Biology subjects

Helm, M.

Publications and source records attributed to Helm, M..

At least 19 recordsLinked to original sources

bioRxiv2026

Harmonized nucleoside mass spectrometry enables reproducible cross-platform RNA modification quantification

RNA modification analysis by LC-MS/MS is central to epitranscriptomics, yet quantitative comparison across laboratories and instrument platforms remains poorly standardized. Here, we performed a community-driven benchmarking study during the first Human RNome Project workshop to systematically evaluate cross-platform reproducibility of ribonucleoside mass spectrometry workflows. Using the same analytical column and gradient, standardized RNA samples, and shared reagents, we compared nucleoside quantification across quadrupole, time-of-flight, and orbitrap-based LC-MS platforms employing distinct acquisition strategies. While chromatographic separation was highly reproducible across systems, nucleoside-specific MS response behavior differed substantially between platforms and limited direct comparability of relative signal intensities. These response differences varied across analytes and concentration ranges, demonstrating that harmonized chromatography alone is insufficient for transferable quantitative analysis. Stable isotope-labeled internal standard (SILIS) normalization substantially reduced platform-and method-dependent response and improved agreement for most evaluated modifications. External calibration improved agreement between qTOF and Orbitrap workflows for a subset of modifications but did not fully resolve residual intersystem differences. Based on these findings, we establish benchmark-derived recommendations for harmonized relative and absolute RNA modification quantification, including guidance for calibration design, quality control, and data reporting. Together, this work provides a methodological framework for reproducible nucleoside LC-MS/MS workflows and establishes a foundation for large-scale comparative epitranscriptomic studies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=107 SRC="FIGDIR/small/739095v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@6e4e98org.highwire.dtl.DTLVardef@1ea13beorg.highwire.dtl.DTLVardef@b65858org.highwire.dtl.DTLVardef@1623337_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2026

DModE: An end-to-end framework for Differential Modification and Expression Analysis of Nanopore direct RNA sequencing data

SummaryNanopore direct RNA sequencing (DRS) enables simultaneous quantification of transcript abundance and RNA modifications from native RNA molecules, providing a unique opportunity to study transcriptional and epitranscriptomic regulation within a single experiment. However, comprehensive analysis of DRS data remains challenging, as existing workflows typically focus on individual processing steps and often require manual integration of multiple software packages for expression analysis, modification detection, statistical testing, and visualization. Furthermore, integrated differential expression and differential RNA modification analysis at both gene and isoform resolution remains poorly supported by current workflows. Here, we present DModE (Differential Modification and Expression Analysis), an end-to-end framework for integrated analysis of Nanopore DRS data. DModE combines an Epi2ME-compatible Nextflow preprocessing workflow with a dedicated Python package for downstream statistical analysis, visualization, and reporting. The framework supports differential gene and isoform expression analysis, differential RNA modification analysis at genome and transcript level, metagene profiling, exploratory epitranscriptomic analyses, and integrated assessment of relationships between expression and modification dynamics. Results are automatically summarized in interactive HTML reports, facilitating reproducible and accessible data interpretation. By integrating transcriptomic and epitranscriptomic analyses within a single framework, DModE substantially simplifies comprehensive DRS data analysis and lowers the barrier for studying RNA modification biology using Nanopore sequencing. Availability and implementationThe DModE Preprocessing pipeline is available on GitHub (https://github.com/johannesmiedema/DModE-preprocessing). The Python package can be installed via pip and is also available on GitHub (https://github.com/johannesmiedema/dmode).

bioinformatics↗

bioRxiv2026

Sex-specific remodeling of the tRNA epitranscriptome in Alzheimers disease

tRNA modifications are critical regulators of RNA stability, decoding fidelity, and cellular stress adaptation, yet their contribution to human neurodegenerative disease remains poorly understood. Beyond their established functions in translational control, emerging evidence shows that RNA modifications influence neurogenesis, neurodevelopment, neuronal function, brain-cell differentiation, and cellular plasticity. Consequently, dysregulation of these molecular processes is increasingly recognized as a mechanistic contributor to neurodegenerative disorders. Alzheimers disease (AD), characterized by amyloid pathology, synaptic dysfunction, and progressive neuronal loss, has recently been linked to disturbances in RNA metabolism, suggesting that alterations in the epitranscriptome may represent an underexplored dimension of AD pathophysiology. Here, we systematically profiled the tRNA epitranscriptome across cellular and animal models of AD, as well as in human postmortem brain tissue from non-demented controls and AD patients, using liquid chromatography-tandem mass spectrometry (LC-MS/MS). This method enables highly sensitive quantification of RNA modifications, with limits of detection in the low femtomole range. Across our models, we identified a conserved yet sex-specific remodeling of the tRNA modification landscape in AD. Because therapeutic options and early diagnostic tools for AD remain limited, we leveraged these findings to develop a tRNA-centered RNA-modification score that integrates both nucleobase-specific modification patterns and neuropathological disease severity into a quantitative metric. Together, our findings identify the tRNA epitranscriptome as a unifying molecular sex-specific signature of AD, linking disease pathology and sex to impaired RNA metabolism. This line of research opens a new path toward establishing early biomarkers or diagnostic tools for AD. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/715648v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@174c203org.highwire.dtl.DTLVardef@6d610aorg.highwire.dtl.DTLVardef@fddeecorg.highwire.dtl.DTLVardef@136700e_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Complete post-transcriptional modification profiles in individual Staphylococcus aureus tRNA species

Transfer RNAs play a critical role in protein synthesis by matching mRNA codons to their corresponding amino acids. Their post-transcriptional modifications, shaping structure, stability, and codon decoding, are now recognized as key regulators of translation and cellular adaptation, including in bacterial pathogens. Here, we provide a comprehensive analysis of tRNA modifications in Staphylococcus aureus using extensive oligonucleotide mass spectrometry and deep-sequencing methods, generating a high-confidence modification map for each individual tRNA species, including the non-proteogenic tRNAGly. While the overall tRNA modification landscape is conserved among Gram-positive bacteria, our data uncovered unexpected S. aureus-specific features. These include the absence of [m2A]37 in tRNAs despite the presence of the methyltransferase RlmN, a single multi-site DusB2 enzyme catalyzing all tRNA dihydrouridylation, and a dedicated pseudouridine synthase responsible for [{Psi}]32 formation. Besides, heterogenous modification patterns were observed in tRNALeu(UAA) and tRNALys(CAA), highlighting a complex interplay in anticodon hypermodification. Integration of ribosome profiling and Nanopore tRNA sequencing offered a global view of the decoding properties of the reduced S. aureus tRNA set, revealing efficient four-way wobble recognition, slower translation of rare codons by low abundant tRNAs, and distinctive decoding dynamics of Gly codons potentially influenced by the unusual modification status of tRNAGly(UCC). This work establishes a framework for future research aimed at dissecting the role of specific tRNA modifications in S. aureus physiology and pathogenesis.

molecular biology↗

bioRxiv2025

Functional Characterization of Zymoseptoria tritici Candidate Effectors Reveals Their Role in Modulating Immunity in Nicotiana benthamiana

Zymoseptoria tritici is a significant wheat pathogen responsible for Septoria tritici blotch (STB) disease. Despite its economic impact, understanding of the molecular interactions between Z. tritici and its host remains limited, particularly the functions of many uncharacterized candidate effectors. Here, we assessed seven Z. tritici candidate effectors with elevated expression during the early biotrophic phase and transition to necrotrophy by transiently expressing them in the heterologous, non-host system Nicotiana benthamiana, with and without their predicted signal peptides. AlphaFold structural predictions revealed that two candidates share similarity with proteins of known function: a sterol-binding protein from Saccharomyces cerevisiae and a necrosis-inducing effector from the apple canker pathogen Valsa mali. Effector-associated phenotypes did not always correlate with expression timing, and signal peptide presence significantly influenced observed activities. Because N. benthamiana is not a natural host of Z. tritici, observed responses may reflect conserved, host-independent activities of effectors or immune recognition by N. benthamiana, rather than their native functions in wheat. Consistent with this distinction, two candidate effectors, Mycgr3107904 and Mycgr394290, induced cell death in N. benthamiana while also modulating ROS burst, likely indicative of immune recognition. Some candidates attenuate ROS production, suggesting potential targeting of conserved cellular processes, whereas none suppress PBR1-mediated cell death. Overall, our results indicate that candidate effector activities cannot be inferred from expression profiles alone, and that responses in the non-host N. benthamiana system reflect a combination of immune recognition and potential conserved activities, and primarily serve to prioritize candidates for validation in the native wheat-Z. tritici interaction.

molecular biology↗

bioRxiv2025

Pus7 mutation links tRNA dysregulation to aggressive behavior through activation of the integrated stress response and glycolytic reprogramming

Pseudouridine ({Psi}) is a prevalent RNA modification found in multiple RNA species. It is deposited by {Psi} synthases (Pus) and it can stabilize RNA structures. Patients carrying alterations in the Pus7 gene suffer from developmental delay, intellectual disability, microcephaly, hyperactivity and increased aggression levels. Here we show that the Pus7 mutation in human patient cells and in a Drosophila model is associated with a specific decrease of tRNA:Aspartate (tRNA-Asp) levels, which leads to slow decoding at Aspartate codons. This in turn activates the integrated stress response and induces a metabolic shift towards increased glycolysis and reduced mitochondrial respiration. Elevating tRNA-Asp expression, inhibiting the integrated stress response or dampening the glycolytic pathway is sufficient to rescue the aggressiveness phenotype, demonstrating the involvement of the tRNA-Asp-ISR-glycolysis axis in this behavior. Together our data provide new insights into the molecular defects associated with the loss of Pus7 and suggest potential new avenues for therapeutic treatment.

genetics↗

bioRxiv2025

The two-step purification method ViREn identifies a single NSUN6-mediated 5-methylcytosine modification promoting dengue virus RNA genome turnover

Chemical modifications on cellular and viral RNAs are new layers of post-transcriptional regulation of cellular processes including RNA stability and translation. Although advances in analytical methods have improved detection sensitivity, the precise mapping of RNA modifications at single-base resolution remains challenging. Especially for low abundant viral RNAs extracted from infected cells, requirements for sensitivity and purity limit accuracy and reproducibility. Here we report the two-step method ViREn for the enrichment of the genomic RNA (gRNA) of dengue virus (DENV), a positive-sense single-stranded RNA virus. This approach enabled the preparation of gRNA with significantly increased purity and led to the identification of a high-confidence 5-methylcytosine (m5C) site in DENV gRNA, orthogonally validated by Illumina-based bisulfite sequencing and direct RNA sequencing by Nanopore Oxford Technologies. Strikingly, this m5C modification was exclusively detected in gRNA extracted from infected cells but not in gRNA extracted from viral particles. We identified NSUN6 as the host methyltransferase catalyzing this modification and demonstrated a role for m5C in regulating DENV gRNA turnover. ViREn thus enables the mapping of m5C on low abundance viral gRNA with unprecedented precision and sensitivity and facilitates mechanistic studies into the role of RNA modification in virus replication.

biochemistry↗

bioRxiv2025

Redefining human pre-rRNA processing at single nucleotide resolution using long read Nanopore sequencing

Ribosome biogenesis requires the synthesis and processing of precursor rRNAs (pre-rRNAs) into mature rRNAs. Traditional methods like northern blotting and metabolic labeling offer limited resolution. We present NanoRibolyzer, a nanopore-based, long-read sequencing approach that enables ab initio identification and quantification of rRNA precursors. Using both supervised and unsupervised mapping, it detects known and novel pre-rRNA species and defines cleavage sites at single-nucleotide resolution. A simple cell fractionation protocol provides spatial separation of nuclear and cytoplasmic pre-rRNAs. Targeted knockdowns quantify intermediate accumulations, revealing condition-specific processing fingerprints with biomarker potential. Pseudouridine mapping shows that the primary 47S transcript is extensively modified, while aberrant products are not. With its high resolution and unique mapping strategy, NanoRibolyzer offers new insights into rRNA processing and modification, enhancing our understanding of ribosome biogenesis.

cell biology↗

bioRxiv2025

DNA-encoded Library Screening Uncovers Potent DNMT2 Inhibitors Targeting a Cryptic Allosteric Binding Site

The human RNA methyltransferase DNMT2 is thought to be involved in various pathophysiological processes, yet, a major challenge in drug targeting DNMT2 is given by the fact that current SAH-derived inhibitors have poor target selectivity and limited cellular permeability. In this study, we have performed a DNA-encoded library (DEL) screening on DNMT2 yielding five non-SAH-like hit structures, three of which feature a peptidomimetic scaffold. All DEL hits could be validated by orthogonal biophysical and biochemical assays for DNMT2 binding. At the same time, the lead structure did not interact with related methyltransferases from the DNMT and NSUN families highlighting an unmatched DNMT2-targeting selectivity profile. Subsequent crystallographic studies revealed the unique ligand binding mode including an active site loop rearrangement and the formation of a cryptic allosteric binding pocket able to modulate the enzymatic activity by non-covalent DNMT2 dimerization. Based on the crystallographic results, we performed a structure-activity relationship study around the inhibitor lead structure resulting in an optimized DNMT2 inhibitor (KD=3.04 {micro}M), which was able to reduce m5C levels in MOLM-13 tRNA. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=64 SRC="FIGDIR/small/632061v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@1bf92b3org.highwire.dtl.DTLVardef@38498borg.highwire.dtl.DTLVardef@1645740org.highwire.dtl.DTLVardef@9c61d5_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

ModiDeC: a multi-RNA modification classifier for direct nanopore sequencing

RNA modifications play a crucial role in various cellular functions. Here, we present ModiDeC, a deep-learning-based classifier able to identify and distinguish multiple RNA modifications (N6-methyladenosine, inosine, pseudouridine, 2'-O-methylguanosine, and N1-methyladenosine) using direct RNA sequencing. Alongside ModiDeC, we provide an extensive database of in vitro-transcribed and synthetic sequences generated with both the new RNA004 chemistry and the old RNA002 kit. We show that RNA modifications can be accurately recognized and distinguished across different sequence motifs using synthetic data as well as in HEK293T cells and human blood samples. ModiDeC comes with a graphical user interface that allows easy customization and adaptation to specific research questions, such as learning and classifying additional RNA modifications and further sequence motifs. The reproducibility across samples, together with the low rate of false positives, underscores the potential of ModiDeC as a powerful tool for advancing the analysis of epitranscriptomes and RNA modification.

bioinformatics↗

bioRxiv2024

The Fusarium graminearum effector protease FgTPP1 suppresses immune responses and facilitates Fusarium Head Blight Disease

Most plant pathogens secrete effector proteins to circumvent host immune responses, thereby promoting pathogen virulence. One such pathogen is the fungus Fusarium graminearum, which causes Fusarium Head Blight (FHB) disease on wheat and barley. Transcriptomic analyses revealed that F. graminearum expresses many candidate effector proteins during early phases of the infection process, some of which are annotated as proteases. However, the contributions of these proteases to virulence remains poorly defined. Here, we characterize a F. graminearum endopeptidase, FgTPP1 (FGSG_11164), that is highly upregulated during wheat spikelet infection and is secreted from fungal cells. To elucidate the potential role of FgTPP1 in F. graminearum virulence, we generated FgTPP1 deletion mutants ({Delta}Fgtpp1) and performed FHB infection assays. While the number of completely bleached spikes infected by F. graminearum wild-type reached 50% of total infected spikes, the number of fully bleached spikes infected by{Delta} Fgtpp1 mutants was 25%, suggesting FgTPP1 contributes to fungal virulence. Transient expression of green fluorescent protein (GFP)-tagged FgTPP1 revealed that FgTPP1 localizes, in part, to chloroplasts and attenuates chitin-mediated activation of mitogen-activated protein kinase (MAPK) signaling, reactive oxygen species production, and cell death induced by an autoactive disease resistance protein when expressed in planta. Notably, the FgTPP1 protein is conserved across the Ascomycota phylum, making it a core effector among ascomycete plant pathogens. These properties make FgTPP1 an ideal candidate for decoy substrate engineering, with the goal of engineering resistance to FHB, and likely other crop diseases caused by ascomycete fungi.

plant biology↗

bioRxiv2024

Direct RNA sequencing (RNA004) allows for improved transcriptome assessment and near real-time tracking of methylation for medical applications

Direct RNA sequencing (DRS) is a nanopore-based technique for analyzing RNA in its native form, promising breakthroughs in diagnostics and biomarker development. Coupled to RNA002 sequencing chemistry, its clinical implementation has been challenging due to low throughput, low accuracy, and lack of large-scale RNA-modification models. In this study, we evaluate the improvements achieved by pairing the latest RNA004 chemistry with novel modified-base-calling models for pseudouridine and N6-methyladenosine using diverse RNA samples from cell lines, synthetic oligos, and human blood. Finally, we present the first clinical application of DRS by confirming the loss of RNA methylation in a patient carrying truncating mutations in the methyltransferase METTL5. Conclusively, the combined use of RNA004 chemistry with the base-calling models significantly improved the throughput, accuracy, and site-specific detection of modifications. From this perspective, we offer an outlook on the potential suitability of DRS for use in routine diagnostics, the first comprehensive benchmark of human peripheral blood and quality assessments of RNA therapeutics.

bioinformatics↗

bioRxiv2024

Real-time transcriptomic profiling in distinct experimental conditions

Nanopore technology offers real-time sequencing opportunities, providing rapid access to sequenced data and allowing researchers to manage the sequencing process efficiently, resulting in cost-effective strategies. Here, we present focused case studies demonstrating the versatility of real-time transcriptomics analysis in rapid quality control for long-read RNA-seq. We illustrate its utility through four experimental setups: 1) transcriptome profiling of distinct human cellular populations, 2) identification of experimentally enriched transcripts, 3) transcriptional analysis of cells under heat shock conditions and 4) identification of experimentally manipulated genes (knockout and overexpression) in several yeast strains. We show how to perform multiple layers of quality control as soon as sequencing has started, addressing both the quality of the experimental and sequencing traits. Real-time quality control measures assess sample/condition variability and determine the number of identified genes per sample/condition. Furthermore, real-time differential gene/transcript expression analysis can be conducted at various time points post-sequencing initiation (PSI), revealing dynamic changes in gene/transcript expression between two conditions. Using real-time analysis, which occurs in parallel to the sequencing run, we identified differentially expressed genes/transcripts as early as 1hr PSI. These changes were consistently observed throughout the entire sequencing process. We discuss the new possibilities offered by real-time data analysis, which have the potential to serve as a valuable tool for rapid and cost-effective quality checks in specific experimental settings and can be potentially integrated into clinical applications in the future.

bioinformatics↗

bioRxiv2024

Genome-informed trophic classification and functional characterization of virulence proteins from the maize tar spot pathogen Phyllachora maydis

Phyllachora maydis is an ascomycete foliar fungal pathogen and the causal agent of tar spot in maize. Though P. maydis is considered one of the most economically important foliar pathogens of maize, our general knowledge of the trophic lifestyle and functional role of effector proteins from this fungal pathogen remains limited. Here, we utilized a genome-informed approach to predict the trophic lifestyle of P. maydis and functionally characterized a subset of candidate effectors from this fungal pathogen. Leveraging the most recent P. maydis genome annotation and the CATAStrophy pipeline, we show this fungal pathogen encodes a predicted Carbohydrate-active enzymes (CAZymes) repertoire consistent with that of biotrophs (monomertrophs). To investigate fungal pathogenicity, we selected eighteen candidate effector proteins that were previously shown to be expressed during primary disease development. We assessed whether these putative effectors share predicted structural similarity with other characterized fungal effectors and determined whether any suppress plant immune responses. Using AlphaFold2 and Foldseek, we showed one candidate effector, PM02_g1115, adopts a predicted protein structure similar to that of an effector from Verticillium dahlia. Furthermore, transient expression of candidate effector-fluorescent protein fusions in Nicotiana benthamiana revealed that most effector proteins localize to both the nucleus and the cytosol. Importantly, three candidate effectors consistently attenuated chitin-mediated reactive oxygen species production in N. benthamiana. Collectively, these results presented herein provide valuable insights into the predicted trophic lifestyle and putative functions of effectors from P. maydis and will likely stimulate continued research to elucidate the molecular mechanisms used by P. maydis to induce tar spot.

plant biology↗

bioRxiv2023

Mechanisms of infection and response of the fungal wheat pathogen Zymoseptoria tritici during compatible, incompatible and non-host interactions

Zymoseptoria tritici is responsible for Septoria tritici blotch, a disease causing significant annual yield losses in wheat. To investigate infection phase-specific gene expression in the pathogen, we analyzed gene expression during infection of susceptible (Taichung 29) and resistant (Veranopolis and Israel 493) wheat cultivars, plus the non-host species barley at 1, 3, 6, 10, 17 and 23 days post inoculation (DPI). There were dramatic differences in pathogen gene expression at 10 DPI in the susceptible compared to both resistant interactions. The most pronounced differences in pathogen gene expression were observed at 3 DPI in both the susceptible and resistant host interactions compared to the non-host interaction. Thirty-one putative effectors showed early expression during the susceptible interaction compared to the non-host interaction, and six effectors were selected for subcellular localization studies. Using Agrobacterium-mediated transient expression in Nicotiana benthamiana, subcellular localization assays revealed that two candidate effectors, Mycgr3109710 and Mycgr394290, localized to putative mobile cytosolic bodies when expressed without their signal peptides, suggesting potential roles in intracellular signaling or host gene regulation. When expressed with their native signal peptides, four candidate effectors localized to the cytosol, while one effector, Mycgr3107904, did not accumulate to detectable levels, as shown by immunoblot analysis, indicating degradation. Comparison of pathogen gene expression in the susceptible host to expression in the resistant hosts, allowed us to identify genes that are expressed during the transition from biotrophic to necrotrophic growth at 10 DPI. Comparison of pathogen gene expression in resistant and susceptible hosts, versus in the non-host barley, allowed us to identify genes involved in initial colonization and host recognition. In addition, our study contributes to understanding candidate effectors that are activated early during infection and may play a role in the initial suppression of plant immunity, making them strong candidates for functional characterization.

plant biology↗

bioRxiv2023

Pseudomonas syringae pv. tomato DC3000 induces defense responses in diverse maize inbred lines

Many phytopathogens translocate virulence (effector) proteins into plant cells to circumvent host immune responses during infection. One such pathogen is Pseudomonas syringae pv. tomato DC3000, which secretes at least twenty-nine effectors into host cells, of which a subset elicits host defense responses in crop plant species. However, it is unknown whether P. syringae pv. tomato DC3000 activates immune responses in diverse maize inbreds. Here, we screened a diverse maize germplasm collection for effector-dependent recognition of this bacterial pathogen. As a control, we infiltrated Pseudomonas syringae DC3000(D36E), a derivative of P. syringae pv. tomato DC3000 that lacks all endogenous effectors. In our evaluations, we observed a variety of responses to P. syringae pv. tomato DC3000 in maize and scored the phenotypes as either no observable response (N) or as one of three responses: weak chlorosis (WC), chlorosis (C) with minimal cell death, and hypersensitive reaction (HR)-like cell death. Of the twenty-six maize inbreds screened, 13 were scored as N, 2 as WC, 2 as C, and 9 as HR-like cell death. Importantly, no maize line responded to P. syringae DC3000(D36E), demonstrating the responses observed are likely dependent upon recognition of one or more Pseudomonas effectors. Importantly, maize inbreds that recognize P. syringae pv. tomato DC3000 accumulated detectable hydrogen peroxide as well as an increase in transcript expression of a subset of maize defense genes. Collectively, our results will likely stimulate new research aimed at identifying the cognate maize disease resistance proteins that recognize the activities of one or more bacterial effectors.

plant biology↗

bioRxiv2023

N1-methylation of adenosine (m1A) in ND5 mRNA leads to complex I dysfunction in Alzheimer's disease

One mechanism of particular interest to regulate mRNA fate post-transcriptionally is mRNA modification. Especially the extent of m1A mRNA methylation is highly discussed due to methodological differences. However, one single m1A site in mitochondrial ND5 mRNA was unanimously reported by different groups. ND5 is a subunit of complex I of the respiratory chain. It is considered essential for the coupling of oxidation and proton transport. Here we demonstrate that this m1A site might be involved in the pathophysiology of Alzheimers disease (AD). One of the pathological hallmarks of this neurodegenerative disease is mitochondrial dysfunction, mainly induced by Amyloid {beta} (A{beta}). A{beta} mainly disturbs functions of complex I and IV of the respiratory chain. However, the molecular mechanism of complex I dysfunction is still not fully understood. We found enhanced m1A methylation of ND5 mRNA in an AD cell model as well as in AD patients. Formation of this m1A methylation is catalyzed by increased TRMT10C protein levels, leading to translation repression of ND5. As a consequence, here demonstrated for the first time, TRMT10C induced m1A methylation of ND5 mRNA leads to mitochondrial dysfunction. Our findings suggest that this newly identified mechanism might be involved in A{beta}-induced mitochondrial dysfunction.

neuroscience↗

bioRxiv2023

Adenine methylation is very scarce in the drosophila genome and not erased by the Ten Eleven Translocation dioxygenase.

N6-methyladenine (6mA) DNA modification has recently been described in metazoans, including in drosophila, for which the erasure of this epigenetic mark has been ascribed to the Ten Eleven Translocation (TET) enzyme. Here, we re-evaluated 6mA presence and TET impact on drosophila genome. Using axenic or conventional breeding conditions, we found traces of 6mA by LC-MS/MS and no significant increase in 6mA levels in the absence of TET, suggesting that this modification is present at very low levels in the drosophila genome but not regulated by TET. Consistent with this latter hypothesis, further molecular and genetic analyses showed that TET does not demethylate 6mA but acts essentially in an enzymatic-independent manner. Our results call for further caution concerning the role and regulation of 6mA DNA modification in metazoans and underline the importance of TET non-enzymatic activity for fly development.

molecular biology↗