Biology subjects

Basso, M.

Publications and source records attributed to Basso, M..

8 recordsLinked to original sources

bioRxiv2025

Empowering multiplexed ultra-throughout ribosome profiling with RiboWich

Ribosome profiling (RiboSeq) improved the understanding of mRNA translation, enabling the precise mapping of ribosome positioning along transcripts at single-nucleotide resolution. Although various library preparation protocols overcome the need for high input material, their technical complexity and limited efficiency hinder robust profiling, preventing them from keeping pace with other sequencing techniques and applications. To move towards high-throughput and single-cell RiboSeq technologies, we developed RiboWich (Ribosome sandWich). By directly ligating adaptors to ribosome-embedded RNA fragments, RiboWich eliminates the need for ribosome purification and size-selecting ribosome footprints and tackles two major bottlenecks, expanding RiboSeq for more advanced technologies. RiboWich offers robustness in profiling and excels in detecting upstream translons in immortalized and primary cells alike. By exploiting a dual-step multiplexing strategy, RiboWich enables the simultaneous profiling of at least 96 samples while retaining sensitivity and capturing condition-specific differences in translation. By enabling scalable, low-input translatome profiling, this advancement empowers proteogenomic approaches and AI/ML-driven data analysis to uncover regulatory dynamics, neoantigens, functional small translons, and drug-responsive signatures across diverse biological contexts and users. Altogether, RiboWich represents a straightforward, versatile, and scalable ribosome profiling conceptual platform that combines accessibility, sensitivity, and throughput potential, laying the foundation for advanced single-cell RiboSeq applications.

molecular biology↗

bioRxiv2025

Natural variants of von Willebrand factor R1205 causing von Willebrand disease with accelerated von Willebrand factor clearance: in silico docking models and energetics of the interaction with both LRP1 and GpIb A1 domain

Type 1 von Willebrand disease (VWD) is often caused by variants in von Willebrand factor (VWF), including p.R1205H ("Vicenza mutation"), which accelerate VWF clearance via macrophage receptor LRP1 and impair platelet adhesion. However, the structural mechanisms underlying these phenotypes remain unclear. Here, we use integrative computational modeling (I-TASSER, HADDOCK2.4, and PRODIGY) to predict how p.R1205H/C/L/S variants alter VWF interaction with LRP1 and GPIb. Our models reveal that R1205 acts as a structural hinge: its variants disrupt polar networks in VWFs D3 domain, exposing neo-epitopes that enhance LRP1 binding ({Delta}{Delta}G up to -5.3 kcal/mol) while destabilizing the A1 domains 1-{beta}2 loop, reducing GPIb affinity (33-fold for R1205L/C). These findings explain clinical observations, p.R1205H rapid clearance yet retained platelet adhesion, and establish R1205 as a dual-functional switch regulating VWF circulatory lifetime and hemostatic activity. This analytical procedute provides a template for predicting pathogenicity of VWF variants and designing targeted therapies for VWD. Author SummaryIn our study, we explored how specific genetic changes in von Willebrand factor (VWF), a protein crucial for haemostsis, lead to a bleeding disorder called von Willebrand disease (VWD). We focused on mutations at position R1205 in VWF, known to cause rapid clearance of the protein from the bloodstream, resulting in low levels and impaired clotting. Using advanced computer modeling, we investigated how these mutations alter VWFs interactions with two key partners: LRP1, a receptor that removes VWF from circulation, and GpIb, a platelet receptor essential for clotting. Our models revealed that R1205 acts like a structural hinge. Mutations disrupt its stabilizing role, exposing regions that bind more tightly to LRP1, thus explaining the accelerated clearance seen in patients. Simultaneously, these changes can also weaken VWFs ability to interact with platelets, further impairing platelet haemostatic function. This work provides a blueprint for understanding how some VWF variants cause disease and could guide the design of targeted therapies to correct or compensate for these defects, improving treatment for VWD patients.

biochemistry↗

bioRxiv2025

Targeting Neuromuscular Junction Regeneration is a Therapeutic Strategy in ALS

Instability and denervation of the neuromuscular junction (NMJ) are early events in Amyotrophic Lateral Sclerosis (ALS), likely reflecting a progressive decline in the regenerative capacity of motor neurons (MNs) and their environment. To investigate this, we evaluated NMJ regeneration throughout disease progression in SOD1G93A mice following reversible axon terminal degeneration induced by -Latrotoxin. In parallel, we monitored the expression of CXCR4, a GPCR upregulated during axonal regeneration, and tested whether its pharmacological activation could mitigate ALS- related functional decline. We found that NMJ regenerative capacity is largely preserved during pre- and early symptomatic stages, and remains active in subsets of NMJs even at later stages. CXCR4 is expressed at axon terminals from early disease stages, declining only at end stage. Its expression is conserved across ALS models, including SOD1G93A pigs, hiPSC-derived MN with ALS mutations, and biopsies from sporadic ALS patients. CXCR4 stimulation improved motor function, NMJ innervation, MN survival, and respiratory performance in ALS mice, and axon outgrowth in iPSC-derived MN. These findings identify the NMJ and CXCR4 as viable therapeutic targets in ALS.

neuroscience↗

bioRxiv2025

Translation-specific disruption of Col1a1 expression in multiple models of Spinal Muscular Atrophy can be rescued by Risdiplam.

Spinal muscular atrophy (SMA) is a monogenic neurodegenerative disorder caused by decreased levels of Survival of Motor Neuron (SMN) protein. If left untreated, SMA patients have a poor prognosis, marked by the degeneration of motor neurons, progressive muscle weakness and atrophy. The approval of SMN-restoring therapies that improve symptoms and lifespan in patients with SMA has created emerging, non-neuronal phenotypes and an urgent need for deepening our understanding of disease pathogenesis. Leveraging the knowledge that SMN loss drives alterations in translation, we used multiple tissues from a mouse model of SMA to uncover early translational alterations in key mRNAs and proteins, which act as contributors to pathogenesis and hallmarks of the disease. Among hundreds of differentially translated mRNAs, Col1a1 emerged as a translation-specific manifestation of early defects in the mouse model. These findings were confirmed in fibroblasts derived from patients with varying levels of disease severity. Notably, treatment with SMN-restoring therapies rescued COL1A1 protein levels, particularly in fibroblasts from patients with the most severe forms of the disease. Overall, our study identifies COL1A1 as an indicator of disease severity in SMA, which captures early molecular alterations and respond to SMN-modifying therapies.

molecular biology↗

bioRxiv2025

Model-independent reorganization of translation in TDP-43 Amyotrophic Lateral Sclerosis

The RNA-binding protein TDP-43 is a major contributor and a pathological hallmark of Amyotrophic Lateral Sclerosis (ALS), yet how TDP-43 dysregulation mechanistically alters protein synthesis across neuronal compartments and disease models remains unclear. Here, we dissected TDP-43-driven translational alterations in both in vitro and in vivo TDP-43 models of ALS. Through ribosome and polysome profiling, computational, and biochemical analyses, we observed robust TDP-43-associated translational remodelling at cellular and subcellular resolution. Our findings reveal a conserved mechanism across models, characterized by enhanced ribosome recruitment on polysomes, elongation impairment, axonal downregulation and instability of TDP-43 target mRNAs and redistribution toward non-target transcripts. Notably, TDP-43 dysregulation alters ribosome dynamics and selectively impairs translation of TDP-43 target mRNAs, whilst favouring the translation of other transcripts. This process reflects a compensatory but maladaptive response to TDP-43-induced mRNA destabilization. Together, these data demonstrate that alterations in TDP-43 disrupts neuronal proteostasis through ribosome reorganization and loss of mRNA homeostasis, providing a unifying mechanistic framework for translational imbalance in ALS and revealing early molecular events that may underlie motor neuron vulnerability.

molecular biology↗

bioRxiv2024

Small molecule inhibitors of hnRNPA2B1-RNA interactions reveal a predictable sorting of RNA subsets into extracellular vesicles

Extracellular vesicles (EVs) are cell-secreted membranous particles contributing to intercellular communication. Coding and non-coding RNAs are widely detected EV cargo, and RNA-binding proteins (RBPs), such as hnRNPA2B1, have been circumstantially implicated in sorting vesicular RNAs. However, the contribution of competitive RBP-RNA interactions responsible for RNA-sorting outcomes still needs to be deciphered, especially for EV-RNA interference and predictability. We conducted a reverse proteomic analysis that prioritized heterogeneous nuclear ribonucleoproteins recognizing purine-rich RNA sequences representing a subset of previously identified EXO motifs. A screening campaign using a full-length human hnRNPA2B1 protein and artificial purine-rich RNA brought to small molecule inhibitors orthogonally validated through biochemical and cell-based approaches. Selected drugs effectively interfered with a post-transcriptional layer impacting secreted EV- RNAs, reducing the vesicular pro-inflammatory miR-221 while counteracting the hnRNPA2B1- or TDP43Q331K-dependent paracrine activation of NF-{kappa}B in EV-recipient cells. This study demonstrates the possibility of predicting the EV-RNA quality for developing innovative strategies targeting discrete paracrine functions. SummaryExtracellular vesicles (EVs) are cell-released, heterogeneous lipid particles conceived as vehicles for intercellular communication. RNA is a widely detected cargo, and the comprehension of EV-sorting mechanisms represents a step forward in predicting EV quality and associated paracrine effects. While it is known that specific RNA-binding proteins (RBPs) play a role in EV-RNA sorting, the quantitative contribution of competing RBP-RNA interactions and the predictability of RNA-sorting outcomes are poorly understood. Here, we show that a core of hnRNPs compete for the binding to the heterogeneous EV-RNA in vitro. Given prioritized interactions with purine-rich RNA motifs, we set up a pharmacological screen platform to find inhibitors of protein-RNA interactions. Our results suggest that selected small molecules can interfere with EV-RNA quality, altering the distribution of specific miRNA cargoes and associating with a discriminant NF-kB activation in EV-recipient cells. This work highlights the role of RBP-RNA interactions in influencing the EV-RNA quality and paracrine functions.

molecular biology↗

bioRxiv2023

Actin-based deformations of the nucleus control multiciliated ependymal cell differentiation.

Ependymal cells (ECs) are multi-ciliated cells in the brain that contribute to cerebrospinal fluid flow. ECs are specified at embryonic stages but differentiate later in development. Differentiation depends on genes such as GEMC1 and MCIDAS with E2F4/5, but also on cell cycle related factors. In the mouse brain, we observe that nuclear deformation accompanies EC differentiation. Tampering with these deformations either by decreasing F-actin levels or by severing the link between the nucleus and the actin cytoskeleton blocks differentiation. Conversely, increasing F-actin by knocking out the Arp2/3 complex inhibitor Arpin or artificially deforming the nucleus activates differentiation. These data are consistent with actin polymerization triggering nuclear deformation and jump-starting the signaling that produces ECs. A player in this process is the retinoblastoma1 (RB1) protein whose phosphorylation prompts MCIDAS activation. Overall, this study reveals an important role for actin-based mechanical inputs to the nucleus as controlling factors in cell differentiation.

developmental biology↗

bioRxiv2023

Antero-posterior gradients of cell plasticity and proliferation modulate posterior regeneration in the annelid Platynereis

Regenerative abilities are extremely variable among animals and may be substantial in some phyla, such as the annelids. So far, the cellular mechanisms underlying regeneration in annelids remain elusive. To precisely determine the origin(s), plasticity and fate of the cells participating in the blastema formation during posterior regeneration in the annelid Platynereis dumerilii, we developed specific tools to track proliferative cells as well as gut epithelial cells. We showed that two populations of progenitors are at play during regeneration and that, among them, gut progenitors from differentiated tissues are lineage-restricted. Strikingly, gut progenitors from less differentiated and more proliferative tissues are much more plastic and can produce ectodermal and mesodermal derivatives, in addition to gut cells. However, their plasticity is de facto limited as exemplified by their inability to regenerate populations of stem cells responsible for the constant growth of the worms. We evidenced that those stem cells are from local origin (i.e. from the segment abutting the amputation plan) as most of the blastema cells. Our results are in favour of a hybrid and flexible cellular model for posterior regeneration in Platynereis relying on a gradient of cell plasticity along the antero-posterior axis of the animal.

developmental biology↗