Raman spectroscopy and biochemical characterizations to define sphingolipid remodeling in replicative senescence. Senescent cells exhibit elevated ceramide levels and depletion of very-long-chain sphingomyelins, despite unaltered sphingomyelin synthase 1 expression, implicating impaired ceramide-sphingomyelin turnover. Pharmacological inhibition of ceramide transfer protein (CERT), the ER-to-Golgi ceramide transporter, phenocopies this sphingolipid remodeling and enhances senescence markers, suggesting disrupted ceramide trafficking as a driver of senescence. Raman spectroscopy suggests ceramide accumulation localized to the ER, with no enrichment at mitochondria, consistent with a non-apoptotic phenotype. Analysis of ER-enriched fractions confirm increased ceramide levels in ER fractions of senescent cells. Mechanistically, ceramide accumulation at the ER can contribute to ER stress, a key component of the senescence program. These findings identify altered ceramide trafficking as a contributor to ER stress and highlight ER-localized ceramide as a critical component of senescence-associated sphingolipid remodeling. SignificanceHow ceramides function during replicative senescence has remained poorly understood. In this study, we demonstrate that senescent cells accumulate ceramides at the endoplasmic reticulum (ER), revealing a fundamentally different mode of ceramide regulation and function in senescence. By integrating lipidomics, enzymatic assays, Raman-BCA imaging, and biochemical assays, we show that impaired ER-to-Golgi ceramide transport leads to ER-localized ceramide retention. Disrupting ceramide trafficking through inhibition of the ceramide transfer protein CERT phenocopies senescence-associated lipid remodeling and enhances senescence markers, establishing altered ceramide transport as a mechanistic contributor to this process. These findings link impaired ceramide trafficking to replicative senescence and suggest that ceramide function in senescence is linked to its retention at the ER.

cell biology↗

bioRxiv2024

Systemic inhibition of de novo purine biosynthesis prevents weight gain and improves metabolic health by increasing thermogenesis and decreasing food intake

ObjectiveObesity is a major health concern, largely because it contributes to type 2 diabetes mellitus (T2DM), cardiovascular disease, and various malignancies. Increase in circulating amino acids and lipids, in part due to adipose dysfunction, have been shown to drive obesity-mediated diseases. Similarly, elevated purines and uric acid, a degradation product of purine metabolism, are found in the bloodstream and in adipose tissue. These metabolic changes are correlated with metabolic syndrome, but little is known about the physiological effects of targeting purine biosynthesis. MethodsTo determine the effects of purine biosynthesis on organismal health we treated mice with mizoribine, an inhibitor of inosine monophosphate dehydrogenase 1 and 2 (IMPDH1/2), key enzymes in this pathway. Mice were fed either a low-fat (LFD; 13.5% kcal from fat) or a high-fat (HFD; 60% kcal from fat) diet for 30 days during drug or vehicle treatment. We ascertained the effects of mizoribine on weight gain, body composition, food intake and absorption, energy expenditure, and overall metabolic health. ResultsMizoribine treatment prevented mice on a HFD from gaining weight, but had no effect on mice on a LFD. Body composition analysis demonstrated that mizoribine significantly reduced fat mass but did not affect lean mass. Although mizoribine had no effect on lipid absorption, food intake was reduced. Furthermore, mizoribine treatment induced adaptive thermogenesis in skeletal muscle by upregulating sarcolipin, a regulator of muscle thermogenesis. While mizoribine-treated mice exhibited less adipose tissue than controls, we did not observe lipotoxicity. Rather, mizoribine-treated mice displayed improved glucose tolerance and reduced ectopic lipid accumulation. ConclusionsInhibiting purine biosynthesis prevents mice on a HFD from gaining weight, and improves their metabolic health, to a significant degree. We also demonstrated that the purine biosynthesis pathway plays a previously unknown role in skeletal muscle thermogenesis. A deeper mechanistic understanding of how purine biosynthesis promotes thermogenesis and decreases food intake may pave the way to new anti-obesity therapies. Crucially, given that many purine inhibitors have been FDA-approved for use in treating various conditions, our results indicate that they may benefit overweight or obese patients. HighlightsO_LIA purine biosynthesis inhibitor, mizoribine, protects against diet-induced weight gain C_LIO_LIMizoribine prevents fat mass gain in high-fat diet-fed male mice C_LIO_LIMizoribine reduces food intake and increases thermogenesis C_LIO_LIMizoribine induces expression of sarcolipin, a regulator of thermogenesis C_LIO_LIMizoribine treatment reduces ectopic lipids and increases glucose tolerance C_LI

physiology↗

bioRxiv2024

TGFb-ANGPT2-Tie2 axis in cancer-associated fibroblasts reprograms oral cancer cells to embryonic-like cell state with predictive significance of poor prognosis

Myofibroblastic cancer-associated fibroblasts (CAFs) in tumor stroma is identified as poor-prognostic indicator in oral cancer; however, biological mechanisms are largely unexplored. Here, we discovered the role of autocrine or exogenous transforming growth factor beta (TGF{beta}) in inducing Tunica Interna Endothelial cell kinase 2 (Tie2) -signaling through histone deacetylase-mediated downregulation of Tie2-antagonist, Angiopoietin-2 in CAFs, responsible for induction and maintenance of myofibroblastic differentiation. To understand the influence of CAF-specific Tie2-signaling on cancer cell properties, we performed CAF-Cancer cell co-culture and its single-cell RNA sequencing (scRNA-Seq). Distinct clustering of CAFs suggested their transcriptional heterogeneity, driven by TGF{beta}-Tie2 activation. Interestingly, CAF-specific Tie2-signaling was responsible to reprogram cancer cells, producing embryonic-like cell state with increased stemness and EMT signatures. Importantly, both the Tie2-specific gene expression signature as well as reprogrammed cancer cell specific gene expression modules were validated respectively in fibroblasts clusters and malignant cell clusters in two independent earlier reported scRNAseq studies of HNSCC tumors. Highlighting the translatability of our study, the gene expression signature derived from reprogrammed cancer cells showed significant association with poor prognosis in HNSCC patient of TCGA cohort. Pharmacological inhibition of Tie2-signaling in CAFs, significantly abrogated the tumor initiating ability of co-cultured oral cancer cell lines. Overall, combining our molecular and computational analysis, we may propose Tie2 as a novel factor responsible for CAF mediated cancer cell plasticity, associated with aggressive nature of oral cancer. TeaserTie2-signaling is activated in myofibroblasts which impacts the behaviour of malignant cells by inducing cancer cell plasticity to acquire stemness.

cancer biology↗

bioRxiv2024

Restoring hippocampal glucose metabolism rescues cognition across Alzheimer's disease pathologies

Impaired cerebral glucose metabolism is a pathologic feature of Alzheimer Disease (AD), and recent proteomic studies highlight a disruption of glial carbohydrate metabolism with disease progression. Here, we report that inhibition of indoleamine-2,3-dioxygenase 1 (IDO1), which metabolizes tryptophan to kynurenine (KYN) in the first step of the kynurenine pathway, rescues hippocampal memory function and plasticity in preclinical models of amyloid and tau pathology by restoring astrocytic metabolic support of neurons. Activation of IDO1 in astrocytes by amyloid-beta42 and tau oligomers, two major pathological effectors in AD, increases KYN and suppresses glycolysis in an AhR-dependent manner. Conversely, pharmacological IDO1 inhibition restores glycolysis and lactate production. In amyloid-producing APPSwe-PS1{Delta}E9 and 5XFAD mice and in tau-producing P301S mice, IDO1 inhibition restores spatial memory and improves hippocampal glucose metabolism by metabolomic and MALDI-MS analyses. IDO1 blockade also rescues hippocampal long-term potentiation (LTP) in a monocarboxylate transporter (MCT)-dependent manner, suggesting that IDO1 activity disrupts astrocytic metabolic support of neurons. Indeed, in vitro mass-labeling of human astrocytes demonstrates that IDO1 regulates astrocyte generation of lactate that is then taken up by human neurons. In co-cultures of astrocytes and neurons derived from AD subjects, deficient astrocyte lactate transfer to neurons was corrected by IDO1 inhibition, resulting in improved neuronal glucose metabolism. Thus, IDO1 activity disrupts astrocytic metabolic support of neurons across both amyloid and tau pathologies and in a model of AD iPSC-derived neurons. These findings also suggest that IDO1 inhibitors developed for adjunctive therapy in cancer could be repurposed for treatment of amyloid- and tau-mediated neurodegenerative diseases.

neuroscience↗

bioRxiv2024

MICOS Complex Loss Governs Age-Associated Murine Mitochondrial Architecture and Metabolism in the Liver, While Sam50 Dictates Diet Changes

Background & AimsAging is associated with a significant decline in mitochondrial function in the liver, leading to an increased risk of liver disease. This study examines age-related changes in the mitochondrial structure of human and murine livers using a combination of Serial Block-Face Scanning Electron Microscopy (SBF-SEM) and mass spectrometry approaches. MethodsThis study integrates mitochondrial structure analysis in a murine model with an analysis of liver architecture, lipogenesis, and genetically regulated gene expression in human cohorts. We explored the Mitochondrial Contact Site and Cristae Organizing System (MICOS) complex using SBF-SEM, three-dimensional reconstruction with Amira software, and mass spectrometry techniques. ResultsAging leads to a reduction in mitochondrial size and complexity, resulting in changes in the metabolomic and lipidomic profiles of murine liver cells that are comparable to those observed in aged human samples. We find that genetically modeled expression of MICOS complex genes OPA1 and CHCHD3 is associated with chronic liver disease phenotypes within a large biobank population. Furthermore, we observed dysregulated mitochondrial calcium handling and increased oxidative stress due to the disruption of the MICOS complex. ConclusionOur study highlights the age-associated decline in mitochondrial complexity and metabolic regulation within the aging murine liver and the human population. We have identified that these changes are partially attributable to the age-related loss of the MICOS complex. Impact and implicationsThis study offers new insights into the changes to mitochondrial ultrastructure that occur during aging. Using SBF-SEM, the quantification of young and aged murine mitochondrial structure was performed, which had previously been an underexplored avenue for measuring mitochondrial changes. The discovery of mitochondrial ultrastructural changes, in conjunction with measurements of age-associated metabolic alterations and gene association data, provides a model for how changes in MICOS expression may modulate age-related impairment of hepatic mitochondria. These results provide a new model by which changes in MICOS protein expression may both cause and be a potential therapeutic target for age-related impairment in hepatic function. HighlightsDecreased modeled expression of CHCHD3 in individuals of European genetic ancestry is linked to liver transplant and cirrhosis, while decreased modeled expression of OPA1 in individuals of African genetic ancestry is associated with chronic liver disease and cirrhosis. Aging alters liver lipid accumulation, MICOS mRNA levels, and disease markers. Aging reduces the volume and complexity of murine liver ultrastructure. Aging and diet significantly alter the MICOS complex in mice. Knockdown of Mic60 and Chchd6 lowers Ca2+ uptake, retention, and induces oxidative stress in HepG2 cells. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=139 SRC="FIGDIR/small/599846v3_ufig1.gif" ALT="Figure 1000"> View larger version (47K): org.highwire.dtl.DTLVardef@1cdd61corg.highwire.dtl.DTLVardef@a3fb74org.highwire.dtl.DTLVardef@1d1ad36org.highwire.dtl.DTLVardef@c2e55f_HPS_FORMAT_FIGEXP M_FIG C_FIG Liver aging causes metabolic, lipidomic, and mitochondrial structural alterations, reflecting age-dependent losses in the MICOS complex. Key components of the MICOS complex (MIC60, CHCHD3 and CHCHD6) are illustrated.

physiology↗

bioRxiv2024

The MICOS Complex Regulates Mitochondrial Structure and Oxidative Stress During Age-Dependent Structural Deficits in the Kidney

Due to aging, the efficiency of kidney function begins to decrease. Dysfunction in mitochondria and their cristae is a hallmark of aging. Therefore, age-related decline in kidney function could be attributed to changes in mitochondrial ultrastructure, increased reactive oxygen species, and alterations in metabolism and lipid composition. We sought to understand how mitochondrial ultrastructure is altered over time in tubular kidney cells. A serial block facing-scanning electron microscope and manual segmentation using the Amira software were employed to visualize murine kidney samples during the aging process at 3 months (young) and 2 years (old). We found that 2-year mitochondria are more fragmented with many uniquely shaped mitochondria observed across aging, concomitant with shifts in ROS, metabolomics, and lipid homeostasis. Furthermore, we demonstrate that the mitochondrial contact site and cristae organizing system (MICOS) complex is impaired in the kidney during aging. Disruption of the MICOS complex resulted in altered mitochondrial metabolic function and increased ROS levels. We found significant, detrimental structural changes in the mitochondria of aged kidney tubules, suggesting a potential mechanism underlying the increased frequency of kidney disease with aging. We hypothesize that disruption of the MICOS complex exacerbates mitochondrial dysfunction, creating a vicious cycle of mitochondrial degradation and oxidative stress, which impacts kidney health. Impact and ImplicationsDue to aging, the efficiency of kidney function begins to decrease, and the risk of kidney diseases may increase; however, the specific regulators of mitochondrial age-related changes are poorly understood. This study demonstrates that the MICOS complex may be a target for mitigating age-related mitochondrial changes. The MICOS complex is associated with oxidative stress and calcium dysregulation, which also arise in many kidney pathologies. HighlightsO_LIAging alters the MICOS mRNA levels and disease markers. C_LIO_LIAging reduces cristae architecture, mitochondrial volume and complexity in murine kidney ultrastructure C_LIO_LIReducing MIC60 and CHCHD6 lowers Ca2+ uptake and retention and induces oxidative stress in HEK cells. C_LIO_LIMetabolomic Profiling revealed that NAD+ and amino acid metabolism were altered in aged kidneys. C_LIO_LIMICOS deficiency alters the reduced basal, ATP-linked, maximal capacity and spare capacity. C_LIO_LIDecreased modeled expression of CHCHD6 in individuals of European genetic ancestry is linked to chronic kidney disease, whereas decreased modeled expression of OPA1 in individuals of African genetic ancestry is associated with chronic kidney disease. C_LI Graphical AbstractKidney aging causes a decline in the MICOS complex, concomitant with metabolic, lipidomic, and mitochondrial structural alterations. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/598108v3_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@1a868f0org.highwire.dtl.DTLVardef@1817bfborg.highwire.dtl.DTLVardef@1f2a1a3org.highwire.dtl.DTLVardef@520692_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2024

Impedance-based assay for pan-cancer early and rapid detection of cell-free DNA

Aberrant DNA methylation is a hallmark of cancer, and plasma cell-free DNA (cfDNA) containing these abnormal methylation patterns has emerged as a promising non-invasive biomarker for pan-cancer detection. However, intrinsic challenges remain that continue to limit its broad clinical application. Here we show a simple and rapid impedance biosensor called Asima Rev that can detect cancer cfDNA in under 5 min without the need for any molecular labelling, electrode modification, signal amplification, or target enrichment steps. Using 216 clinical samples (50 healthy) from 15 different cancer types (all stages) we show an overall sensitivity and specificity of 96.4% and 94.0%, respectively. Differences in methylation content between cancerous and healthy cfDNA lead to distinct solvation behaviour and electro-physicochemical property that remain consistent across cancer types regardless of the distribution patterns of methyl cytosine. Our test exploits this inherent difference.

biophysics↗

bioRxiv2024

Metabolite accumulation from oral NMN supplementation drives aging-specific kidney inflammation

The mitochondrial-rich renal tubule cells are key regulators of blood homeostasis via excretion and reabsorption of metabolic waste. With age, tubules are subject to increasing mitochondrial dysfunction and declining nicotinamide adenine dinucleotide (NAD+) levels, both hampering ATP production efficiency. We tested two mitochondrial interventions in young (6-mo) and aged (26-mo) adult male mice: (ELAM), a tetrapeptide in clinical trials that improves mitochondrial structure and function, and nicotinamide mononucleotide (NMN), an NAD+ intermediate and commercially available oral supplement. Kidneys were analyzed from young and aged mice after eight weeks of treatment with ELAM (3 mg/kg/day), NMN (300 mg/kg/day), or from aged mice treated with the two interventions combined (ELAM+NMN). We hypothesized that combining pharmacologic treatments to ameliorate mitochondrial dysfunction and boost NAD+ levels, would more effectively reduce kidney aging than either intervention alone. Unexpectedly, in aged kidneys, NMN increased expression of genetic markers of inflammation (IL-1{beta} and Ccl2) and tubule injury (Kim-1). Metabolomics of endpoint sera showed that NMN-treated aged mice had higher circulating levels of uremic toxins than either aged controls or young NMN-treated mice. ELAM+NMN- treated aged mice accumulated uremic toxins like NMN-only aged mice, but reduced IL-1{beta} and Ccl2 kidney mRNA. This suggests that pre-existing mitochondrial dysfunction in aged kidney underlies susceptibility to inflammatory signaling with NMN supplementation in aged, but not young, mice. These findings demonstrate age and tissue dependent effects on downstream metabolic accumulation from NMN and highlight the need for targeted analysis of aged kidneys to assess the safety of anti-aging supplements in older populations. Summary StatementDeclining levels of NAD+ and increasing mitochondrial dysfunction with age are functionally linked and are popular mechanistic targets of commercially available anti-aging therapeutics. Studies have focused on nicotinamide mononucleotide (NMN), nicotinamide riboside (NR) and nicotinamide (NAM) supplementation to boost cellular NAD+, but a consensus on the dosage and regimen that is beneficial or tolerable has not been reached. We show that although high levels of sustained NMN supplementation are beneficial to liver and heart in aged mice, the same dosing regimen carries age-associated signs of kidney inflammation. Our findings underscore a complex state of age- and tissue-specific metabolic homeostasis and raise questions not only about how much, and for how long, but at what age is NAD+ boosting safe.

physiology↗

bioRxiv2024

Experimental type 1 diabetes metabolically rejuvenates CD8+ T cells for improved control of tumor growth through an IGF1-IGF1R axis

AbstractEpidemiological studies suggest that patients with pre-existing type 1 diabetes (T1D) have a decreased risk of developing melanoma, prostate cancer, and breast cancer, although the underlying mechanism remains to be elucidated. In translational modelling, we observed that streptozotocin (STZ) induced T1D mice exhibited restricted melanoma and carcinoma (mammary, lung and colon) growth in association with extended overall survival. Tumor-infiltrating CD8+ T cells were found to be responsible for tumor growth restriction. Tumor infiltrating CD8+ T cells but not tumor cells themselves exhibited higher glycolytic and cytotoxic activities in T1D hosts. Such improved anti-tumor T cell function was linked to selective upregulated expression of insulin-like growth factor 1, insulin-like growth factor 1 receptor, and phospho-mTOR in CD8+ T cells in the TME. T1D patient derived CD8+ T cells displayed superior activation in vitro after tumor antigen stimulation vs. non-diabetic CD8+ T cells. Activation of T1D patient derived CD8+ T cells was sensitive to targeted antagonism of IGF1R and mTOR, supporting the operational involvement of the IGF1R-mTOR signaling axis. Our results suggest that selective activation of the intrinsic IGF1R-mTOR signaling axis in CD8+ T cells represents a preferred endpoint to achieving more effective immunotherapy outcomes and improved cancer patient management. SignificanceExperimental type 1 diabetes decelerates tumor growth through metabolic activation of cytotoxic T cells dependent on an IGF1R-mTOR signaling pathway. CD8+IGF1R+IGF1+ T cells play a crucial role in T1D dependent tumor control.

cancer biology↗

bioRxiv2024

Integrative single-cell analysis of neural stem/progenitor cells reveals epigenetically dysregulated interferon response in progressive multiple sclerosis

Progressive multiple sclerosis (PMS) is characterized by a primary smouldering pathological disease process associated with a superimposed inflammatory activity. Cellular and molecular processes sustaining the pathobiology of PMS remain to be identified. We previously discovered senescence signatures in neural stem/progenitor cells (NSCs) from people with PMS. Applying direct reprogramming to generate directly induced NSCs (iNSCs) from somatic fibroblasts, we retain epigenetic information and observe hypomethylation of genes associated with lipid metabolic processes and IFN signalling only in PMS lines. Single-cell/nucleus transcriptomic and epigenetic profiling reveal an inflammatory, senescent-like, IFN-responsive radial glia (RG)-like cell subcluster mainly in PMS iNSCs that is driven by IFN-associated transcription factors. Lastly, we identify a population of senescent, IFN-responsive, disease-associated RG-like cells (DARGs) in the PMS brain that share pseudotime trajectories with iNSCs in vitro. We describe the existence of a non-neurogenic, dysfunctional DARG population that has the potential to fuel smouldering inflammation in PMS.

neuroscience↗

bioRxiv2024

Increased Cholesterol Synthesis Drives Neurotoxicity in Patient Stem Cell-Derived Model of Multiple Sclerosis

Senescent neural progenitor cells have been identified in brain lesions of people with progressive multiple sclerosis (PMS). However, their role in disease pathobiology and contribution to the lesion environment remains unclear. By establishing directly induced neural stem/progenitor cell (iNSC) lines from PMS patient fibroblasts, we studied their senescent phenotype in vitro. Senescence was strongly associated with inflammatory signaling, hypermetabolism, and the senescence associated secretory phenotype (SASP). PMS-derived iNSCs displayed increased glucose-dependent fatty acid and cholesterol synthesis, which resulted in the accumulation of cholesteryl ester-enriched lipid droplets. An HMG-CoA reductase-mediated lipogenic state was found to induce secretion of the SASP in PMS iNSC conditioned media via transcriptional regulation by cholesterol-dependent transcription factors. SASP from PMS iNSCs induced neurotoxicity. Chemical targeting of HMG-CoA reductase using the cholesterol-lowering drug simvastatin (SV) prevented SASP release and resulting neurotoxicity. Our findings suggest a disease-associated, cholesterol-related, hypermetabolic phenotype of PMS iNSCs that leads to neurotoxic signaling and is rescuable pharmacologically.

neuroscience↗