Constitutive activation of hypoxia-inducible factor 1 (HIF-1) in vhl-1(ok161) mutant Caenorhabditis elegans induces a characteristic phenotype marked by extended lifespan, reduced body size, and broad metabolic reprogramming. However, the underlying determinants mediating these adaptations remain incompletely understood. Here, we show that HIF-1 activation drives a distinct sulfur metabolic state characterized by the accumulation of hydrogen sulfide (H₂S) and persulfide species, accompanied by extensive remodeling of the mitochondrial proteome and altered stress sensitivity. Strikingly, these organismal and metabolic phenotypes require the mitochondrial sulfide:quinone oxidoreductase SQRD-1 (SQOR), which catalyzes the first step of sulfide oxidation. Loss of sqrd-1 abolishes the lifespan extension and small-body phenotype of vhl-1 mutants and reverses their proteomic, mitochondrial, and metabolic profiles toward wild-type levels. Mechanistically, SQRD-1-dependent sulfide oxidation establishes a redox state that shapes mitochondrial function, including suppression of respiratory capacity and modulation of oxidative stress responses. Consistent with this, quantitative proteomics revealed coordinated regulation of mitochondrial pathways and sulfur oxidation enzymes in vhl-1 mutants that is lost in vhl-1;sqrd-1 double mutants. Functional analyses identified mitochondrial complex I subunits (NUO-4 and NUO-5) as downstream effectors linking this metabolic state to lifespan and body-size regulation. Together, these findings identify SQRD-1 as a critical determinant of the metabolic and physiological outcomes of chronic HIF-1 activation, defining a sulfide-dependent redox state that couples hypoxia signaling to mitochondrial function and organismal adaptation. This work highlights mitochondrial sulfide oxidation as a key regulatory node in hypoxia responses and a potential target for modulating stress resistance and metabolic health.

Kidney transplantation (KTx) is the preferred treatment for kidney failure. However, post-transplant management is challenging due to the limited lifespan of transplanted organs. Current methods for monitoring post-transplant complications are invasive and have limitations. Therefore, there is an urgent need for novel non-invasive biomarkers. This study investigates the proteomic composition of urine to understand renal biology during the process of transplantation and to identify potential markers for outcome prediction. Urine samples were collected from donors before transplantation and from recipients 4 weeks and 1 year after transplantation. Proteomic analysis was performed using mass spectrometry and label-free quantification. Statistical analyses included principal component analysis (PCA) and enrichment analysis. The resulting key findings were confirmed in an independent validation cohort. In addition, correlative regression models to evaluate the relationship between protein abundance and clinical outcomes in the further course after transplantation were performed. 106 urine samples in the setting of 70 kidney transplantations were analyzed. PCA revealed distinct clustering of donor and recipient samples, indicating significant proteomic changes after transplantation. Hierarchical clustering and gene ontology analysis identified molecular changes as a response to transplantation and showed an over-representation of relevant pathways related to inflammation, cell immune response and coagulation in both the original and validation cohorts. Multivariate regression analysis, including linear and logistic regression, identified 11 potential protein biomarkers, including ORM2, IL1RAP, APP, and FABP4 as predictors of eGFR 12 months after transplantation and 1 HP as a predictor of infections within the first year after transplantation, respectively. This study underscores the potential of noninvasive urine proteomics for identifying biological processes involved in kidney transplantation and for enhancing post-transplant monitoring and outcome prediction. We identified 12 potential biomarkers with added value to standard clinical parameters linked to transplant outcomes, which will be promising candidates for future outcome monitoring after KTx.

Aging reduces cellular resilience and increases susceptibility to organ injury, notably acute kidney injury (AKI). Ischemia-reperfusion injury (IRI) influences outcomes after kidney transplantation. In animal models, short-term calorie restriction (CR) extends lifespan and protects kidneys from IRI, but translation to patients is limited due to incomplete mechanistic insight. This study examined clinical and molecular effects of short-term CR in living kidney donors. Twelve donors were alternately assigned to CR or an ad libitum diet. CR participants consumed a formula diet providing 50% of individual caloric needs for seven days before donation. Clinical parameters and biosamples from perirenal fat, renal arteries, ureters, kidney biopsies, blood, and urine were collected. Primary outcomes were CR-induced molecular changes; clinical outcomes were secondary. CR was well tolerated and caused significant weight loss without affecting AKI incidence or increasing adverse events. Lipidomic and proteomic analyses showed enhanced lipolysis and proteostasis, reduced insulin signaling, sex-specific effects, and decreased inflammatory factors in donor arteries and ureter tissue. This hypothesis-generating study indicates that short-term CR before donation is feasible and suggests that CR promotes organ protection in humans by dampening insulin signaling and inflammation, providing a basis for future targeted interventions in clinical and transplant research moving forward.

Real-life medical data usage is currently limited by lack of structured databases and analysis tools. Autosomal dominant polycystic kidney disease (ADPKD) remains the most common monogenic cause of kidney failure. We present MEDA-PKD, an interactive, real-time, web-based analytics platform that integrates clinical routine with research data from a large, multicenter ADPKD cohort into a harmonized database (https://shiny.cecad.uni-koeln.de/ADPKD_registry/). Importantly, MEDA-PKD provides automated real-time analysis and dynamic visualization using ShinyApps, replacing traditional static approaches to clinical cohorts. As of March 2026, MEDA-PKD visualized data from 1,735 patients (mean follow-up 796 days). Analysis of data from patients initiating tolvaptan therapy demonstrate the platform’s capacity to evaluate therapeutic impact in real time. Other tabs include family history, medication, extrarenal manifestations, lifestyle, quality of life and a wide array of lab values (20,894 entries). Besides, MEDA-PKD integrates proteomics data with deep clinical phenotyping and allows for secure individual patient-level data access to participating centers. MEDA-PKD represents a paradigm shift by transforming static cohort data into a living, actionable knowledge resource providing real-time insight in ADPKD and will serve as a template for other diseases.

BACKGROUND:RNA-binding proteins (RBPs) are fundamental regulators of cellular biology that affect all steps in the generation and processing of RNA molecules. Recent evidence suggests that regulation of RBPs that modulate both RNA stability and translation may have a profound effect on the proteome. However, regulation of RBPs in clinically relevant experimental conditions has not been studied systematically.METHODS:We used RNA interactome capture, a method for the global identification of RBPs to characterize the global RNA-binding proteome (RBPome) associated with polyA-tailed RNA species in murine ciliated epithelial cells of the inner medullary collecting duct. To study regulation of RBPs in a clinically relevant condition, we analyzed hypoxia-associated changes of the RBPome.RESULTS:We identified >1000 RBPs that had been previously found using other systems. In addition, we found a number of novel RBPs not identified by previous screens using mouse or human cells, suggesting that these proteins may be specific RBPs in differentiated kidney epithelial cells. We also found quantitative differences in RBP-binding to mRNA that were associated with hypoxia versus normoxia.CONCLUSIONS:These findings demonstrate the regulation of RBPs through environmental stimuli and provide insight into the biology of hypoxia-response signaling in epithelial cells in the kidney. A repository of the RBPome and proteome in kidney tubular epithelial cells, derived from our findings, is freely accessible online, and may contribute to a better understanding of the role of RNA-protein interactions in kidney tubular epithelial cells, including the response of these cells to hypoxia.

The cellular response to hypoxia is crucial to organismal survival, and hypoxia-inducible factors (HIF) are the key mediators of this response. HIF-signaling is central to many human diseases and mediates longevity in the nematode. Despite the rapidly increasing knowledge on RNA-binding proteins (RBPs), little is known about their contribution to hypoxia-induced cellular adaptation. We used RNA interactome capture (RIC) in wild-type Caenorhabditis elegans and vhl-1 loss-of-function mutants to fill this gap. This approach identifies more than 1,300 nematode RBPs, 270 of which can be considered novel RBPs. Interestingly, loss of vhl-1 modulates the RBPome. This difference is not primarily explained by protein abundance suggesting differential RNA-binding. Taken together, our study provides a global view on the nematode RBPome and proteome as well as their modulation by HIF-signaling. The resulting RBP atlas is also provided as an interactive online data mining tool (http://shiny.cecad.uni-koeln.de:3838/celegans_rbpome).

AbstractThe extracellular matrix (ECM) is a pivotal three-dimensional network crucial for tissue organization, cellular communication, and fundamental cellular processes, where collagens are the major chemical entity in amount. ECM deregulation is directly involved with several pathologies, such as tumour growth and invasiveness, atherosclerosis, and diabetic nephropathy. Mutations in the von Hippel-Lindau tumour suppressor (pVHL) cause VHL syndrome, a multi-tumour syndrome commonly associated with clear cell renal carcinoma (ccRCC). Loss of pVHL is associated with the activation of hypoxia-inducible factor (HIF) signaling. Mutation of VHL-1 in the nematodeCanorhabditis eleganshas been shown to increase lifespan and stress resistance. Interestingly, considering recent findings on the involvement of collagens in the regulation of lifespan, we also observed these animals to show defects in body morphology in a HIF-1 dependent manner. Based on this finding, we established a link between HIF-1 activation upon loss of VHL-1 and ECM defects associated with alterations in collagen expression. An RNAi screen examining genes upregulated invhl-1mutant worms revealed the sulfide quinone oxidoreductasesqrd-1to mediate the change in body morphology. SQRD-1 is essential to the HIF-1 dependent increase in several collagen genes. One of these genes,col-88, partly mediates both the impact of loss of VHL-1 on lifespan extension and body length. The downregulation of the uncharacterisedcol-88partially restores lifespan extension and reduces body size ofvhl-1/sqrd-1tovhl-1(ok161)single mutant. This study contributes to the increasing body of evidence linking lifespan extension and the ECM and now implicates this axis in hypoxia-signaling. These findings are of special interest considering the role of ECM integrity in tumour growth and metastasis.Author SummaryThe extracellular matrix and its composing collagens are associated with a wide number of diseases, including cancer. The von Hippel-Lindau tumour suppressor (pVHL) is known to work by regulating the Hypoxia Inducible Factor (HIF) to help the organism to adapt to lack of oxygen. Mutations in pVHL are associated with clear cell renal carcinoma (ccRCC). Interestingly, a small number of studies have shown that pVHL can be directly associated with collagens, a function that is independent of its classical role regulating HIF. However, there is no further knowledge about which role the hypoxia pathway has when it comes to extracellular matrix formation and function, what would be useful since the invasiveness of cancers, such as ccRCC, are directly connected to their matrix/collagen composition. Here we observed that the model organismC. eleganshas drastically different collagen composition and body size upon a mutation on itsvhl-1gene. Furthermore, a protein previously only known to be involved in sulfide metabolism, SQRD-1, connects body size and lifespan in this animal model, revealing a surprising link between the hypoxia pathway and sulfur metabolism to control lifespan. Further studies could target sulfur metabolism in ccRCC to modulate collagen production and tumour invasiveness.

Chronic Kidney Disease (CKD), a global health burden, is strongly associated with age-related renal function decline, hypertension, and diabetes, which are all frequent consequences of obesity. Despite extensive studies, the mechanisms determining susceptibility to CKD remain insufficiently understood. Clinical evidence together with prior studies from our group showed that perinatal metabolic disorders after intrauterine growth restriction or maternal obesity adversely affect kidney structure and function throughout life. Since obesity and aging processes converge in similar pathways we tested if perinatal obesity caused by high-fat diet (HFD)-fed dams sensitizes aging-associated mechanisms in kidneys of newborn mice. The results showed a marked increase of γH2AX-positive cells with elevated 8-Oxo-dG (RNA/DNA damage), both indicative of DNA damage response and oxidative stress. Using unbiased comprehensive transcriptomics we identified compartment-specific differentially-regulated signaling pathways in kidneys after perinatal obesity. Comparison of these data to transcriptomic data of naturally aged kidneys and prematurely aged kidneys of genetic modified mice with a hypomorphic allele of Ercc1, revealed similar signatures, e.g., inflammatory signaling. In a biochemical approach we validated pathways of inflammaging in the kidneys after perinatal obesity. Collectively, our initial findings demonstrate premature aging-associated processes as a consequence of perinatal obesity that could determine the susceptibility for CKD early in life.

SIGNIFICANCE STATEMENT:AKI is a major clinical complication leading to high mortality, but intensive research over the past decades has not led to targeted preventive or therapeutic measures. In rodent models, caloric restriction (CR) and transient hypoxia significantly prevent AKI and a recent comparative transcriptome analysis of murine kidneys identified kynureninase (KYNU) as a shared downstream target. The present work shows that KYNU strongly contributes to CR-mediated protection as a key player in the de novo nicotinamide adenine dinucleotide biosynthesis pathway. Importantly, the link between CR and NAD+ biosynthesis could be recapitulated in a human cohort. BACKGROUND:Clinical practice lacks strategies to treat AKI. Interestingly, preconditioning by hypoxia and caloric restriction (CR) is highly protective in rodent AKI models. However, the underlying molecular mechanisms of this process are unknown. METHODS:Kynureninase (KYNU) knockout mice were generated by Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and comparative transcriptome, proteome and metabolite analyses of murine kidneys pre- and post-ischemia-reperfusion injury in the context of CR or ad libitum diet were performed. In addition, acetyl-lysin enrichment and mass spectrometry were used to assess protein acetylation. RESULTS:We identified KYNU as a downstream target of CR and show that KYNU strongly contributes to the protective effect of CR. The KYNU-dependent de novo nicotinamide adenine dinucleotide (NAD+) biosynthesis pathway is necessary for CR-associated maintenance of NAD+ levels. This finding is associated with reduced protein acetylation in CR-treated animals, specifically affecting enzymes in energy metabolism. Importantly, the effect of CR on de novo NAD+ biosynthesis pathway metabolites can be recapitulated in humans. CONCLUSIONS:CR induces the de novo NAD+ synthesis pathway in the context of IRI and is essential for its full nephroprotective potential. Differential protein acetylation may be the molecular mechanism underlying the relationship of NAD+, CR, and nephroprotection.

Mycophenolate Mofetil (MMF) has an established role as a therapeutic agent in childhood nephrotic syndrome. While other immunosuppressants have been shown to positively affect podocytes, direct effects of MMF on podocytes remain largely unknown. The present study examines the effects of MMF’s active component Mycophenolic Acid (MPA) on the transcriptome of podocytes and investigates its biological significance. We performed transcriptomics in cultured murine podocytes exposed to MPA to generate hypotheses on podocyte-specific effects of MPA. Accordingly, we further analyzed biological MPA effects on actin cytoskeleton morphology after treatment with bovine serum albumin (BSA) by immunofluorescence staining, as well as on cell survival following exposure to TNF-α and cycloheximide by neutral red assay. MPA treatment significantly (adjusted p < 0.05) affected expression of 351 genes in podocytes. Gene Ontology term enrichment analysis particularly clustered terms related to actin and inflammation-related cell death. Indeed, quantification of the actin cytoskeleton of BSA treated podocytes revealed a significant increase of thickness and number of actin filaments after treatment with MPA. Further, MPA significantly reduced TNFα and cycloheximide induced cell death. MPA has a substantial effect on the transcriptome of podocytes in vitro, particularly including functional clusters related to non-immune cell dependent mechanisms. This may provide a molecular basis for direct beneficial effects of MPA on the structural integrity and survival of podocytes under pro-inflammatory conditions.

Caloric Restriction (CR) extends lifespan and augments cellular stress-resistance from yeast to primates, making CR an attractive strategy for organ protection in the clinic. Translation of CR to patients is complex, due to problems regarding adherence, feasibility, and safety concerns in frail patients. Novel tailored dietary regimens, which modulate the dietary composition of macro- and micronutrients rather than reducing calorie intake promise similar protective effects and increased translatability. However, a direct head-to-head comparison to identify the most potent approach for organ protection, as well as overlapping metabolic consequences have not been performed. We systematically analyzed six dietary preconditioning protocols - fasting mimicking diet (FMD), ketogenic diet (KD), dietary restriction of branched chained amino acids (BCAA), two dietary regimens restricting sulfur-containing amino acids (SR80/100) and CR - in a rodent model of renal ischemia-reperfusion injury (IRI) to quantify diet-induced resilience in kidneys. Of the administered diets, FMD, SR80/100 and CR efficiently protect from kidney damage after IRI. Interestingly, these approaches show overlapping changes in oxidative and hydrogen sulfide (H2S)-dependent cysteine catabolism as a potential common mechanism of organ protection.

Acute kidney injury is a frequent complication in the clinical setting and associated with significant morbidity and mortality. Preconditioning with short-term caloric restriction is highly protective against kidney injury in rodent ischemia reperfusion injury models. However, the underlying mechanisms are unknown hampering clinical translation. Here, we examined the molecular basis of caloric restriction-mediated protection to elucidate the principles of kidney stress resistance. Analysis of an RNAseq dataset after caloric restriction identified Cyp4a12a, a cytochrome exclusively expressed in male mice, to be strongly downregulated after caloric restriction. Kidney ischemia reperfusion injury robustly induced acute kidney injury in male mice and this damage could be markedly attenuated by pretreatment with caloric restriction. In females, damage was significantly less pronounced and preconditioning with caloric restriction had only little effect. Tissue concentrations of the metabolic product of Cyp4a12a, 20-hydroxyeicosatetraenoic acid (20-HETE), were found to be significantly reduced by caloric restriction. Conversely, intraperitoneal supplementation of 20-HETE in preconditioned males partly abrogated the protective potential of caloric restriction. Interestingly, this effect was accompanied by a partial reversal of caloric restriction-induced changes in protein but not RNA expression pointing towards inflammation, endoplasmic reticulum stress and lipid metabolism. Thus, our findings provide an insight into the mechanisms underlying kidney protection by caloric restriction. Hence, understanding the mediators of preconditioning is an important prerequisite for moving towards translation to the clinical setting.

JADE family proteins (JADE1/2/3) have been implicated in diverse cellular functions and signaling pathways ranging from WNT signaling and cell cycle control to cell death and complex transcriptional regulation through histone acetyl-transferase complexes. JADE proteins show a high degree of sequence similarity and share two PHD zinc finger domains. JADE1 interacts with cilia-associated proteins and has been implicated in cilia-related genetic disorders with kidney phenotypes. However, the function of the widely expressed JADE proteins at the molecular level is still elusive. Here we show that JADE proteins regulate proteasome abundance and activity. Using kidney cells as a model, we demonstrate that loss of either JADE protein resulted in increased expression of almost all components of the 26S proteasome. Regulation occurred at the post-translational level and was not the consequence of transcriptional activation. Consistent with a role for JADE proteins in regulating overall proteasomal abundance, proteasomal activity was elevated in Jade-deficient cells, while exogenous expression of JADE1/2/3 decreased the level of proteasome activity. Coimmunoprecipitation experiments confirmed the interaction of proteasomal subunits with Jade1 suggesting a direct role of JADE proteins in regulating turnover, stability and abundance of the 26s proteasome. These data may now explain the plethora of cellular roles that have been attributed to JADE proteins.