Biology subjects

Bizet, M.

Publications and source records attributed to Bizet, M..

6 recordsLinked to original sources

bioRxiv2025

m3C is a mitochondrial mRNA modification which promotes tumor progression

Mitochondrial mRNA modifications are suggested to play a role in fine-tuning mitochondrial gene expression and function. However, the epitranscriptomic landscape of mitochondrial mRNA (mt-mRNA) remains poorly explored. Here, we uncover N3-methylcytosine (m3C) as a novel mt-mRNA that is catalyzed by METTL8, an enzyme previously known to modify mt-tRNA. Transcriptome-wide mapping reveals that METTL8-dependent m3C is enriched in mt-mRNAs encoding complex I subunits of the respiratory chain. Additionally, METTL8 is highly expressed in various cancers, notably in cervical cancer. METTL8 depletion impairs cell migration in vitro and reduces tumor growth in mouse xenografts. Finally, transcriptomic analyses further link METTL8 expression to oncogenic pathways, mitochondrial functions and complex I activity. Together, our results reveal a novel mitochondrial mRNA modification that promotes cancer progression.

cell biology↗

bioRxiv2024

Expanding the diversity of nitroxide-based paramagnetic probes conjugated to non-canonical amino acids for SDSL-EPR applications

Understanding protein structure requires studying its dynamics, which is critical to elucidating its functional role. Over time, biophysical techniques have revolutionized this field, offering remarkable insights into the structure-function relationship. Among these, Site-Directed Spin Labelling (SDSL) combined with Electron Paramagnetic Resonance (EPR) is a powerful method delivering structural data at the residue level, irrespective of protein size or environment. Traditional nitroxide labels, which target cysteine residues, often face limitations when these residues are essential for protein structure or function. To address this, alternatives have been proposed as the use of non-canonical amino acids (ncaa) coupled with specific nitroxide labels. This study introduces 14N-HO-5223, a novel nitroxide label specific to the pAzPhe ncaa, alongside its 15N-derivative. These labels were grafted at two sites of the model protein, the diflavin Cytochrome P450 reductase. For comparative purpose, two already reported labels were also used. Continuous wave (cw) EPR spectroscopy validated the HO-5223 label as an effective reporter of protein dynamics. Additionally, Double Electron-Electron Resonance (DEER) measurements provided distance distributions between the semi-quinone FMNH* state of the CPR and all nitroxide labels. These results expand the toolkit of the ncaa-nitroxide pairs, enabling EPR-based structural studies of proteins where cysteine modification is impractical, further advancing our ability to decode protein dynamics and function.

biophysics↗

bioRxiv2023

Tet-dependent 5-hydroxymethyl-Cytosine modification of mRNA regulates the axon guidance genes robo2 and slit in Drosophila

Modifications of mRNA, especially methylation of adenosine, have recently drawn much attention. The much rarer modification, 5-hydroxymethylation of cytosine (5hmC), is not well understood and is the subject of this study. Vertebrate Tet proteins are 5-methylcytosine (5mC) hydroxylases and catalyze the transition of 5mC to 5hmC in DNA. These enzymes have recently been shown to have the same function in messenger RNAs in both vertebrates and in Drosophila. The Tet gene is essential in Drosophila as Tet knock-out animals do not reach adulthood. We describe the identification of Tet-target genes in the embryo and larval brain by mapping one, Tet DNA-binding sites throughout the genome and two, the Tet-dependent 5hmrC modifications transcriptome-wide. 5hmrC modifications are distributed along the entire transcript, while Tet DNA-binding sites are preferentially located at the promoter where they overlap with histone H3K4me3 peaks. The identified mRNAs are preferentially involved in neuron and axon development and Tet knock-out led to a reduction of 5hmrC marks on specific mRNAs. Among the Tet-target genes were the robo2 receptor and its slit ligand that function in axon guidance in Drosophila and in vertebrates. Tet knock-out embryos show overlapping phenotypes with robo2 and both Robo2 and Slit protein levels were markedly reduced in Tet KO larval brains. Our results establish a role for Tet-dependent 5hmrC in facilitating the translation of modified mRNAs primarily in cells of the nervous system.

neuroscience↗

bioRxiv2022

The RNA demethylase FTO controls m6A marking on SARS-CoV-2 and classifies COVID-19 severity in patients

The RNA modification N6-methyladenosine (m6A) plays a key role in the life cycles of several RNA viruses. Whether this applies to SARS-CoV-2 and whether m6A affects the outcome of COVID-19 disease is still poorly explored. Here we report that the RNA demethylase FTO strongly affects both m6A marking of SARS-CoV-2 and COVID-19 severity. By m6A profiling of SARS-CoV-2, we confirmed in infected cultured cells and showed for the first time in vivo in hamsters that the regions encoding TRS_L and the nucleocapsid protein are multiply marked by m6A, preferentially within RRACH motifs that are specific to {beta}-coronaviruses and well conserved across SARS-CoV-2 variants. In cells, downregulation of the m6A demethylase FTO, occurring upon SARS-CoV-2 infection, increased m6A marking of SARS-CoV-2 RNA and slightly promoted viral replication. In COVID-19 patients, a negative correlation was found between FTO expression and both SARS-CoV-2 expression and disease severity. FTO emerged as a classifier of disease severity and hence a potential stratifier of COVID-19 patients.

molecular biology↗

bioRxiv2021

Novel Role of UHRF1 in DNA methylation-mediated repression of latent HIV-1

The multiplicity, heterogeneity and dynamic nature of HIV-1 latency mechanisms are reflected in the current lack of functional cure for HIV-1 and in the various reported ex vivo potencies of latency- reversing agents. Here, we investigated the molecular mechanisms underlying the potency of the DNA methylation inhibitor 5-aza-2-deoxycytidine (5-AzadC) in HIV-1 latency reversal. Doing so, we uncovered specific demethylation CpG signatures induced by 5-AzadC in the HIV-1 promoter. By analyzing the binding modalities to these CpG, we revealed the recruitment of the epigenetic integrator UHRF1 to the HIV-1 promoter. We further demonstrated the role of UHRF1 in DNA methylation- mediated silencing of the latent HIV-1 promoter. As a proof-of-concept to this molecular characterization, we showed that pharmacological downregulation of UHRF1 in ex vivo HIV+ patient cell cultures resulted in potent reactivation of latent HIV-1. Together, we identify UHRF1 as a novel actor in HIV-1 gene silencing and highlight that it constitutes a new molecular target for HIV-1 curative strategies.

microbiology↗

bioRxiv2020

The chromatin remodeler LSH controls genome-wide cytosine hydroxymethylation

TET proteins convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), leading to a dynamic epigenetic state of DNA that can influence transcription. While TET proteins have been associated with either epigenetic repression or activation complexes, the overall understanding of the molecular mechanisms involved in TET-mediated regulation of gene transcription still remains limited. Here, we show that TET proteins interact with lymphoid-specific helicase (LSH), a chromatin remodeling factor belonging to the SNF2 super family. Lsh knock-out leads to a significant reduction of 5-hydroxymethylation global level in mouse embryonic fibroblasts (MEFs) and in embryonic stem cells (ESC). Whole genome sequencing of 5hmC in wild-type versus Lsh knock-out MEFs and ESCs showed that in absence of Lsh, some regions of the genome gain 5hmC while others lose it, with not much effect on gene expression. We further show that 5hmC modifications upon Lsh loss is not a direct consequence of 5mC decrease, as differentially hydroxymethylated regions (DhMR) did not overlap with DMR (differentially methylated regions), underlying that these modifications occurred at different genomic loci. Altogether, our results suggest that LSH is a key regulator of 5hmC in both MEFs and ESC and that TET proteins rely on specific factors to establish genome-wide 5hmC patterns.

molecular biology↗