Biology subjects

Boulais, J.

Publications and source records attributed to Boulais, J..

14 recordsLinked to original sources

bioRxiv2026

Comparative profiling of SARS-CoV-2 variant infections reveals diverse impacts on host cell RNA and RNA binding protein distribution and regulation

RNA viruses perturb cellular RNA metabolism to promote viral transcript expression and genome replication. However, the extent of host cell RNA reprogramming and the consequences on RNA binding protein (RBP) expression and localization remain poorly understood. Here, we utilized an integrated set of transcriptomics, proteomics, and high-content imaging approaches to compare the impact of different SARS-CoV-2 variants, including an early viral isolate and later emerging variants of concern (Alpha, Delta and Omicron) on the host cellular RNA and RBP landscapes. While all variants deeply remodelled the host transcriptome in a subcellular compartment-specific manner, we found that Omicron variants express lower levels of viral RNA products compared to earlier variants, while still driving efficient production of viral proteins. The surge in viral RNA expression correlated with altered subcellular RNA biotype distribution, cytoplasmic mRNA exclusion, the selective disruption of the localization of polyA-tail interacting proteins, and the nuclear accumulation of non-coding transcripts such as RN7SL family members and MALAT-1. Unexpectedly, we found many RBPs that exhibiting altered subcellular localization following abiotic innate immune stimulation with poly(I:C) but remained unaltered in cells infected with SARS-CoV-2 variants, revealing a capacity for these viruses to limit the overall impact on the RBP subcellular distribution. Our results further indicate that Omicron infection sustains higher expression levels of proteins components of the signal recognition particle and translation factors, which may enable a more effective production of immune response and viral components. Altogether, this work highlights the deep impact of SARS-CoV-2 infection on host cell RNA regulatory landscape and provides new insights to explain the differential impact of Omicron compared to early SARS-CoV-2 variants. HighlightsO_LISARS-CoV-2 variants drive varying degrees of viral RNA expression in infected Calu-3 cells C_LIO_LIInfection reshapes the host cell RNA landscape in a cell-compartment specific manner C_LIO_LIRN7SL RNA exhibits a strong increase in relative abundance and accumulates in the nucleus of infected cells C_LIO_LISARS-CoV-2 infections impact the localization of polyA-tail binding proteins while blocking the nuclear accumulation of immune response-sensitive RBPs C_LIO_LIOmicron infections are less disruptive to cellular function and sustain the production of translation factors and components of the SRP complex C_LI

molecular biology↗

bioRxiv2025

Multiple FGFR1 mutations modulate tumorigenic mechanisms in glioneuronal tumors.

FGFR1 genetic alterations are associated with human diseases, including brain tumors. We reported multiple FGFR1 mutations in familial and sporadic cases of low-grade glioneuronal tumors, suggesting intrinsic mechanisms of selective pressure toward FGFR1 multiple events arising in the context of a quiet genome. To decipher the molecular mechanisms triggered by multiple FGFR1 mutations, we have mapped the proximal interactome of wild-type, single-and double-mutant FGFR1 proteins through a BioID-MS approach. Our data reveals novel oncogenic functionality for the two hotspots N546K and K656E, linked to evasion of lysosomal degradation. We identified a modulatory role played by the susceptibility variant R661P, which hampers the oncogenic potential of both hotspot mutations by rescuing receptor degradation and reducing N546K affinity for the downstream effector PLC{gamma}. The R661P variant alone abolished the self-renewal capacity of oligodendroglioma cells and showed downregulation of genes involved in neurodevelopment and neuro-glial cell fate decisions, both aspects overcome in the double mutants. This study sheds light on the oncogenic effects associated with FGFR1 alterations and their recurrence in low-mutation burden and therapy naive tumors.

cancer biology↗

bioRxiv2024

Investigating the Impacts of Sphingomyelinases on Extracellular Vesicle Cargo Sorting

Extracellular vesicles (EVs) form through regulated biogenesis processes involving sphingomyelinases (SMases), enzymes that metabolize sphingomyelin to produce ceramide--a lipid influencing membrane rigidity and essential for EV generation. This study explores the impact on EV protein and RNA cargoes resulting from inhibiting neutral SMase (NSM) and acid SMase (ASM) in human MCF7 cells. Our results revealed that NSM inhibition reduces EV nanoparticles and diminishes RNA and protein cargoes, including endosomal, spliceosomal, and translation-related proteins. Conversely, ASM inhibition increased RNA-binding proteins within and enhanced the expression of ribonucleoprotein complex-associated RNA in released EVs, including several snRNAs and 7SL RNA. Intriguingly, ASM-inhibited EVs enhanced the migration and translational activity of recipient MCF10A cells. These findings suggest an important role for SMase-dependent vesiculation in governing RNA and protein trafficking to the extracellular space, unveiling potential implications for cellular communication and function.

molecular biology↗

bioRxiv2024

Localized translation of cell junction mRNAs is required for epithelial cell polarity

Epithelial cells exhibit a highly polarized organization along their apico-basal axis, a feature that is critical to their function and frequently perturbed in cancer. One less explored process modulating epithelial cell polarity is the subcellular localization of mRNA molecules. In the present study, we report that several mRNAs encoding evolutionarily conserved epithelial polarity regulatory proteins, including Zo-1, Afdn and Scrib, are localized to cell junction regions in Drosophila epithelial tissues and human epithelial cells. Targeting of these mRNAs is coincident with the robust junctional distribution of their encoded proteins, and we demonstrate that they are locally translated at cell junction regions. To identify RNA binding proteins (RBPs) potentially implicated in junctional mRNA regulation, we performed systematic immuno-labeling with a collection of validated RBP antibodies, identifying a dozen RBPs with consistent junctional distribution patterns, several of which directly bind junctional transcripts. Strikingly, depletion of these RBP candidates, including MAGOH, a core component of the exon-junction complex (EJC), perturbed the junctional distribution and localized translation of Zo-1 and Scrib mRNAs, as well as the junctional accumulation of their protein products. Functional disruption of MAGO, or its interaction partner Y14, in Drosophila follicular epithelial cells perturbs the distribution of junctional transcripts and proteins. Finally, tissue microarray analysis of ovarian cancer tumor specimens revealed that expression of MAGOH and ZO-1 is positively correlated and that both proteins are potential biomarkers of good prognosis. Altogether, this work reveals that localized mRNA translation at cell junction regions is important for modulating epithelial cell polarity. HIGHLIGHTSO_LICell junction mRNA targeting is conserved between tissues and species C_LIO_LIThese mRNAs undergo localized translation at areas of cell-cell contact C_LIO_LIA diversity of RBPs localize to cell junction regions and interact with junctional transcripts. C_LIO_LIDisruption of junctional RBPs impacts epithelial cell polarity and localized translation C_LIO_LIMAGOH and ZO-1 expression is correlated in ovarian tumor specimens and are potential biomarkers of good prognosis C_LI

molecular biology↗

bioRxiv2023

GAS6 and AXL promote insulin resistance by rewiring insulin signaling and increasing insulin receptor trafficking to late endosomes

Growth-arrest specific 6 (GAS6) is a secreted protein that acts as a ligand for TAM receptors (TYRO3, AXL and MERTK). In humans, GAS6 circulating levels and genetic variations in GAS6 are associated with hyperglycemia and increased risk of type 2 diabetes. However, the mechanisms by which GAS6 influences glucose metabolism are not understood. Here, we show that Gas6 deficiency in mice increases insulin sensitivity and protects from diet-induced insulin resistance. Conversely, increasing GAS6 circulating levels is sufficient to reduce insulin sensitivity in vivo. GAS6 inhibits the activation of the insulin receptor (IR) and reduces insulin response in muscle cells in vitro and in vivo. Mechanistically, AXL and IR fom a complex, while GAS6 reprograms signaling pathways downstream of IR. This results in increased IR endocytosis following insulin treatment. This study contributes to a better understanding of the cellular and molecular mechanisms by which GAS6 and AXL influence insulin sensitivity. ARTICLE HIGHLIGHTSO_LIGAS6 deficient mice are characterized by improved glucose tolerance, increased insulin sensitivity and are protected from diet-induced insulin resistance. C_LIO_LIThe GAS6 receptor AXL is expressed in muscle cells and forms a complex with the insulin receptor. C_LIO_LIGAS6 reprograms insulin signaling by modulating the phosphorylation of proteins implicated in endocytosis, vesicle-mediated transport, and membrane trafficking. C_LIO_LIAXL and GAS6 promotes insulin receptor endocytosis following insulin treatment. C_LI

physiology↗

bioRxiv2023

The hGID GID4 E3 ubiquitin ligase complex targets ARHGAP11A to regulate cell migration

The human CTLH/GID (hGID) complex emerged as an important E3 ligase regulating multiple cellular processes, including cell cycle progression and metabolic activity. However, the range of biological functions controlled by hGID remains unexplored. Here, we show that the hGID substrate receptor GID4 regulates cell growth and migration. Biochemical and cellular assays combined with proximity-dependent biotinylation (BioID2) revealed that the hGIDGID4 E3-ligase targets the Rho-GAP ARHGAP11A for degradation. Depletion of GID4 or impeding the GID4 substrate binding pocket impairs motility and directed cell movement, whereas knockdown of ARHGAP11A significantly restores the cell migration defect. We found that GID4 controls cell migration by degrading ARHGAP11A thereby preventing its accumulation at the cell periphery where it inactivates RhoA activity. Together, we identified a unique function for GID4, as well as a wide range of substrate profiles beyond Pro/N-degron motifs, which pave the way for deciphering additional pathways regulated by hGID E3 ligase activity through its GID4 substrate receptor.

biochemistry↗

bioRxiv2023

Mapping the MOB proteins' proximity network reveals a unique interaction between human MOB3C and the RNase P complex

Distinct functions mediated by members of the monopolar spindle-one-binder (MOB) family of proteins remain elusive beyond the evolutionary conserved and well-established roles of MOB1A and B in regulating the Hippo pathway. Since MOB proteins are adaptors, understanding how they engage in protein-protein interactions and complexes assembly is essential to define the full scope of their biological functions. To address this, we undertook a proximity-dependent biotin identification (BioID) approach to define the interactomes of all seven human MOB proteins in HeLa and HEK293 cell lines. We uncovered > 200 interactions, of which at least 70% are unreported on BioGrid. The generated dataset reliably recalled the bona fide interactors of the well-studied MOBs. We further defined the common and differential interactome between different MOBs on a subfamily and an individual level. We discovered a unique interaction between MOB3C and 7 out of 10 protein subunits of the RNase P complex, an endonuclease that catalyzes tRNA 5 maturation. As a proof-of-principle for the robustness of the generated dataset, we validated the specific interaction of MOB3C with catalytically active RNase P by using affinity purification-mass spectrometry and pre-tRNA cleavage assays of MOB3C pulldowns. In summary, our data provide novel insights into the biology of MOB proteins and reveal the first interactors of MOB3C, components of the RNase P complex, and hence an exciting nexus with RNA biology.

biochemistry↗

bioRxiv2023

ARF-family global interactome mapping uncovers spatial organization of cellular signaling pathways

The ADP-ribosylation factors (ARFs) and ARF-like (ARLs) GTPases serve as essential molecular switches governing a wide array of cellular processes. In this study, we utilized proximity-dependent biotin identification (BioID) to comprehensively map the interactome of 28 out of 29 ARF and ARL proteins in two cellular models. Through this approach, we identified [~]3000 high-confidence proximal interactors, enabling us to assign subcellular localizations to the family members. Notably, we uncovered previously undefined localizations for ARL4D and ARL10. Clustering analyses further exposed the distinctiveness of the interactors identified with these two GTPases. We also reveal that the expression of the understudied member ARL14 is confined to the stomach and intestines. We identified phospholipase D1 (PLD1) and the ESCPE-1 complex, more precisely SNX1, as proximity interactors. Functional assays demonstrated that ARL14 can activate PLD1 in cellulo and is involved in cargo trafficking via the ESCPE-1 complex. Overall, the BioID data generated in this study provide a valuable resource for dissecting the complexities of ARF and ARL spatial organization and signaling. SUMMARY STATEMENTGeneration of the ARF family interactome allowed the attribution of potential localizations and functions to previously understudied members. We found that ARL14 activates PLD1 and contributes to ESCPE-1-mediated trafficking.

molecular biology↗

bioRxiv2022

C1orf112 is a novel regulator of interstrand crosslink that decreases FIGNL1-RAD51 interaction

Interstrand DNA crosslinks (ICLs) represent complex lesions that block essential biological processes, including DNA replication, recombination, and transcription. Several pathways have been involved in ICL repair, in particular nucleotide excision repair (NER), translesion DNA synthesis (TLS), Fanconi anemia (FA), and homologous recombination (HR). Still, the extent of factors involved in the resolution of ICL-induced DNA double-strand breaks (DSBs) remains poorly defined. Using CRISPR-based genome-wide screening, we identified the poorly characterized C1orf112 (also known as Apolo1) as a novel sensitizer to the clinically relevant ICL-inducing agent mafosfamide. Consistently, we noted that low expression of C1orf112 correlates with increased sensitivity to a series of ICL agents and PARP inhibitors in a panel of cell lines. We showed that lack of C1orf112 does not impact the initial recruitment and ubiquitylation of FANCD2 at the ICL site but rather impairs the resolution of RAD51 from ICL-induced DSBs, thereby compromising homology-directed DNA repair pathways. Our proximal mapping of C1orf112 protein neighbours coupled to structure-function analysis revealed that C1orf112, through its WCF motif, forms a complex with the N-terminal domain of the AAA+ ATPase FIGNL1 and regulates the interaction of FIGNL1 with RAD51. Our work establishes the C1orf112-FIGNL1 complex as an integral part of the HR-mediated response to ICLs by regulating the unloading of RAD51 during ICL repair.

molecular biology↗

bioRxiv2020

ARL15 modulates magnesium homeostasis through N-glycosylation of CNNMs

Cyclin M (CNNM1-4) proteins maintain cellular and body magnesium (Mg2+) homeostasis. Using various biochemical approaches, we have identified members of the CNNM family as direct interacting partners of ADP-ribosylation factor-like protein 15 (ARL15), a small GTP-binding protein. ARL15 interacts with CNNMs at their carboxyl-terminal conserved cystathionine-{beta}-synthase (CBS) domains. In silico modeling of the interaction using the reported structures of both CNNM2 and ARL15 supports that the small GTPase specifically binds the CBS1 domain. Immunocytochemical experiments demonstrate that CNNM2 and ARL15 co-localize in the kidney, with both proteins showing subcellular localization in the Golgi-apparatus. Most importantly, we found that ARL15 is required for forming complex N-glycosylation of CNNMs. Overexpression of ARL15 promotes complex N-glycosylation of CNNM3. Mg2+ uptake experiments with a stable isotope demonstrate that there is a significant increase of 25Mg2+ uptake upon knockdown of ARL15 in multiple kidney cancer cell lines. Altogether, our results establish ARL15 as a novel negative regulator of Mg2+ transport by promoting the complex N-glycosylation of CNNMs.

biochemistry↗

bioRxiv2020

APOBEC2 binds Chromatin and Represses Transcription during Myoblast Differentiation

The activation induced cytidine deaminase/apolipoprotein B editing complex (AID/APOBEC) family comprises several nucleic acid editors with roles ranging from antibody diversification to mRNA editing. APOBEC2, an evolutionarily conserved member of this family, has neither an established substrate nor a mechanism of action, however genetic evidence suggests functional relevance in tissues such as muscle. Here, we demonstrate that in muscle, APOBEC2 does not have any of the attributed molecular functions of the AID/APOBEC family, such as RNA editing, DNA demethylation, or DNA mutation. Instead, we show that APOBEC2 occupies chromatin at promoter regions of certain genes, whose expression is repressed during muscle cell differentiation. We further demonstrate that APOBEC2 on one hand binds promoter region DNA directly and in a sequence specific fashion, while on the other it interacts with HDAC transcriptional corepressor complexes. Therefore, APOBEC2, by actively repressing the expression of non-myogenesis pathway genes, plays a key role in enforcing the proper establishment of muscle cell fate.

molecular biology↗

bioRxiv2020

DNA binding reorganizes the intrinsically disordered C-terminal region of PSC in Drosophila PRC1

Polycomb Group (PcG) proteins regulate gene expression by modifying chromatin. A key PcG complex, Polycomb Repressive Complex 1 (PRC1), has two activities: a ubiquitin ligase activity for histone H2A, and a chromatin compacting activity. In Drosophila, the Posterior Sex Combs (PSC) subunit of PRC1 is central to both activities. The N-terminal homology region (HR) of PSC assembles into PRC1, including partnering with dRING to form the ubiquitin ligase for H2A. The intrinsically disordered C-terminal region of PSC (PSC-CTR) compacts chromatin, and inhibits chromatin remodeling and transcription in vitro. Both the PSC-HR and the PSC-CTR are essential in vivo. To understand how these two activities may be coordinated in PRC1, we used cross-linking mass spectrometry (XL-MS) to analyze the conformations of the PSC-CTR in PRC1 and how they change on binding DNA. XL-MS identifies interactions between the PSC-CTR and the core of PRC1, including between the PSC-CTR and PSC-HR. New contacts and overall more compacted PSC-CTR conformations are induced by DNA binding. Protein footprinting of accessible lysine residues in the PSC-CTR reveals an extended, bipartite candidate DNA/chromatin binding surface. Our data suggest a model in which DNA (or chromatin) follows a long path on the flexible PSC-CTR. Intramolecular interactions of the PSC-CTR detected by XL-MS can bring the high affinity DNA/chromatin binding region close to the core of PRC1 without disrupting the interface between the ubiquitin ligase and the nucleosome. Our approach may be applicable to understanding the global organization of other large IDRs that bind nucleic acids. HighlightsO_LIAn intrinsically disordered region (IDR) in Polycomb protein PSC compacts chromatin C_LIO_LICross-linking mass spectrometry (XL-MS) was used to analyze topology of the PSC IDR C_LIO_LIProtein footprinting suggests a bipartite DNA binding surface in the PSC IDR C_LIO_LIA model for the DNA-driven organization of the PSC IDR C_LIO_LICombining XL-MS and protein footprinting is a strategy to understand nucleic acid binding IDRs C_LI O_FIG O_LINKSMALLFIG WIDTH=149 HEIGHT=200 SRC="FIGDIR/small/130492v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@346c5dorg.highwire.dtl.DTLVardef@1b672b3org.highwire.dtl.DTLVardef@1247bfdorg.highwire.dtl.DTLVardef@48448e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2019

PGC-1α isoforms coordinate to balance hepatic metabolism and apoptosis in inflammatory environments

Liver is regularly exposed to changing metabolic and inflammatory environments. It must sense and adapt to metabolic need while balancing resources required to protect itself from insult. PGC-1 is a transcriptional coactivator that both coordinates metabolic adaptation to diverse stimuli and protects against inflammation. However, it is not known how PGC-1 integrates extracellular signals to balance metabolic and anti-inflammatory outcomes. PGC-1 exists as multiple, alternatively spliced variants expressed from different promoters. Primary mouse hepatocytes were used to evaluate the role(s) of different PGC-1 proteins in regulating hepatic metabolism and inflammatory signaling downstream of TNF. PGC-11 and PGC-14 were expressed in hepatocytes and expression analysis uncovered shared and isoform-specific roles for these variants linked to metabolism and inflammation. PGC-11 primarily impacted gene programs of nutrient and mitochondrial metabolism, while TNF signaling revealed that PGC-14 influenced several pathways related to innate immunity and cell death. Gain- and loss-of-function models illustrated that PGC-14 uniquely enhanced expression of anti-apoptotic gene programs and attenuated hepatocyte apoptosis in response to TNF or LPS. This was in contrast to PGC-11, which reduced the expression of a wide inflammatory gene network, but did not prevent liver cell death in response to the cytokine. We conclude that PGC-1 variants have distinct, yet complimentary roles in hepatic responses to metabolism and inflammation and identify PGC-14 as an important mitigator of apoptosis.

molecular biology↗

bioRxiv2018

Natural selection contributed to immunological differences between human hunter-gatherers and agriculturalists

The shift from a hunter-gatherer (HG) to an agricultural (AG) mode of subsistence is believed to have been associated with profound changes in the burden and diversity of pathogens across human populations. Yet, the extent to which the advent of agriculture may have impacted the evolution of the human immune system remains unknown. Here we present a comparative study of variation in the transcriptional responses of peripheral blood mononuclear cells to bacterial and viral stimuli between Batwa rainforest hunter-gatherers and Bakiga agriculturalists from Uganda. We observed increased divergence between hunter-gatherers and agriculturalists in the transcriptional response to viruses compared to that for bacterial stimuli. We demonstrate that a significant fraction of these transcriptional differences are under genetic control, and we show that positive natural selection has helped to shape population differences in immune regulation. Across the set of genetic variants underlying inter-population immune response differences, however, the signatures of positive selection were disproportionately observed in the rainforest hunter-gatherers. This result is counter to expectations based on the popularized notion that shifts in pathogen exposure due to the advent of agriculture imposed radically heightened selective pressures in agriculturalist populations.

evolutionary biology↗