Biology subjects

Monti, M.

Publications and source records attributed to Monti, M..

At least 19 recordsLinked to original sources

bioRxiv2026

PHANES: an integrated resource of protein features for the human proteome

Protein analysis and machine-learning workflows increasingly rely on large-scale numerical representations of proteins. While protein language models provide powerful embeddings, their limited interpretability and high computational cost hinder their use in many biological applications. We present PHANES, a human proteome database and web server providing a comprehensive, biologically interpretable set of features for 20335 proteins. PHANES integrates sequence-derived physicochemical descriptors, structure-based and graph-theoretical features computed from AlphaFold predictions, thermostability indices, and tissue expression specificity. In addition to precomputed features for the human proteome, an integrated web server enables on-demand feature calculation for user-provided protein structures. PHANES supports data export, visualization and comparative analysis at the level of individual proteins and protein groups, bridging the gap between complex numerical representations and biological interpretation while providing an interpretable foundation for exploratory analyses and downstream machine-learning applications. PHANES is freely available at https://tools.tartaglialab.com/phanes.

bioinformatics↗

bioRxiv2026

Growth, metabolism, and the benefits of clock-enabled anticipation

Circadian clocks are found in myriad species across the tree of life. They are commonly assumed to provide a selective advantage by enabling organisms to anticipate daily environmental changes. It has proven remarkably difficult, however, to move beyond this general idea to explain precisely what advantage anticipation confers and to quantify how large this benefit is. Here, we address these challenges by developing a mathematical model of diurnally varying growth and metabolism in the cyanobacterium Synechococcus elongatus, which fixes carbon through photosynthesis during the day and stores a portion of it as glycogen for use during the night. Specifically, we extend the framework introduced by Hwa and coworkers (Scott et al., Science 330, 1099 (2010)), which quantifies the relationship between growth and proteome composition, to a circadian context. We find that the fastest growth is achieved when glycogen production during the day is minimized to the amount required to sustain essential nighttime processes; growth then occurs exclusively during the day, in agreement with experimental observations. However, the slow relaxation of the proteome, arising from the intrinsically slow growth rate of S. elongatus, severely constrains the implementation of this optimal strategy: In the absence of anticipation, the cell must grow at night in order to produce proteins essential for nighttime metabolism. In contrast, with anticipation enabled by a circadian clock, the bacterium can instead adjust its proteome late in the day, allowing it to avoid less efficient nighttime growth altogether, thereby significantly enhancing its growth rate.

microbiology↗

bioRxiv2026

Warming exacerbates fungal pathogenicity to drive physiological collapse and microbiome reorganization in octocorals

Corals are increasingly threatened by climate change, yet the interplay between ocean warming and fungal infection remains experimentally underexplored. Here, we demonstrate that thermal stress exacerbates the pathogenicity of the fungus Aspergillus sydowii, driving physiological collapse and functional reorganization in the octocoral Sclerophytum sp. We deployed a 34-day mesocosm heat-stress experiment and quantified performance, mortality, and taxonomic and functional microbiome shifts. Combined heat and fungal exposure proved highly lethal, causing 45% mortality compared to 7% under heat exposure alone. Re-isolation from diseased nubbins implicates A. sydowii as the causal agent. Metagenomic profiling indicated that physiological collapse was underpinned by a functional transition from mutualism to antagonism in the microbiome. Specifically, combined heat and fungal stress triggered a depletion of ankyrin- and WD40-repeat proteins - hallmarks of symbiotic stability - concurrent with a surge in genes involved in fungal cell wall degradation and secondary metabolite biosynthesis. While surviving holobionts recovered full photosynthetic efficiency after combined stress, their microbiomes did not revert to baseline. Instead, they assembled into a taxonomically distinct configuration characterized by enrichments of sulfate-reducers (Thermodesulfobacteriota) and thermotolerant phototrophs (Thermosynechococcales). This suggests that despite the rapid photosystem recovery, the holobiont retained a complex legacy of thermal and biotic stress across both its internal chemical microenvironment and its microbiome. These findings highlight the decoupled recovery processes of different holobiont components, demonstrating that even though some corals may survive severe climate-driven disease, they emerge as ecologically reorganized entities.

ecology↗

bioRxiv2026

Functional characterisation of an essential neo-chromosome III in Sc2.0 strain reveals opportunities and challenges for genome minimisation in Sc3.0

AbstractLarge-scale genome minimisation in eukaryotes remains a major challenge due to essential genes embedded within deletion-refractory regions and pervasive synthetic lethal interactions. Here, we address these limitations by engineering an essential neo-chromosome III that relocates all 14 essential genes from synthetic chromosome III onto a separate chromosome, thereby enabling further minimisation of synIII. To further expand design space, we created highly synthetic neo-chromosome variants with sequences absent from natural genomes. We refactored essential gene expression using both native and orthogonal promoter-terminator pairs from Saccharomyces paradoxus and S. eubayanus. Reporter assays showed that orthogonal regulatory elements largely recapitulate S. cerevisiae activity. Both architectures restored viability in essential gene deletion libraries, demonstrating robust cross-species complementation. Engineered linear and circular forms of essential neo-chromosomes were highly stable over 100 generations and supported a near wild-type phenotype. Relocating essential functions enabled SCRaMbLE-mediated deletion of previously inaccessible regions, substantially expanding the deletion landscape. To improve the screening efficiency of SCRaMbLEd strains, we developed a SCRaMbLE reporter, ERICA (Elementary Random Integration Cassette), a loxPsym-flanked URA3 cassette that integrates randomly and enables iterative selection. Nanopore sequencing confirmed complex rearrangements, including deletions of up to [~]40 kb and loss of essential loci. Together, this work establishes a modular and extensible platform for orthogonal essential gene engineering and SCRaMbLE-enabled genome reduction, providing key design principles for next-generation synthetic eukaryotic genomes. These findings have broad implications beyond yeast, providing transferable design principles for genome minimisation in more complex eukaryotic systems, including mammalian and human cells.

genomics↗

bioRxiv2026

Host tRNA modifications drive efficient translation of influenza A virus genome and impact host antiviral stress responses

Influenza A virus (IAV), as all other viruses, is completely dependent on the host translation machinery components, including host transfer RNAs (tRNAs), to effectively decode its genome. However, while the human genome is biased towards cytosine (C) and guanosine (G)-ending codons, the IAV genome is skewed towards adenine (A) and uridine (U)-ending codons. Nevertheless, translation of IAVs RNA genome is highly efficient. Here we show that host tRNA and tRNA epitranscriptome dynamics are important regulators of IAV RNA translation and host antiviral responses. We show that the levels of several tRNA modifications, including 5-methylcarboxymethyluridine (mcm5U34) and 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U34), and their cognate writers, vary over the course of IAV infection. Additionally, we demonstrate that a set of tRNAs are preferentially recruited to ribosomes upon IAV infection, in line with IAV codon usage requirements. We further show that loss of ELP3, the catalytic subunit of the elongator complex, which is involved in the catalysis of mcm5U34 and of mcm5s2U34, induces tRNA hypomodifications, impairs translation of codon biased IAV genes and triggers the integrated stress response (ISR), interfering with IAV propagation. Taken together, our results uncover the relevance of host tRNAs and their modifications for optimal expression of viral genomes and host antiviral responses, setting the tRNA epitranscriptome as a promising target for the development of host-based antiviral therapies.

molecular biology↗

bioRxiv2026

Pathogen priming reveals host immune training and microbiome conditioning in corals

UNSTRUCTURED ABSTRACTIn several species, vaccine-like approaches, where hosts are primed through controlled pathogen exposure, have proven effective in enhancing responses to subsequent infections. This principle remains unexplored in corals. Here, we demonstrate that chronic exposure to non-lethal concentrations of live or inactivated Vibrio coralliilyticus primes the coral holobiont to counter subsequent infections under heat stress. Non-primed corals experienced greater heat stress pathogen-driven bleaching and a significant decline in their photosynthetic efficiency compared to primed samples. These results were linked to microbiome conditioning and host gene expression modulation, including a layered, fine-tuned immune and cellular response to microbial invasion. This proof-of-concept challenges strictly innate immune responses in corals and positions immune priming and microbiome conditioning as integrated mechanisms of coral holobiont resilience. Together, these findings can contribute to redefine coral immunity concepts and lay the groundwork for developing new microbiome-based strategies to enhance coral health for reef conservation under climate change. LAY SUMMARYCoral reefs are increasingly threatened by rising seawater temperatures and disease. Unlike vertebrates, corals do not possess a classical adaptive immune system, so they have been thought to rely only on innate defenses. However, our study shows that corals may be more capable than previously believed. We demonstrate that exposing corals to non-lethal doses of a widespread bacterial pathogen can train them to better withstand subsequent infections under heat stress. Corals that were not pre-exposed suffered more pathogen caused bleaching and showed a stronger decline in physiological performance, while those that were primed were more resilient. This improved resistance appears to come from two coordinated processes. First, the corals associated bacterial community shifted in a way that seems to help protect the host. Second, the coral itself adjusted its gene expression, mounting a more effective and controlled response to infection. These findings suggest that corals may be able to remember past exposures and respond more effectively to future infections, even without a traditional adaptive immune system, providing a foundation for developing new strategies to support coral resilience.

microbiology↗

bioRxiv2026

Coral Probiotics Buffer Adjacent Ecosystem-Level Responses to Extreme Marine Heatwave

Probiotics can enhance coral thermal tolerance, yet their ecosystem-level effects remain unknown. Here, we present the first long-term in-situ test of whether coral-targeted probiotics influence adjacent cryptobenthic reef communities during a record marine heatwave. Probiotics were applied to Pocillopora favosa and Acropora spp. coral colonies for 18 months, spanning the fourth global bleaching event. Cryptobenthic communities were assessed using biomimetic monitoring structures integrating biodiversity surveys, molecular profiling, microbial network analyses, and metabolic assays. Before the heatwave, probiotic and control patches were comparable across structural, microbial, and functional metrics. Following thermal stress, control patches exhibited pronounced losses of cryptobenthic invertebrate abundance and taxonomic breadth, microbial network fragmentation, and net carbonate dissolution. In contrast, probiotic-treated patches retained higher biodiversity, cohesive microbial interaction architectures, and positive calcification. These findings demonstrate that coral-targeted probiotics can scale from host-level intervention to buffer adjacent ecosystem-level responses to extreme marine heatwaves under accelerating climate change. TeaserA coral-targeted probiotic strategy enhances multi-trophic resilience under heat stress.

microbiology↗

bioRxiv2026

HERCULES: an integrative deep-learning framework for predicting RNA-binding propensity and mutation effects at single-residue resolution

RNA-binding proteins (RBPs) regulate essential aspects of RNA metabolism, yet accurately identifying RNA-binding domains (RBDs) and quantifying the impact of sequence variation on RNA-binding ability remain challenging. Here, we present HERCULES (Hybrid framEwoRk for RNA-binding domain loCalization and mUtation anaLysis using physicochemical and languagE modelS), a unified sequence-based framework for simultaneous RBD localization, global RNA-binding propensity prediction and mutation effect assessment. HERCULES integrates a fine-tuned protein language model with an explicit residue-level physicochemical module, combining global contextual representations with local mutation-sensitive descriptors. On an independent test set, the HERCULES global score discriminates RBPs from non-RBPs with an AUROC of 0.86. At residue resolution, HERCULES outperforms state-of-the-art sequence-based predictors in identifying canonical, non-canonical and putative RBDs across Pfam-annotated proteins. Using a curated dataset of experimentally validated RNA-binding-disrupting mutations, HERCULES correctly classifies 87% of deleterious variants, including single-amino acid substitutions. Evaluation on experimentally resolved protein-RNA complexes further demonstrates robust residue-level performance and improved generalization when contact annotations are augmented with AlphaFold3-predicted complexes. By unifying domain localization and mutation sensitivity within a single sequence-only framework, HERCULES provides a mechanistically interpretable approach for studying RNA-protein interactions. HERCULES is freely available at https://tools.tartaglialab.com/hercules and as an open-source Python package at https://github.com/tartaglialabIIT/hercules.git.

biochemistry↗

bioRxiv2026

Selective profiling of translationally active tRNAs and their dynamics under stress

During translation, transfer RNAs (tRNAs) deliver specific amino acids to the ribosome in a coordinated manner with the sequence encoded by the mRNA. Despite their central role in protein synthesis, their precise contribution to the modulation of translation remains poorly understood, primarily due to lack of methods to characterise tRNA abundances and their modifications from actively translating ribosomes. Here we develop tRIBO-seq, a simple and robust nanopore-based method to selectively capture ribosome-associated native tRNA populations (ribo-tRNAs) from actively translating ribosomes, providing tRNA abundance, modification and fragmentation information from a single experiment. Using tRIBO-seq, we find that ribo-tRNAs, but often not total tRNAs, are significantly altered upon stress. Notably, we find that tRNAome alterations strongly vary depending on the stress type: while viral infection and leucine deprivation leads to changes in ribo-tRNA abundances, methionine starvation causes a dramatic loss of methyl-based tRNA modifications. By contrast, we find that arsenite exposure does not alter tRNA abundances nor modification patterns, but rather causes major fragmentation events in selected subsets of ribo-tRNAs. Altogether, tRIBO-seq offers a robust and reproducible approach to map the full tRNA landscape -capturing tRNA abundance, modification and fragmentation patterns-, across both total and actively translating tRNA populations, revealing the dynamics of the tRNAome with unprecedented resolution, in a single experiment.

molecular biology↗

bioRxiv2025

Novel ciliary protein TRIM8 is a multifunctional workhorse during mitosis

TRIM8 is an E3 ubiquitin ligase that functions as both a tumour suppressor and an oncoprotein. Earlier, we reported that TRIM8 interacts with key regulators of mitotic spindle assembly, and that TRIM8 knockdown results in mitotic delay and aneuploidy. In this study, we implemented a multi-omics strategy with differential transcriptomic (single-cell RNA sequencing or scRNA-seq), translatomic (polysome profiling with RNA-seq), and proteomic (LC-MS/MS) approaches to elucidate the involvement of TRIM8 in different levels (transcription, translation, post-translation) and stages (G0/G1, S, G2/M) of mitotic cell cycle regulation and progression. With the aid of differential transcriptomic (scRNA-seq) and proteomic (LC-MS/MS) approaches, we show that depletion of TRIM8 perturbs the canonical "Cell Cycle Control of Chromosomal Replication" pathway and demonstrate that TRIM8 negatively regulates the expression of TOP2A, known to be essential for genomic integrity. We also show that TRIM8 downregulation induces substantial alterations in the translation activity of cells and results in the upregulation of polysome-bound MALAT1 lncRNA by means of significant changes in polysome profiling coupled with RNA-sequencing. Moreover, we unveil endogenous TRIM8 as a novel ciliary protein that co-localizes with CEP170, required for ciliary function, in the centrosomal region throughout all mitotic phases. Our work shows the dynamic role played by a TRIM family protein across various stages of mitosis for the first time, laying the foundation for exploring the therapeutic potential of TRIM8 in addressing cell cycle-related diseases, including cancer. HIGHLIGHTSO_LITRIM8 is involved in transcriptional and post-translational regulation of "Cell Cycle Control of Chromosomal Replication" pathway and oversees the expression of TOP2A, essential for mitotic chromosome structure maintenance. C_LIO_LIThe silencing of TRIM8 induces changes in cellular translation activity and alters the expression pattern of key translational proteins. Additionally, TRIM8-silencing leads to an elevation of long non-coding RNA (lncRNA) MALAT1 in the polysome-bound fraction. C_LIO_LITRIM8 is identified as a novel ciliary protein. C_LIO_LISilencing of TRIM8 results in the upregulation of the centrosomal protein CEP170, and both proteins co-localize in the centrosomal region throughout all stages of mitosis. C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/646005v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@1b03f1borg.highwire.dtl.DTLVardef@1ff16e4org.highwire.dtl.DTLVardef@37f07borg.highwire.dtl.DTLVardef@8d28af_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

Decoding RNA-RNA Interactions: The Role of Low-Complexity Repeats and a Deep Learning Framework for Sequence-Based Prediction

RNA-RNA interactions (RRIs) are fundamental to gene regulation and RNA processing, yet their molecular determinants remain unclear. In this work, we analyze several large-scale RRI datasets and identify low-complexity repeats (LCRs), including simple tandem repeats, as key drivers of RRIs. Our findings reveal that LCRs enable thermodynamically stable interactions with multiple partners, positioning them as key hubs in RNA-RNA interaction networks. These RRIs appear to be important for several aspects of RNA metabolism. Sequencing-based analysis of the lncRNA Lhx1os interactors validates the importance of LCRs in shaping contacts potentially involved in neuronal development. Recognizing the pivotal role of sequence determinants, we develop RIME, a deep learning model that predicts RRIs by leveraging embeddings from a nucleic acid language model. RIME outperforms traditional thermodynamics-based tools, successfully captures the role of LCRs and prioritizes high-confidence interactions, including those established by lncRNAs. RIME is freely available at https://tools.tartaglialab.com/rna_rna.

biochemistry↗

bioRxiv2024

Low fidelity DNA polymerase IV accelerates genome evolution in Pseudomonas aeruginosa

Specialized DNA polymerases are crucial for bypassing lesions and facilitating various cellular processes. Despite extensive research, the mutagenic effects of these error-prone enzymes on genomes are still not fully understood. In this study, we examined the genomic instability caused by Pol IV in the bacterial pathogen Pseudomonas aeruginosa. Pol IV mutagenesis primarily involved the misincorporation of oxidized guanine nucleotides opposite template adenine. This activity led to a distinctive mutational signature, characterized by A to C transversions occurring preferentially at AT sites flanked by a 5G and/or 3C. Furthermore, Pol IV preferentially targeted specific chromosomal locations near the replication termination region and rRNA-encoding operons. Several genes associated with virulence, motility, antibiotic resistance and chemotaxis are located in these difficult-to-replicate regions, making them hotspots for Pol IV-mutagenesis. Notably, half of the mutation events catalyzed by Pol IV effectively impaired gene function. This can be attributed to the strong bias of Pol IV for mutating specific codons with its preferred sequence contexts, leading to substitutions primarily to the unreactive Ala and Gly residues. Remarkably, mutation signatures identified for Pol IV were also present in genomes from clinical isolates of P. aeruginosa, providing compelling evidence for its role in genetic diversification during pathogen adaptation.

genomics↗

bioRxiv2024

Alterations in large-scale resting-state network nodes following transcranial focused ultrasound of deep brain structures

BackgroundLow-intensity transcranial focused ultrasound (tFUS) is a brain stimulation approach that holds immense promise for the treatment of brain-based disorders. Several studies in humans have shown that tFUS can successfully modulate perfusion in focal sonication targets including the amygdala; however, limited research has explored how tFUS impacts the function of large-scale neural networks. ObjectiveThe aim of the current study was to address this gap and examine changes in resting-state connectivity between large-scale network nodes using a randomized, double-blind, within-subject crossover study design. MethodsHealthy adults (n=18) completed two tFUS sessions, 14 days apart. Each session included tFUS of either the right amygdala or the left entorhinal cortex (ErC). The inclusion of two active targets allowed for within-subjects comparisons as a function of the locus of sonication. Resting-state functional magnetic resonance imaging was collected before and after each tFUS session. ResultstFUS altered resting-state functional connectivity (rsFC) within and between rs-network nodes. Specifically, pre-to-post sonication of the right amygdala modulated connectivity within nodes of the salience network (SAN) and between nodes of the SAN and the default-mode network (DMN) and fronto-parietal network (FRP). A decrease in SAN to FPN connectivity was specific to the amygdala target. Pre-to-post sonication of the left ErC was found to modulate connectivity between the dorsal attention network (DAN) and FPN and DMN. An increase in DAN to DMN connectivity was specific to the ErC target. ConclusionThese preliminary findings may suggest that tFUS induces neuroplastic changes beyond the immediate sonication target.

neuroscience↗

bioRxiv2024

Accurate Predictions of Phase Separating Proteins at Single Amino Acid Resolution

Liquid-liquid phase separation (LLPS) is a molecular mechanism that leads to the formation of membraneless organelles inside the cell. Despite recent advances in the experimental probing and computational prediction of proteins involved in this process, the identification of the protein regions driving LLPS and the prediction of the effect of mutations on LLPS are lagging behind. Here, we introduce catGRANULE 2.0 ROBOT (R - Ribonucleoprotein, O - Organization, in B - Biocondensates, O - Organelle, T - Types), an advanced algorithm for predicting protein LLPS at single amino acid resolution. Integrating physico-chemical properties of the proteins and structural features derived from AlphaFold models, catGRANULE 2.0 ROBOT significantly surpasses traditional sequence-based and state-of-the-art structure-based methods in performance, achieving an Area Under the Receiver Operating Characteristic Curve (AUROC) of 0.76 or higher. We present a comprehensive evaluation of the algorithm across multiple organisms and cellular components, demonstrating its effectiveness in predicting LLPS propensities at the single amino acid level and the impacts of mutations on LLPS. Our results are robustly supported by experimental validations, including immunofluorescence microscopy images from the Human Protein Atlas. catGRANULE 2.0 ROBOTs potential in protein design and mutation control can improve our understanding of proteins propensity to form subcellular compartments and help develop strategies to influence biological processes through LLPS. catGRANULE 2.0 ROBOT is freely available at https://tools.tartaglialab. com/catgranule2.

biochemistry↗

bioRxiv2023

Benchmarking tRNA-Seq quantification approaches by realistic tRNA-Seq data simulation identifies two novel approaches with higher accuracy

Quantification of transfer RNA (tRNA) using illumina sequencing based tRNA-Seq is complicated due to their degree of redundancy and extensive modifications. As such, no tRNA-Seq method has become well established, while various approaches have been proposed to quantify tRNAs from sequencing reads. Here, we use realistic tRNA-Seq simulations to benchmark tRNA-Seq quantification approaches, including two novel approaches. We demonstrate that these novel approaches are consistently the most accurate, using data simulated to mimic five different tRNA-Seq methods. This simulation-based benchmarking also identifies specific shortfalls for each quantification approach and suggests that up to 13% of the variance observed between cell lines in real tRNA-Seq data could be due to systematic differences in quantification accuracy.

bioinformatics↗

bioRxiv2023

Resolving the pharmacological redox-sensitivity of SARS-CoV-2 PLpro in drug repurposing screening enabled identification of the competitive GRL-0617 binding site inhibitor CPI-169

The SARS CoV-2 Papain-Like protease has multiple roles in the viral replication cycle, related to both its polypeptide cleavage function and its capacity to antagonize host immune response. Targeting PLpro function is recognized as a promising mechanism to modulate viral replication whilst supporting host immune responses. However, development of PLpro specific inhibitors remains challenging. Upcoming studies revealed the limitation of reported inhibitors by profiling them through a pipeline of enzymatic, binding and cellular activity assays showing unspecific activity. GRL-0617 remained the only validated molecule with demonstrated anti-viral activity in cells. In this study we refer to the pitfalls of redox-sensitivity of PLpro. Using a screening-based approach to identify inhibitors of PLpro proteolytic activity, we made extensive efforts to validate the active compounds over a range of conditions and readouts, emphasising the need for comprehensive orthogonal data when profiling putative PLpro inhibitors. The remaining active compound CPI-169, showed to compete with GRL-0617 in NMR-based experiments, suggesting to share a similar binding mode, opening novel design opportunities for further developments as antiviral agents. Author summaryThe increasing knowledge about SARS-CoV-2 allowed the development of multiple strategies to contain the spread of COVID-19 infection. Nevertheless, effective antiviral pharmacological treatments are still rare and viral evolution allowed a fast adaptation and escape from available containment methods. The papain like protease (PLpro) has now become the next most promising SARS-CoV-2 therapeutic due to its multiple functions in virus replication cycle and antagonization of host immune response. However, due to inherent flexibility and sensitivity of this enzyme specific inhibitors are rare. Here we report on a screening strategy using repurposing of known drugs that takes into account PLpro characteristics to identify new inhibitors, showing the success of the approach by identifying CPI-169 that competitive targets the well described GRL-0617 inhibitor binding pocket of PLpro and helping to design further antiviral agents.

molecular biology↗

bioRxiv2023

Interrogation of RNA-protein interaction dynamics in bacterial growth

Characterising RNA-protein interaction dynamics is fundamental to understand how bacteria respond to their environment. In this study, we have analysed the dynamics of 91% of the Escherichia coli expressed proteome and the RNA-interaction properties of 271 RNA-binding proteins (RBPs) at different growth phases. We find that 68% of RBPs differentially bind RNA across growth phases and characterise 17 previously unannotated proteins as bacterial RBPs including YfiF, a ncRNA-binding protein. While these new RBPs are mostly present in Proteobacteria, two of them have human orthologs in the form of mitochondrial proteins associated with rare metabolic disorders. Moreover, we reveal novel RBP functions for proteins such as the chaperone HtpG, a new stationary phase tRNA-binding protein. Altogether, we provide the first dynamic RBPome of a bacterium, showcasing how this approach can reveal the function of uncharacterised proteins, and identify critical RNA-protein interactions for cell growth which could inform new antimicrobial therapies.

microbiology↗

bioRxiv2023

Large-scale genomic rearrangements boost SCRaMbLE in Saccharomyces cerevisiae

Genomic rearrangements contribute to gene copy number alterations, disruption of protein-coding sequences and/or perturbation of cis-regulatory networks. SCRaMbLE, a Cre/loxP-based system implanted in synthetic yeast chromosomes, can effectively introduce genomic rearrangements, and is thus a potential tool to study genomic rearrangements. However, the potential of SCRaMbLE to study genomic rearrangements is currently hindered, because a strain containing all 16 synthetic chromosomes is not yet available. Here, we constructed a yeast strain, SparLox83, containing 83 loxPsym sites distributed across all 16 chromosomes, with at least two sites per chromosome. Inducing Cre recombinase expression in SparLox83 produced versatile genome-wide genomic rearrangements, including inter-chromosomal events. Moreover, SCRaMbLE of the hetero-diploid strains derived from crossing SparLox83 with strains possessing synthetic chromosome III (synIII) from the Sc2.0 project led to increased diversity of genomic rearrangements and relatively faster evolution of traits compared to a strain with only synIII. Analysis of these evolved strains demonstrates that genomic rearrangements can perturb the transcriptome and 3D genome structure and can consequently impact phenotypes. In summary, a genome with sparsely distributed loxPsym sites can serve as a powerful tool to study the consequence of genomic rearrangements and help accelerate strain engineering in Saccharomyces cerevisiae.

synthetic biology↗