Biology subjects

Coute, Y.

Publications and source records attributed to Coute, Y..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Nucleocytoviricota viral factories are transient organelles made by phase separation

Phase separation is a widespread mechanism in viral processes, mediating replication, host manipulation and virion morphogenesis. The phylum Nucleocytoviricota encompasses diverse and ubiquitous viruses, including Poxviridae, the climate-modulating Emiliania huxleyi virus and other so-called Giant Viruses. Cytoplasmic members of this phylum form viral factories but their nature has remained unresolved. Here, we demonstrate that these viral factories are formed by liquid-liquid phase separation. We prove that mimivirus viral factories are formed by multilayered phase separation, orchestrated by at least two scaffold proteins. To extend these findings across the phylum Nucleocytoviricota, we developed a bioinformatic pipeline to predict scaffold proteins based on a conserved molecular grammar, despite major primary sequence variability. Scaffold candidates were validated in Marseilleviridae and Poxviridae, highlighting a role of H5 as a scaffold protein in the vaccinia virus. Finally, we provide a repertoire of client proteins of the nucleus-like viral factory of mimivirus and demonstrate important sub-compartmentalization of functions, including those related to the central dogma. Overall, we reveal a new mechanism for an organelle to deploy nuclear-like functions entirely based on phase separation and re-classified phylum Nucleocytoviricota viral factories as biomolecular condensates.

microbiology↗

bioRxiv2024

Proteomic profiling of UV damage repair patches uncovers histone chaperones with central functions in chromatin repair

DNA damage compromises not only genome stability but also the integrity of the chromatin template, which plays a central role in controlling cell identity. Our understanding of chromatin repair mechanisms is very incomplete. To bridge this knowledge gap, here we devise a novel proteomic strategy to characterize dynamic changes in the chromatin landscape during the repair of UV-induced DNA lesions in human cells, in a quantitative, unbiased and time-resolved manner. Thus, we identify the histone chaperones DNAJC9 and MCM2 as central players in chromatin repair. We demonstrate that DNAJC9 and MCM2 are independently recruited to sites of UV damage repair. DNAJC9 provides new H3-H4 histones to CAF-1 and HIRA chaperones for deposition into chromatin and also stimulates old H3-H4 histone recovery. DNAJC9 cooperates with MCM2 to coordinate old and new histone dynamics during UV damage repair. Together, our proteomic dataset provides a molecular framework for further dissecting epigenome maintenance mechanisms.

molecular biology↗

bioRxiv2024

AtVPS13M1 is involved in lipid remodeling in low phosphate and is located at the mitochondria surface in plants

VPS13 are conserved lipid transporters with multiple subcellular localizations playing key roles in many fundamental cellular processes. While the localization and function of VPS13 have been extensively investigated in yeast and animals, little is known about their counterparts in plants, particularly regarding their role in stress response. In this study, we characterized AtVPS13M1, one of the four VPS13 paralogs of the flowering plant Arabidopsis thaliana. We show that AtVPS13M1 binds and transports glycerolipids with a low specificity in vitro. AtVPS13M1 interferes with phospholipids degradation in response to phosphate starvation, a nutrient stress that triggers a massive remodeling of membrane lipids. AtVPS13M1 is mainly expressed in young dividing and vascular tissues. Finally, we show that AtVPS13M1 is mainly located at the surface of mitochondria in leaves. Overall, our work highlights the conserved role in lipid transport of VPS13 in plants, reveals their importance in nutrient stress response and opens important perspectives for the understanding of lipid remodeling mechanisms and for the characterization of this protein family in plants.

cell biology↗

bioRxiv2024

Deciphering hepatitis B virus phospho-signature in primary human hepatocytes

Phosphorylation is a major post-translation modification (PTM) of proteins which is finely tuned by the activity of several hundred kinases and phosphatases. It controls most if not all cellular pathways including anti-viral responses. Accordingly, viruses often induce important changes in the phosphorylation of host factors that can either promote or counteract viral replication. Among more than 500 kinases constituting the human kinome only few have been described as important for the Hepatitis B virus (HBV) infectious cycle, and most of them intervene during early or late infectious steps by phosphorylating the viral Core protein (HBc) protein. In addition, little is known on the consequences of HBV infection on the activity of cellular kinases. The objective of this study was to investigate the global impact of HBV infection on the cellular phosphorylation landscape early after infection. For this, primary human hepatocytes (PHHs) were challenged or not with HBV, and a mass spectrometry (MS)-based quantitative phosphoproteomic analysis was conducted two- and seven-days post-infection. The results indicated that while, as expected, HBV infection only minimally modified the cell proteome, significant changes were observed in the phosphorylation state of several host proteins at both times points. Gene enrichment and ontology analyses of up- and down-phosphorylated proteins revealed common and distinct signatures induced by infection. In particular, HBV infection resulted in up-phosphorylation of proteins involved in DNA damage signaling and repair, RNA metabolism, in particular splicing, and cytoplasmic cell-signaling. Down-phosphorylated proteins were mostly involved in cell signaling and communication. Validation studies carried out on selected up-phosphorylated proteins, revealed that HBV infection induced a DNA damage response characterized by the appearance of 53BP1 foci, the inactivation of which by siRNA increased cccDNA levels. In addition, among up-phosphorylated RNA binding proteins (RBPs), SRRM2, a major scaffold of nuclear speckles behaved as an antiviral factor. In accordance with these findings, kinase prediction analysis indicated that HBV infection upregulates the activity of major kinases involved in DNA repair. These results strongly suggest that HBV infection triggers an intrinsic anti-viral response involving DNA repair factors and RBPs that contribute to reduce HBV replication in cell culture models.

microbiology↗

bioRxiv2024

Proteomics-based characterization of ribosome heterogeneity in adult mouse organs

While long thought to be invariable in all cellular organisms, evidence has emerged that the translation process, i.e. protein assembly from mRNA sequence decoding, is regulated by variable features of the translation machinery. Notably, ribosomes, the functional units of protein synthesis, display variations in their composition, depending on the developmental stage, cell type or physiopathological context, thus providing a new level of actionable regulation of gene expression. Yet, a comprehensive map of the heterogeneity of ribosome composition in ribosomal proteins (RPs) in different organs and tissues is not available. In this work, we explored tissue-specific ribosome heterogeneity using mass spectrometry-based quantitative proteomic characterization of ribosomal fractions purified from 14 adult mouse organs and tissues. We performed crossed clustering and statistical analyses of RP composition to highlight stable, variable and tissue-specific RPs across organs and tissues. Focusing on specific RPs, we validated their relative abundance with a targeted proteomic approach, which gave further insight into the tissue-specific ribosome RP signature. Finally, we investigated the origin of RP variations in ribosome fraction of the different tissues, by comparing RP relative abundances in our proteomic dataset and in three independent transcriptomic datasets. Interestingly, we found that, in some tissues, the RP abundance in purified ribosomes does not always correlate with the corresponding RP transcript level, arguing for a translational regulation of RP expression, and/or a regulated incorporation of RPs into ribosomes. Altogether, our data support the notion of a tissue-specific RP signature of ribosomes, which opens avenues to study how specific ribosomal composition provides an additional level of regulation to control gene expression in different tissues and organs.

biochemistry↗

bioRxiv2024

Photoreceptor-induced LHL4 protects photosystem II in Chlamydomonas reinhardtii

Photosynthesis, the fundamental process using light energy to convert CO2 to organic matter, is vital for life on Earth. It relies on capturing light through light-harvesting complexes in evolutionarily well-conserved photosystems (PS) I and II and on the conversion of light energy into chemical energy. Composition and organization of both photosystem core complexes are well conserved across evolution. PSII is particularly sensitive to photodamage but benefits from a large diversity of photoprotective mechanisms, finely tuned for the specific light conditions. Light Harvesting Complex protein family members (LHC and LHC-like families) have acquired a dual function during evolution. Members of the LHC antenna complexes of photosystems capture light energy whereas others dissipate excess energy that cannot be harnessed for photosynthesis. This process mainly occurs through non photochemical quenching (NPQ). In this work, we focus on the LHL4 protein, which is a LHC-like protein induced by UV-B and blue light photoreceptor signaling pathways in the model green microalgae Chlamydomonas reinhardtii. We demonstrate that alongside established NPQ effectors, LHL4 plays a key role in photoprotection, preventing singlet oxygen accumulation in PSII and promoting cell survival upon light stress. LHL4 protective function is distinct from that of NPQ-related proteins, as it specifically and uniquely binds to the transient monomeric form of the core PSII complex, safeguarding its integrity. LHL4 characterization expands our understanding of the interplay between light harvesting and photoprotection mechanisms upon light stress in photosynthetic microalgae.

plant biology↗

bioRxiv2024

Phototropin connects blue light perception to starch metabolism in green algae

In photosynthetic organisms light acts as an environmental signal to control their development and physiology, and as energy source to drive the conversion of CO2 into carbohydrates used for growth or storage. The main storage carbohydrate in green algae is starch, which accumulates during the day and is broken down at night to meet cellular energy demands. The signalling role of light quality in the regulation of starch accumulation remains unexplored. Here, we identify PHOTOTROPIN-MEDIATED SIGNALLING KINASE 1 (PMSK1) as a key regulator of starch metabolism in Chlamydomonas reinhardtii. In its phosphorylated form (PMSK1-P), it activates GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE (GAP1), promoting starch biosynthesis. We show that blue light, perceived by PHOTOTROPIN, induces PMSK1 dephosphorylation that in turn represses GAP1 mRNA levels and reduces starch accumulation. These findings reveal a novel blue light-mediated signaling pathway that advances our understanding of photoreceptor-controlled carbon metabolism in microalgae. One-Sentence SummaryBlue light perception by PHOTOTROPIN triggers kinase-mediated signaling to inhibit starch accumulation in the green alga Chlamydomonas.

plant biology↗

bioRxiv2023

The α/β hydrolase domain-containing protein 1 (ABHD1) acts as a lysolipid lipase and is involved in lipid droplet formation

Lipid droplets (LDs) are the major sites of lipid and energy homeostasis. However, few LD biogenesis proteins have been identified. Here, using Chlamydomonas as a model, we show that ABHD1, a member of the /{beta} hydrolase domain-containing protein family, is a novel type of LD-associated protein which stimulates LD formation through two distinct actions on the LD surface, one enzymatic and the other structural. ABHD1 was localized to LD surface in Chlamydomonas cells. The knockout mutants contained similar amounts of triacylglycerols (TAG) but their LDs showed an increased content in lyso- derivatives of the betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS). Over-expression of ABHD1 in Chlamydomonas induced LD formation and boosted TAG content, suggesting a key role in LD biogenesis. The purified recombinant ABHD1 protein hydrolyzed lyso-DGTS, producing a free fatty acid and a glyceryltrimethylhomoserine moiety. In vitro experiments using droplet- embedded vesicles showed that ABHD1 promoted LD emergence. Taken together, these results identify ABHD1 as a new player in LD formation by its lipase activity on lyso-DGTS and by its distinct biophysical property. This study further suggests that lipases targeted to LDs and able to act on their polar lipid coat may be interesting tools to promote LD assembly in eukaryotic cells. Significant statementLipid droplets are subcellular organelles specialized for triacylglycerol storage. Their dynamic turnover is key to managing energy homeostasis in response to cell cycle states and environmental cues. To gain insights into LD biogenesis, we characterized a putative /{beta}- hydrolase (ABHD1) in the model algae Chlamydomonas reinhardtii and show it is located at the LD surface. We found that ABHD1 overexpression promotes LD formation and acts as a lipase mainly on lyso derivatives of the betaine lipid diacylglyceryl-N,N,N-trimethylhomoserine (DGTS), the major lipid constituent of the LD hemi-membrane. We also show that ABHD1 has a remarkable biophysical property favoring LD budding. This work thus identifies a novel type of lipase acting on betaine lipid and provides a first example of a protein with a dual function nvolved in LD formation.

biochemistry↗

bioRxiv2023

Cytoskeleton remodeling induced by SMYD2 methyltransferase drives breast cancer metastasis

Malignant forms of breast cancer refractory to existing therapies remain a major unmet health issue, primarily due to metastatic spread. A better understanding of the mechanisms at play will provide better insights for alternative treatments to prevent breast cancer cells dispersion. Here, we identify the lysine methyltransferase SMYD2 as a clinically actionable master regulator of breast cancer metastasis. While SMYD2 is overexpressed in aggressive breast cancers, we notice that it is not required for primary tumor growth. However, mammary-epithelium specific SMYD2 ablation increases mouse overall survival by blocking the primary tumor cells ability to metastasize. Mechanistically, we identify BCAR3 as a genuine physiological substrate of SMYD2 in breast cancer cells. BCAR3 monomethylated at lysine K334 (K334me1) is recognized by a novel methyl-binding domain present in FMNLs proteins. These actin cytoskeleton regulators are recruited at the cell edges by the SMYD2 methylation signaling and modulates lamellipodia properties. Breast cancer cells with impaired BCAR3 methylation loose migration and invasiveness capacity in vitro and are ineffective in promoting metastases in vivo. Remarkably, SMYD2 pharmacologic inhibition efficiently impairs the metastatic spread of breast cancer cells, PDX and aggressive mammary tumors from genetically engineered mice. This study provides a rationale for innovative therapeutic prevention of malignant breast cancer metastatic progression by targeting the SMYD2-BCAR3-FMNL axis.

cancer biology↗

bioRxiv2023

Comparative transcriptomics reveal a novel tardigrade specific DNA binding protein induced in response to ionizing radiation

Tardigrades, microscopic animals found in virtually all ecosystems, are renowned for their remarkable ability to withstand extreme conditions. Recent studies have identified novel tardigrade specific protein families that aid in resistance to desiccation and ionizing radiation (IR). Notably, a tardigrade specific DNA binding protein called Dsup (for DNA damage suppressor) has been found to protect from X-ray damage in human cells and from hydroxyl radicals in vitro. However, Dsup has only been found in two species within the Hypsibioidea superfamily. To better understand mechanisms underlying radio-resistance in the Tardigrada phylum, we first characterized DNA damage and repair in response to IR in the model species Hypsibius exemplaris. By analysis of phosphorylated H2AX, we demonstrated the induction and repair of DNA double-strand breaks after IR exposure. Importantly, the rate of single-strand breaks induced was roughly equivalent to that in human cells, suggesting that DNA repair plays a predominant role in the remarkable radio-resistance of tardigrades. In order to identify novel tardigrade specific genes involved, we next conducted a comparative transcriptomics across three species, H. exemplaris, Acutuncus antarcticus and Paramacrobiotus fairbanksi, the latter belonging to the Macrobiotoidea superfamily known to lack Dsup homologs. In all three species, many genes of DNA repair were among the most strongly overexpressed genes alongside a novel tardigrade specific gene, named Tardigrade DNA damage Response protein 1 (TDR1). We found that TDR1 protein interacts with DNA and forms aggregates at high concentration suggesting it may condensate DNA and act by preserving chromosome organization until DNA repair is accomplished. Remarkably, when expressed in human cells, TDR1 improved resistance to Bleomycin, a radiomimetic drug. Based on these findings, we propose that TDR1 is a novel tardigrade specific gene responsible for conferring resistance to IR. Our study sheds light on mechanisms of DNA repair helping to cope with high levels of DNA damage. Furthermore, it suggests that at least two tardigrade specific genes, respectively for Dsup and TDR1, have independently evolved DNA-binding functions that contribute to radio-resistance in the Tardigrada phylum.

molecular biology↗

bioRxiv2023

Huntingtin bundles and changes the local proteome of actin filaments in neurons

Huntingtin (HTT) is a large protein whose best-known function being the facilitation of intracellular dynamics along the microtubule network by scaffolding molecular motors complexes. Our recent finding that the defective axonal growth in HD was due to altered growth cone architecture led us to ask whether HTT also influences the cytoskeleton itself. In developing neurons, we found that a large proportion of HTT associates with F-actin in growth cones. Using cell free system and purified recombinant proteins, we observed that HTT binds directly filamentous actin (F-actin) and organizes filaments into bundles. Transmission electron microscopy shows that HTT dimers crosslink adjacent filaments 20 nm apart. We also provide evidence that HTT binding on F-actin modulates the association of other proteins to this cytoskeleton. Notably, HTT limits the association of the growth cone protein Drebrin1 with F-actin. HTT depletion leads to abnormal cytoskeletal organization, localization of Drebrin1 in growth cones, and axonal growth. HTT therefore serves a scaffolding function for the cytoskeleton itself, what might be relevant for HD pathophysiology.

cell biology↗

bioRxiv2023

Pseudomonas aeruginosa MipA-MipB envelope proteins act as new sensors of polymyxin

Due to the rising incidence of antibiotic resistant infections, the last-line antibiotics polymyxins have resurged in the clinics in parallel with new bacterial strategies of escape. The Gram-negative opportunistic pathogen Pseudomonas aeruginosa develops resistance to colistin/polymyxin by distinct molecular mechanisms, mostly through modification of the lipid A component of the LPS by proteins encoded within the arnBCDATEF-ugd (arn) operon. In this work, we characterized a polymyxin-induced operon, named mipBA, notably present in P. aeruginosa strains devoid of the arn operon. We showed that mipBA is activated by ParR/ParS two-component regulatory system in response to polymyxin. MipA and MipB localize to bacterial outer membrane and form a complex in vitro. Structural modeling revealed that the lipoprotein MipB adopts a {beta}-lactamase fold with two additional C-terminal domains,while MipA folds as an outer-membrane {beta}-barrel, harboring an internal negatively charged channel, able to host a polymyxin molecule. Nano differential scanning fluorimetry (DSF) showed that polymyxin stabilized MipA protein in vitro. Mass spectrometry-based quantitative proteomics on whole bacterial membranes demonstrated that the {Delta}mipBA mutant synthesized less MexXY-OprA proteins in response to polymyxin compared to the wild-type strain, as a consequence of impaired transcriptional activation of the mex operon. We propose MipA/MipB to act as membrane (co)sensors working in concert to activate ParS histidine kinase and help the bacterium to cope with polymyxin-mediated envelope stress through synthesis of the efflux pomp, MexXY-OprA.

microbiology↗

bioRxiv2023

Molecular crosstalk between plant translation initiation complexes influences the outcome of virus infection

Successful subversion of translation initiation factors 4E and 4G determines the infection success of potyviruses, the largest group of viruses affecting plants. In the natural variability of many plant species, resistance to potyvirus infection is provided by polymorphisms at 4E and 4G that renders them inadequate for virus hijacking but still functional in translation initiation. In crops where such natural resistance alleles are limited, the genetic inactivation of 4E has been proposed for the engineering of potyvirus resistance. However, recent findings indicate that knockout 4E and 4G alleles may be deleterious for plant health and could jeopardize resistance efficiency in comparison to functional resistance proteins. Here, we explored the cause of these adverse effects by studying the role of the Arabidopsis eIF4E1, whose inactivation was previously reported as conferring resistance to the potyvirus clover yellow vein virus (ClYVV) while also promoting susceptibility to another potyvirus called turnip mosaic virus (TuMV). We report that eIF4E1 is required to maintain global plant translation and to restrict TuMV accumulation during infection, and its absence is associated with a favoured virus multiplication over host translation. Furthermore, our findings demonstrate that eIF4E1 plays a crucial role in inhibiting the TuMV-induced degradation of the translation initiation factor eIFiso4G1, thereby preventing the generation of a truncated protein. Finally, we demonstrate a role for eIFiso4G1 in TuMV accumulation and in supporting plant fitness during infection. These findings suggest that eIF4E1 counteracts the hijacking of the plant translational apparatus during TuMV infection and underscore the importance of preserving the functionality of translation initiation factors 4E and 4G when implementing potyvirus resistance strategies. Author summaryPlants are constantly under threat from viruses that can damage crops and reduce yield. Among these viruses, potyviruses are a major concern, and a small group of genes known as eIF4E are key factors in making a plant susceptible to them. To combat these viruses, it is possible to either use naturally-selected variants of eIF4E that provide resistance, or to disable the gene altogether. However, new research has shown that inactivating eIF4E genes may have unintended consequences for the plants development while compromise resistance to other potyviruses. To investigate this further, we focus in this work on the role of the Arabidopsis eIF4E1 whose inactivation confers resistance to one potyvirus, clover yellow vein virus (ClYVV). We looked why this same mutation at eIF4E1 makes the plants more susceptible to another potyvirus, turnip mosaic virus (TuMV). Our study reveals that eIF4E1 acts in safeguarding the plant translational machinery during TuMV infection. By preventing the degradation of the translation initiation protein eIFiso4G1, eIF4E1 enables the plant to maintain its normal translation activity and ultimately prevents the accumulation of virus proteins. Our findings provide valuable insights into how potyviruses hijack the plants translation process, and emphasizes the need of preserving the functionality of translation initiation factors when developing potyvirus resistances.

plant biology↗

bioRxiv2023

Knockout of GMC-oxidoreductase genes reveals functional redundancy in mimivirus

The mimivirus 1.2Mb genome was shown to be organized into a nucleocapsid-like genomic fiber encased in the nucleoid compartment inside the icosahedral capsid (1). The genomic fiber protein shell is composed of a mixture of two GMC-oxidoreductase paralogs, one of them being the main component of the glycosylated layer of fibrils at the surface of the virion (2). In this study, we determined the effect of the deletion of each of the corresponding genes on the genomic fiber and the layer of surface fibrils. First, we deleted the GMC-oxidoreductase the most abundant in the genomic fiber, and determined its structure and composition in the mutant. As expected, it was composed of the second GMC-oxidoreductase and contained 5- and 6-start helices similar to the wild-type fiber. This result led us to propose a model explaining their coexistence. Then, we deleted the GMC-oxidoreductase the most abundant in the layer of fibrils to analyze its protein composition in the mutant. Second, we showed that the fitness of single mutants and the double mutant were not decreased compared to the wild-type viruses in laboratory conditions. Third, we determined that deleting the GMC-oxidoreductase genes did not impact the glycosylation or the glycan composition of the layer of surface fibrils, despite modifying their protein composition. Since the glycosylation machinery and glycan composition of members of different clades are different (3, 4), we expanded the analysis of the protein composition of the layer of fibrils to members of the B and C clades and showed that it was different among the three clades and even among isolates within the same clade. Taken together, the results obtained on two distinct central processes (genome packaging and virion coating) illustrate an unexpected functional redundancy in members of the family Mimiviridae, suggesting this may be the major evolutionary force behind their giant genomes. One-Sentence SummaryFunctional redundancy preserves mimivirus genomic fiber and layer of fibrils formation.

microbiology↗

bioRxiv2023

Oxidation-sensitive cysteines drive IL-38 amyloid formation

Cytokines of the interleukin (IL)-1 family are widely expressed in epithelial surfaces, including the epidermis, where they play a key role in the maintenance of barrier integrity and host defense. A recent report associated the IL-1 family member IL-33 with stress granules (SGs) in epithelial cells. Formation of SGs is promoted by the aggregation of proteins harboring low complexity regions (LCRs). In this study, using computational analyses, we predicted the presence of LCRs in six of the eleven IL-1 family members. Among these, IL-38 contained a long LCR and localized to Ras GTPase-activating protein binding protein 1 (G3BP1) positive SGs, as well as to G3BP1 negative intracellular protein condensates in keratinocytes exposed to oxidative stress (OS). In addition, we identified two highly aggregation-prone amyloid core (AC) motifs in the IL-38 LCR and detected the formation of amyloid IL-38 aggregates in response to OS in cells and in vitro. Disulfide bond mapping, in silico modelling and the analysis of specific cysteine mutants supported a model in which specific oxidation-sensitive cysteines act as redox switches to modify the conformation of IL-38 and thus the surface exposure of its ACs, shuttling it from a soluble state into biomolecular condensates. Finally, the presence of IL-38 granules in human epidermal layers highly exposed to environmental OS suggests that oxidation-induced formation of amyloid aggregates, as a previously unrecognized intrinsic biological property of IL-38, may be physiologically relevant at this epithelial barrier.

cell biology↗

bioRxiv2023

Cell type-specific control of secondary cell wall formation by Musashi-type translational regulators in Arabidopsis

Deciphering the mechanism of secondary cell wall/SCW formation in plants is key to understanding their development and the molecular basis of biomass recalcitrance. Although transcriptional regulation is essential for SCW formation, little is known about the implication of post-transcriptional mechanisms in this process. Here we report that two bonafide RNA-binding proteins homologous to the animal translational regulator Musashi, MSIL2 and MSIL4, function redundantly to control SCW formation in Arabidopsis. MSIL2/4 interactomes are similar and enriched in proteins involved in mRNA binding and translational regulation. MSIL2/4 mutations alter SCW formation in the fibers, leading to a reduction in lignin deposition, and an increase of 4-O-glucuronoxylan methylation. In accordance, quantitative proteomics of stems reveal an overaccumulation of glucuronoxylan biosynthetic machinery, including GXM3, in the msil2/4 mutant stem. We showed that MSIL4 immunoprecipitates GXM mRNAs, suggesting a novel aspect of SCW regulation, linking post-transcriptional control to the regulation of SCW biosynthesis genes.

plant biology↗

bioRxiv2023

SIN3 acts in distinct complexes to regulate the germline transcriptional program in C. elegans

The SIN3 transcriptional coregulator influences gene expression through multiple interactions that include histone deacetylases (HDACs). Haploinsufficiency and mutations in SIN3 are the underlying cause of Witteveen-Kolk syndrome and related intellectual disability (ID)/autism syndromes, emphasizing its key role in development. However, little is known about the diversity of its interactions and functions in developmental processes. Here we show that loss of SIN-3, the single SIN3 homologue in Caenorhabditis elegans, results in maternal effect sterility associated with deregulation of the germline transcriptome, including desilencing of X-linked genes. We identify at least two distinct SIN3 complexes containing specific HDACs, and show that they differentially contribute to fertility. Single cell smFISH reveals that in sin-3 mutants, the X chromosome becomes re-expressed prematurely and in a stochastic manner in individual germ cells. Furthermore, we identify histone residues whose acetylation increases in the absence of SIN3. Together, this work provides a powerful framework for the in vivo study of SIN3 and associated proteins.

genetics↗

bioRxiv2023

Proteomic analyses of the Arabidopsis cap-binding complex define core set and TOR-dependent protein components

The eukaryotic cap-binding complex (CBC) is a hub for regulations affecting mRNA behaviour including translation, degradation and storage. Beside the core eukaryotic translation initiation factors, other proteins, many of which are yet unknown, are thought to interact stably or transiently with the CBC depending on cell status. The prototype of these regulators is the animal eIF4E binding protein (4E-BP), a direct target of the TOR (Target of Rapamycin) kinase that competes with the cap-binding protein eIF4E, thus repressing translation. In plants, no functional homologs of 4E-BP have so far been characterized. In this work we performed several deep proteomic analyses of the Arabidopsis CBC after cap-affinity purification from wild-type plants. We also investigated the CBC in eIF4E mutant plants, Arabidopsis lines with lower TOR activity, or during infection with eIF4E-dependent potyviruses, conditions which are all affecting translation at the initiation level. These analyses allowed us to define a limited core set of CBC components, which were detected in all samples. Interestingly, we identified proteins, like AGO1 or VCS, which were always detected in conditions where either TOR or mRNA translation were reduced. Meta-analysis of these data revealed several new plant interactors of the CBC, potentially defining pathways related to mRNA stability and degradation, metabolism and viral life cycle. A search for eIF4E binding motifs identified several new potential 4E-BP relatives in plants.

plant biology↗