Biology subjects

Witting, M.

Publications and source records attributed to Witting, M..

9 recordsLinked to original sources

bioRxiv2026

Plasma metabolite responses to an oral protein tolerance test differ between young and sarcopenic participants and suggest altered anabolic sensitivity

BackgroundSarcopenia is associated with anabolic resistance, a blunted muscle protein synthesis response to protein ingestion. Here, we hypothesized that anabolic resistance may be associated with a delayed postprandial decline in circulating plasma amino acids following protein ingestion. We therefore wanted to investigate whether an oral protein tolerance test (OPTT) combined with untargeted plasma metabolomics can detect age-related or sarcopenia-related differences in amino acid time courses consistent with altered postprandial amino acid handling, which could potentially reflect reduced anabolic sensitivity. Moreover, we investigated whether metabolites other than amino acids reacted to the OPTT. MethodsTwelve young healthy adults (controls: 22-28 years) and 12 older adults with clinically diagnosed probable or confirmed sarcopenia (70-91 years) ingested 20 g of whey protein after an overnight fast. We collected venous blood at baseline, 1 h, and 2 h post-ingestion and analyzed the samples by untargeted LC-HRMS plasma metabolomics. Linear mixed-effects models were fitted for 2,968 metabolic features with Benjamini-Hochberg FDR correction. For each category (branched-chain amino acid, essential amino acid [EAA], total amino acid) we summed the within-subject log2 fold changes (FC) of the constituent amino acids. This composite is reported as the summed log2FC Results201 metabolites were structurally annotated including 58 amino acid-related metabolites and 97 lipids. Fourteen of 17 proteinogenic amino acids increased significantly after protein ingestion (FDR<0.05). In young controls, essential amino acids rose more steeply at 1 h than in sarcopenic individuals (+10.06 {+/-} 1.05 vs. +7.84 {+/-} 1.58 summed log2FC) and declined more between 1 and 2 h (-4.93 {+/-} 1.29 vs. -0.20 {+/-} 2.27 summed log2FC). Leucine exemplified this pattern best, rising 1.74 log2FC in controls and declining to 0.96 at 2 h, while remaining elevated at 1.61 log2FC in the sarcopenic group at 2 h (p=0.009). Beyond amino acids, whey protein lowered circulating free fatty acids in both groups (FA 18:2, FA 18:1, FA 16:0; all FDR<0.05). Medium- and long-chain acylcarnitines (Car 8:0, Car14:2) declined postprandially in controls but remained elevated in sarcopenic individuals (p<0.05), suggesting altered postprandial lipid metabolism. ConclusionIn this proof-of-concept study, an OPTT showed that plasma EAAs declined more slowly from their postprandial peak in older adults with sarcopenia than in young adults, consistent with altered postprandial amino acid handling that may reflect anabolic resistance. Whey protein ingestion additionally modulates lipid and acylcarnitine metabolism in an age-dependent manner, suggesting broader alterations in postprandial metabolic regulation in older adults with sarcopenia.

physiology↗

bioRxiv2026

miR-940 suppresses ferroptosis by controlling expression of key regulatory genes

Ferroptosis is a form of regulated cell death that is characterized by iron-dependent lipid peroxidation. This process is regulated by specific metabolites, the lipid composition of the cells, redox-active iron, and antioxidant mechanisms. Although numerous regulators have been identified over the past decade, exploring other mechanisms, particularly from non-coding genomic regions, can build a thorough understanding of the multifaceted regulatory processes underlying ferroptosis. MicroRNAs (miRNAs) play a crucial role in gene regulation and cellular functions. Through a CRISPR KO screen, we identified miR-940 as a negative regulator of ferroptosis. Overexpression of miR-940 in several cell lines consistently suppressed ferroptosis induced by system xc- inhibition. Notably, multiple cancer patient cohorts with elevated miR-940 levels exhibit reduced survival. Integrated bioinformatic, transcriptomic, and proteomic analyses revealed that miR-940 decreases the expression of ACSL4, LPCAT3, DMT1, and NCOA4, and simultaneously increases levels of GPX4. Pharmacological inhibition of GPX4 attenuated the protective effect of miR-940, indicating that its primary anti-ferroptotic activity is mediated through GPX4. Overall, this gene rewiring is associated with reduced levels of redox-active iron and diminished lipid peroxidation, consistent with ferroptosis suppression. These findings suggest that miR-940 coordinates ferroptosis inhibition, which presents a novel regulatory layer for therapeutic exploration in susceptible cancers.

cell biology↗

bioRxiv2025

Methionine synthase reductase regulates heterochromatin independently of methionine synthesis through mitochondrial homeostasis

Metabolic enzymes can influence chromatin organization by modulating the availability of key metabolites, yet how specific metabolic reactions affect chromatin function remains poorly understood. Here, we show that in Caenorhabditis elegans, the methionine-cycle enzyme methionine synthase reductase (MTRR-l/MSR) regulates heterochromatin independently of methionine synthesis. Loss of MTRR-1, but not of the methionine synthase METR-l/MS, specifically reduces heterochromatic histone methylation, derepresses repetitive elements, and causes developmental delay. Multi-omics profiling revealed that mtrr-1 mutants activate transcriptional programs associated with mitochondrial stress and accumulate long-chain acylcamitines, indicating disrupted mitochondrial homeostasis. Functional assays confirmed altered mitochondrial respiration in mtrr-1 mutants, while genetic suppression of the PMK-3/MAPK mitochondrial retrograde signaling pathway partially restored repeat silencing. Consistently, direct perturbation of mitochondrial function was sufficient to induce heterochromatin derepression in wild type animals. Together, our results reveal a previously unrecognized mitochondria-to-chromatin axis controlled by the methionine-cycle enzyme MTRR-l/MSR, demonstrating that mitochondrial homeostasis is required for heterochromatin maintenance independently of canonical methionine metabolism.

cell biology↗

bioRxiv2024

LipiDetective - a deep learning model for the identification of molecular lipid species in tandem mass spectra

Lipids are involved in many vital processes within the cell, and alterations in lipid homeostasis have been associated with various diseases such as cancer or type 2 diabetes. Confidently identifying lipids in samples is a prerequisite for understanding the multiple functions lipids fulfill in health and disease. However, the accurate identification of molecular lipid species based on tandem mass spectrometry data is still a key challenge in lipidomics. Most current approaches rely on using a custom pipeline to process and match the measured spectra against an in-house spectra reference library, which hinders the comparability of results. To address this challenge, a transformer model called LipiDetective was developed and trained on a dataset composed of reference spectra measured from lipid standards, spectra from databases, and published experiments, utilizing both shotgun as well as liquid-chromatography mass spectrometry. LipiDetective demonstrates, for the first time, that artificial neural networks can learn the characteristic lipid fragmentation patterns to automatically and accurately annotate molecular lipids species in tandem mass spectra independently of the experimental setup. The model can even correctly predict lipid species for which it has never seen a spectrum before as it is able to generalize the learned lipid fragmentation patterns. Analysis of the integrated gradients reveals that LipiDetective focuses on relevant peaks that can be matched to known fragments and are thus humanly interpretable. Therefore, LipiDetective has the potential to be a valuable tool to aid in the lipid identification process and support the comparability of results from different sources. Aside from Lipidetective as a "ready-to-use" application, this work primarily offers a deeper understanding of how the model functions and how future deep learning models for lipid identification in mass spectra could be improved.

bioinformatics↗

bioRxiv2024

Polyketide synthase-derived sphingolipids determine microbiota-mediated protection against pathogens in C. elegans

Protection against pathogens is a major function of the gut microbiota. Although bacterial natural products have emerged as crucial components of host-microbiota interactions, their exact role in microbiota-mediated protection is largely unexplored. We addressed this knowledge gap with the nematode Caenorhabditis elegans and its microbiota isolate Pseudomonas fluorescens MYb115 that is known to protect against Bacillus thuringiensis (Bt) infection. We find that MYb115-mediated protection depends on sphingolipids that are derived from an iterative type I polyketide synthase (PKS), thereby describing a noncanonical pathway of bacterial sphingolipid production. We provide evidence that MYb115-derived sphingolipids affect C. elegans tolerance to Bt infection by altering host sphingolipid metabolism. This work establishes sphingolipids as structural outputs of bacterial PKS and highlights the role of microbiota-derived sphingolipids in host protection against pathogens.

molecular biology↗

bioRxiv2023

SIN-3 transcriptional coregulator maintains mitochondrial hormesis and polyamine flux

Mitochondrial function relies on the coordinated transcription of mitochondrial and nuclear genomes to assemble respiratory chain complexes. Across species, the SIN3 coregulator influences mitochondrial functions, but how its loss impacts mitochondrial homeostasis and metabolism in the context of a whole organism is unknown. Exploring this link is important because SIN3 haploinsufficiency causes intellectual disability/autism syndromes and SIN3 plays an important role in tumor biology. Here we show that loss of C. elegans SIN-3 results in transcriptional deregulation of mitochondrial- and nuclear encoded mitochondrial genes, potentially leading to mito-nuclear imbalance. Consistent with impaired mitochondrial function, sin-3 mutants show extensive mitochondrial fragmentation by transmission electron microscopy (TEM) and in vivo imaging, and altered oxygen consumption. Metabolomic analysis of sin-3 mutant animals identifies a signature of mitochondria stress, and deregulation of methionine flux resulting in decreased S-adenosyl methionine (SAM), and increased polyamine levels. Our results identify SIN3 as a key regulator of mitochondrial dynamics and metabolic flux, with important implications for human pathologies.

developmental biology↗

bioRxiv2021

Assigning confidence to structural annotations from mass spectra with COSMIC

Untargeted metabolomics experiments rely on spectral libraries for structure annotation, but these libraries are vastly incomplete; in silico methods search in structure databases but cannot distinguish between correct and incorrect annotations. As biological interpretation relies on accurate structure annotations, the ability to assign confidence to such annotations is a key outstanding problem. We introduce the COSMIC workflow that combines structure database generation, in silico annotation, and a confidence score consisting of kernel density p-value estimation and a Support Vector Machine with enforced directionality of features. In evaluation, COSMIC annotates a substantial number of hits at small false discovery rates, and outperforms spectral library search for this purpose. To demonstrate that COSMIC can annotate structures never reported before, we annotated twelve novel bile acid conjugates; nine structures were confirmed by manual evaluation and two structures using synthetic standards. Second, we annotated and manually evaluated 315 molecular structures in human samples currently absent from the Human Metabolome Database. Third, we applied COSMIC to 17,400 experimental runs and annotated 1,715 structures with high confidence that were absent from spectral libraries.

bioinformatics↗

bioRxiv2020

HLH-30 dependent rewiring of metabolism during starvation in C. elegans

One of the most fundamental challenges for all living organisms is to sense and respond to alternating nutritional conditions in order to adapt their metabolism and physiology to promote survival and achieve balanced growth. Here, we applied metabolomics and lipidomics to examine temporal regulation of metabolism during starvation in wildtype Caenorhabditis elegans and in animals lacking the transcription factor HLH-30. Our findings show for the first time that starvation alters the abundance of hundreds of metabolites and lipid species in a temporal- and HLH-30-dependent manner. We demonstrate that premature death of hlh-30 animals under starvation can be prevented by supplementation of exogenous fatty acids, and that HLH-30 is required for complete oxidation of long-chain fatty acids. We further show that RNAi-mediated knockdown of the gene encoding carnitine palmitoyl transferase I (cpt-1) only impairs survival of wildtype animals and not of hlh-30 animals. Strikingly, we also find that compromised generation of peroxisomes by prx-5 knockdown renders hlh-30 animals hypersensitive to starvation, which cannot be rescued by supplementation of exogenous fatty acids. Collectively, our observations show that mitochondrial functions are compromised in hlh-30 animals and that hlh-30 animals rewire their metabolism to largely depend on functional peroxisomes during starvation, underlining the importance of metabolic plasticity to maintain survival. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/170555v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@a7339borg.highwire.dtl.DTLVardef@12d36b0org.highwire.dtl.DTLVardef@1615d6aorg.highwire.dtl.DTLVardef@13bfaa8_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2019

In-vivo targeted tagging of RNA isolates cell specific transcriptional responses to environmental stimuli and identifies liver-to-adipose RNA transfer

Bio-fluids contain various circulating cell-free RNA transcripts (ccfRNAs). The composition of these ccfRNAs varies between bio-fluids and constitute tantalizing biomarker candidates for several pathologies. ccfRNAs have also been demonstrated as mediators of cellular communication, yet little is known about their function in physiological and developmental settings and most works are limited to in-vitro studies. Here, we have developed iTAG-RNA, a novel method for the unbiased tagging of RNA transcripts in mice in-vivo. We used this method to isolate hepatocytes and kidney proximal epithelial cells-specific transcriptional response to a dietary challenge without interfering with the tissue architecture, and to identify multiple hepatocyte-secreted ccfRNAs in plasma. We also identified transfer of these hepatic derived ccfRNAs to adipose tissue, where they likely serve as a buffering mechanism to maintain cholesterol and lipid homeostasis. Our findings directly demonstrate in-vivo transfer of RNAs between tissues and highlight its implications for endocrine signaling and homeostasis.

molecular biology↗