Biology subjects

Decourcelle, M.

Publications and source records attributed to Decourcelle, M..

6 recordsLinked to original sources

bioRxiv2026

A generalizable normalization framework to decouple protocol and instrument effects: Application to high-sensitivity proteomics multicentric study (PME13)

Multicenter studies are essential for benchmarking analytical workflows, yet their interpretation is often confounded by the combined effects of experimental protocols and instrumentation. To address this challenge, we introduce a simple normalization-based analytical framework, the recovery metric ({rho}), designed to decouple protocol driven effects from instrument dependent variability. We applied this framework to the 13th Proteomics Multicentric Experiment (PME13), a large multicentric proteomics dataset generated across 27 laboratories using high sensitivity workflows and varying sample preparation protocols. By leveraging a common digested reference sample, {rho} enables direct cross-comparison of all datasets on a unified scale, effectively minimizing instrument-related biases. Using this approach, we demonstrate that apparent instrument dependent trends are largely removed when evaluated through {rho}, revealing consistent protocol driven effects across laboratories. Statistical modeling identified key variables influencing {rho}, including sample input amount, reduction and alkylation, and the use of n-dodecyl-{beta}-D-maltoside (DDM). While DDM was associated with improved {rho}, reduction and alkylation and additional handling steps led to reduced performance, particularly at low input levels. We further highlight practical considerations for the application of ratio based normalization, including the occurrence of values exceeding theoretical bounds, which reflect deviations from underlying assumptions and require appropriate filtering. Overall, this work establishes a generalizable analytical strategy for disentangling confounding factors in multicentric datasets and provides practical guidelines for optimizing high sensitivity proteomics (HSP) workflows. The proposed framework is broadly applicable to other analytical fields where cross laboratory comparability is required.

bioinformatics↗

bioRxiv2025

A reciprocal interplay between 5-HT2A and mGlu5 receptors underlies neuroplasticity

The serotonin (5-HT)2A receptor is the primary target of numerous psychoactive drugs including serotonergic psychedelics, and mediates psychedelics-induced neuroplasticity, but the signaling mechanisms involved remain poorly characterized. Using quantitative phosphoproteomics, we show that the administration of the hallucinogenic 5-HT2A receptor agonist 2,5-dimethoxy-4-iodoamphetamine (DOI) to mice promotes the phosphorylation of synaptic proteins belonging to a strongly interconnected protein network and comprising the metabotropic glutamate (mGlu)5 receptor and the scaffolding protein Shank3. Functional studies revealed that hallucinogenic and non-hallucinogenic 5-HT2A receptor agonists promote synaptic targeting of mGlu5 receptor and its association with Shank3. Furthermore, they gate neuroplasticity in cortical neurons through a mechanism requiring mGlu5 receptor, protein kinase C and Shank3. Conversely, neuroplasticity elicited by mGlu5 receptor activation depends on 5-HT2A receptor. Collectively, these findings demonstrate that neuroplasticity-promoting properties of psychedelics depend on a functional, reciprocal interplay between 5-HT2A and mGlu5 receptors involving the synaptic scaffolding protein Shank3.

neuroscience↗

bioRxiv2024

Pervasive translation of the downstream ORF from bicistronic mRNAs by human cells: impact of the upstream ORF codon usage and splicing

In this work, we present a straightforward model to study gene expression regulation from virus-like bicistronic mRNAs in human cells, composed by a short shble upstream ORF and a reporter egfp downstream ORF. We have engineered thirteen synonymous versions of the shble ORF to explore a large parameter space in compositional and folding features, as well as differences in propensity to undergo splicing. Our experimental model focuses on the control and modulation of shble translation elongation and their effects on downstream translation: i) regarding translation initiation, all constructs share identical 5UTR, ribosomal binding site and shble start codon context; and ii) regarding egfp translation, all constructs share identical shble stop codon context, spacer between shble and egfp and the full egfp sequence. Our results show first that the human translation machinery can translate the downstream ORF in bicistronic mRNAs, independently of the characteristics of the upstream ORF. Even if GFP protein levels are hundreds of times lower than SHBLE protein levels, our results challenge the textbook interpretation positing that the eukaryotic translation machinery does not handle the downstream ORF in bicistronic transcripts. Second, we show that synonymous recoding of the upstream ORF determines its own translation efficiency, can uncover cryptic splice signals, and largely conditions the probability of translation from the downstream ORF. Our results are consistent with a leaky scanning mechanism at play for downstream ORF translation from bicistronic mRNAs in human cells.

molecular biology↗

bioRxiv2022

The non-neutral, multi-level, phenotypic impact of synonymous mutations revealed through heterologous gene expression in human cells.

Differences in codon frequency between genomes, genes, or positions along a gene, modulate transcription and translation efficiency, leading to phenotypic and functional differences. Here, we present a multiscale analysis of the effects of synonymous codon recoding during heterologous gene expression in human cells, quantifying the phenotypic consequences of codon usage bias at different molecular and cellular levels, with an emphasis on translation elongation. Six synonymous versions of an antibiotic resistance gene were generated, fused to a fluorescent reporter, and independently expressed in HEK293 cells. Multiscale phenotype was analysed by means of: quantitative transcriptome and proteome assessment, as proxies for gene expression; cellular fluorescence, as a proxy for single-cell level expression; and real-time cell proliferation in absence or presence of antibiotic, as a proxy for the cell fitness. We show that differences in codon usage bias strongly impact the molecular and cellular phenotype: (i) they result in large differences in mRNA levels and in protein levels, leading to differences of over fifteen times in translation efficiency; (ii) they introduce unpredicted splicing events; (iii) they lead to reproducible phenotypic heterogeneity; and (iv) they lead to a trade-off between the benefit of antibiotic resistance and the burden of heterologous expression. In human cells in culture, codon usage bias modulates gene expression by modifying mRNA availability and suitability for translation, leading to differences in protein levels and eventually eliciting functional phenotypic changes. IMPORTANCESynonymous codons encode for the same amino acid, but they are not neutral regarding gene expression or protein synthesis. Bias between synonymous codons have evolved naturally and are also applied in biotechnology protein production. We have studied the multilevel impact of codon usage on a human cell system. We show that differences in codon usage lead to transcriptomic, proteomic and functional changes, modulating gene expression and cellular phenotype.

systems biology↗

bioRxiv2021

Human cellular homeostasis buffers trans-acting translational effects of heterologous gene expression with very different codon usage bias

The frequency of synonymous codons in protein coding genes is non-random and varies both between species and between genes within species. Whether this codon usage bias (CUBias) reflects underlying neutral mutational processes or is instead shaped by selection remains an open debate, especially regarding the role of selection for enhanced protein production. Variation in CUBias of a gene (be it natural synonymous mutations or biotechnological synonymous recoding) can have an enormous impact on its expression by diverse cis-acting mechanisms. But expression of genes with extreme CUBias can also lead to strong phenotypic effects by altering the overall intracellular translation homeostasis via competition for ribosomal machinery or tRNA depletion. In this study, we expressed at high levels in human cells six different synonymous versions of a gene and used matched transcriptomic and proteomic data to evaluate the impact of CUBias of the heterologous gene on the translation of cellular transcripts. Our experimental design focused specifically on differences during translation elongation. Response to expression of the different synonymous sequences was assessed by various approaches, ranging from analyses performed on a per-gene basis to more integrated approaches of the cell as a whole. We observe that the transcriptome displayed substantial changes as a result of heterologous gene expression by triggering an intense antiviral and inflammatory response, but that changes in the proteomes were very modest. Most importantly we notice that changes in translation efficiency of cellular transcripts were not associated with the direction of the CUBias of the heterologous sequences, thereby providing only limited support for trans-acting effects of synonymous changes. We interpret that, in human cells in culture, changes in CUBias can lead to important cis-acting effects in gene expression, but that cellular homeostasis can buffer the phenotypic impact of overexpression of heterologous genes with extreme CUBias.

cell biology↗

bioRxiv2019

The PIWI protein Aubergine recruits eIF3 to activate translation in the germ plasm

Piwi-interacting RNAs (piRNAs) and PIWI proteins are essential in germ cells to repress transposons and regulate mRNAs. In Drosophila, piRNAs bound to the PIWI protein Aubergine (Aub) are transferred maternally to the embryo and regulate maternal mRNA stability through two opposite roles. They target mRNAs by incomplete base-pairing, leading to both their destabilization in the soma, and stabilization in the germ plasm. Here, we report a function of Aub in translation. Aub is required for translational activation of nanos mRNA, a key determinant of the germ plasm. Aub physically interacts with the poly(A) binding protein PABP and the translation initiation factor eIF3. Polysome gradient profiling identifies Aub role at the initiation step of translation. In the germ plasm, PABP and eIF3d assemble in foci that surround Aub-containing germ granules, and Aub acts with eIF3d to promote nanos translation. These results reveal a new mode of mRNA regulation by Aub, highlighting PIWI protein versatility in mRNA regulation.

developmental biology↗