Biology subjects

Potel, C. M.

Publications and source records attributed to Potel, C. M..

7 recordsLinked to original sources

bioRxiv2026

Multi-omics and biophysical phosphoproteomics upon BRAF inhibition uncover functional networks of BRAFV600E-driven signaling

Dysregulated kinase activity drives oncogenic signaling, disrupts cellular homeostasis, and promotes tumour progression. The BRAFV600E mutation constitutively activates the MAPK pathway and is a key therapeutic target in melanoma and other cancers, but the functional relevance of most downstream phosphorylation events and mechanisms of drug resistance remain unclear. To address this, a global multi-omic model of BRAF inhibition response was established in BRAFV600E-mutant cells by integrating time-resolved and biophysical phosphoproteomics, transcriptomics, and thermal proteome profiling. Ultradeep phosphoproteomics revealed extensive phosphorylation changes upon BRAF inhibitor treatment, while biophysical phosphoproteomics identified phosphorylation events linked to altered protein solubility and subcellular localization, suggesting changes in nucleic acid interactions and nuclear reorganisation. Network-based integration of these datasets prioritized functionally relevant phosphorylation sites and kinases. Experimental validation identified CDK9, CLK3, and TNIK as critical regulators of BRAFV600E signaling and candidate targets for combinatorial inhibition capable of re-sensitising resistant cells. The transcription factor ETV3 emerged as a previously unrecognised effector of BRAF signaling. Biophysical proteomics data confirmed that ETV3 phosphorylation modulates DNA-binding, while functional assays combining knockdown, metabolomics, and drug screening demonstrated its role in coordinating transcriptional and metabolic adaptations to BRAF inhibition. This study provides a systems-level framework linking phosphorylation dynamics to protein function and phenotype, identifies ETV3 as a new node in oncogenic BRAF signaling, and illustrates how integrated, site-resolved models can reveal mechanisms of kinase-driven oncogenesis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=43 SRC="FIGDIR/small/704793v1_ufig1.gif" ALT="Figure 1"> View larger version (13K): org.highwire.dtl.DTLVardef@1df1401org.highwire.dtl.DTLVardef@9a77a5org.highwire.dtl.DTLVardef@124f819org.highwire.dtl.DTLVardef@1c6b57_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LITime- and cell type-resolved phosphoproteomics maps BRAF inhibition dynamics C_LIO_LIBiophysical phosphoproteomics, combining quantitative phosphoproteomics with solubility profiling or nuclear fractionation, reveals phosphorylation-driven changes of protein solubility and localization C_LIO_LIIntegration of abundance and biophysical phosphoproteomics data identifies functionally relevant phosphorylation events of BRAFV600E signaling C_LIO_LINetwork integration of multimodal phosphoproteomic, transcriptomic and thermal proteome profiling data links signaling to protein function and cellular phenotypes C_LIO_LIBiophysical evidence improves models and identifies non-canonical kinases driving BRAF signaling as well as novel downstream regulators such as ETV3 C_LIO_LIFollow-up experiments reveal a ETV3-GLUT3-mediated metabolic adaptation in BRAFV600E cells C_LI

systems biology↗

bioRxiv2025

High-throughput peptide-centric local stability assay extends protein-ligand identification to membrane proteins, tissues, and bacteria

Systematic mapping of protein-ligand interactions is essential for understanding biological processes and drug mechanisms. Peptide-centric local stability assay (PELSA) is a powerful tool for detecting these interactions and localizing potential binding sites. However, its original workflow is limited in throughput, sample compatibility and accessible protein targets. Here, we introduce a high-throughput adaptation - HT-PELSA - that increases sample processing efficiency 100-fold while maintaining high sensitivity and reproducibility. HT-PELSA substantially extends the capabilities of the original method by enabling sensitive protein-ligand profiling in crude cell, tissue and bacterial lysates, making it possible to identify membrane protein targets in diverse biological systems. We demonstrate that HT-PELSA can precisely and accurately determine binding affinities of small molecule inhibitors, sensitively detect direct and allosteric ATP binding, and reveal off-target interactions of a marketed kinase inhibitor in heart tissue. By enhancing scalability, reducing costs, and enabling system-wide drug screening across a wide range of sample types, HT-PELSA - when combined with next-generation mass spectrometry - offers a powerful platform poised to accelerate both drug discovery and basic biological research.

cell biology↗

bioRxiv2024

Pervasive sublethal effects of agrochemicals as contributing factors to insect decline

Insect biomass is declining across the globe at an alarming rate. Climate change and the widespread use of pesticides have been hypothesized as two underlying drivers. However, the lack of systematic experimental studies across chemicals and species limits our causal understanding of this problem. Here, we employed a chemical library encompassing 1024 different molecules--including insecticides, herbicides, fungicides, and plant growth inhibitors --to investigate how insect populations are affected by varying concentrations of pesticides, focusing on sublethal doses. Using a controlled laboratory pipeline for Drosophila melanogaster, we found that 57% of these chemicals affect the behavior of larvae at sublethal concentrations, and an even higher proportion compromises long-term survivability after acute exposure. Consistent with these results, we observed that exposure to chemicals at doses orders of magnitude below lethality induced widespread phosphorylation changes across the larval proteome. The effects of agrochemicals were amplified when the ambient temperature was increased by four degrees. We also tested the synergistic effects of multiple chemicals at doses found widely in nature and observed fitness-reducing changes in larval developmental time, behavior, and reproduction. Finally, we expanded our investigation to additional fly species, mosquitos, and butterflies and detected similar behavioral alterations triggered by pesticides at sublethal concentrations. Our results provide experimental evidence that strongly suggests sublethal doses of agrochemicals coupled with changes in environmental temperatures are contributing to the global decline in insect populations. We anticipate that our assays can contribute to improving chemical safety assessment, better protect the environment, secure food supplies, and safeguard animal and human health, as well as understand our rapidly changing world.

ecology↗

bioRxiv2023

Deep quantitative glycoproteomics reveals gut microbiome induced remodeling of the brain glycoproteome

HighlightsO_LIHigh throughput glycoproteomics method with multiplexed quantification C_LIO_LI25-fold improvement of the mouse brain glycoproteome coverage C_LIO_LIStructural features dictate level of glycosite micro-heterogeneity C_LIO_LIGut microbiome composition extensively impacts the brain glycoproteome C_LIO_LIModulation of glycosylation is site-specific C_LI Protein glycosylation is a highly diverse post-translational modification, modulating key cellular processes such as cell signaling, adhesion and cell-cell interactions. Its deregulation has been associated with various pathologies, including cancer and neurological diseases. Methods capable of quantifying glycosylation dynamics are essential to start unraveling the biological functions of protein glycosylation. Here we present Deep Quantitative Glycoprofiling (DQGlyco), a method that combines high-throughput sample preparation, high-sensitivity detection, and precise multiplexed quantification of protein glycosylation. We used DQGlyco to profile the mouse brain glycoproteome, in which we identify 158,972 and 15,056 unique N- and O-glycopeptides localized on 3,199 and 2,365 glycoproteins, respectively - this amounts to 25-fold more glycopeptides identified compared to previous studies. We observed extensive heterogeneity of glycoforms and determined their functional and structural preferences. The presence of a defined gut microbiota resulted in extensive remodeling of the brain glycoproteome when compared to that of germ-free animals, exemplifying how the gut microbiome may affect brain protein functions.

molecular biology↗

bioRxiv2020

Global mapping of Salmonella enterica-host protein-protein interactions during infection

Intracellular bacterial pathogens inject effector proteins into host cells to hijack diverse cellular processes and promote their survival and proliferation. To systematically map effector-host protein-protein interactions (PPIs) during infection, we generated a library of 32 Salmonella enterica serovar Typhimurium (STm) strains expressing chromosomally encoded affinity-tagged effector proteins, and quantified PPIs in macrophages and epithelial cells by Affinity-Purification Quantitative Mass-Spectrometry. Thereby, we identified 25 previously described and 421 novel effector-host PPIs. While effectors converged on the same host cellular processes, most had multiple targets, which often differed between cell types. Using reciprocal co-immunoprecipitations, we validated 13 out of 22 new PPIs. We then used this host-pathogen physical interactome resource to demonstrate that SseJ and SseL collaborate in redirecting cholesterol to the Salmonella Containing Vacuole (SCV) via NPC1, PipB directly recruits the organelle contact site protein PDZD8 to the SCV, and SteC promotes actin bundling by directly phosphorylating formin-like proteins.

microbiology↗

bioRxiv2020

Impact of phosphorylation on thermal stability of proteins

Reversible protein phosphorylation regulates virtually every cellular process and is arguably the most well-studied post-translational modification. Still, less than 3% of the phosphorylation sites identified in humans have annotated functions. Functionally-relevant phosphorylation sites are known to trigger conformational changes to proteins and/or to regulate their interactions with other proteins, nucleic acids and small molecules - all of which can be reflected in the thermal stability of a protein. Thus, combining thermal proteome profiling (TPP) with phosphoproteomics (phospho-TPP) provides a way to assess the functional relevance of identified phosphorylation sites on a proteome-wide scale by comparing the melting behavior of a protein and its phosphorylated form(s). We performed phospho-TPP experiments in HeLa cells with an optimized protocol, and conclude that phosphorylation does affect protein thermal stability, but to a much lesser extent than previously reported.

systems biology↗

bioRxiv2019

Towards a systematic map of the functional role of protein phosphorylation

Phosphorylation is a critical post-translational modification involved in the regulation of almost all cellular processes. However, less than 5% of thousands of recently discovered phosphorylation sites have a known function. Here, we devised a chemical genetic approach to study the functional relevance of phosphorylation in S. cerevisiae. We generated 474 phospho-deficient mutants that, along with the gene deletion library, were screened for fitness in 102 conditions. Of these, 42% exhibited growth phenotypes, suggesting these phosphosites are likely functional. We inferred their function based on the similarity of their growth profiles with that of gene deletions, and validated a subset by thermal proteome profiling and lipidomics. While some phosphomutants showed loss-of-function phenotypes, a higher fraction exhibited phenotypes not seen in the corresponding gene deletion suggestive of a gain-of-function effect. For phosphosites conserved in humans, the severity of the yeast phenotypes is indicative of their human functional relevance. This study provides a roadmap for functionally characterizing phosphorylation in a systematic manner.

systems biology↗