Biology subjects

Giuliani, A.

Publications and source records attributed to Giuliani, A..

13 recordsLinked to original sources

bioRxiv2025

Mutant FUS perturbs m6A regulation by repressing ALKBH5, while restoring m6A levels alleviates ALS pathology in vivo

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder marked by the progressive loss of motor neurons, where aberrant stress granule (SG) dynamics has emerged as a hallmark, especially in cases involving mutations in RNA-binding proteins such as FUS and TDP-43. We previously showed that m6A RNA modification is elevated in FUS-associated ALS and that restoring its physiological levels through METTL3 inhibition is sufficient to rescue the aberrant stress granule phenotypes in different human cellular models. Importantly, here we show that reducing m6A levels ameliorates locomotor defects and neuromuscular junction (NMJ) abnormalities in Drosophila melanogaster models of ALS expressing the human FUSP525L mutant. Moreover, we demonstrate that the increase in m6A is attributable to reduced expression of the m6A eraser ALKBH5, resulting from impaired translation caused by direct FUS binding to its transcript. We further show that ALKBH5 downregulation contributes to RNA and protein alterations observed in FUS-associated ALS, driving the abnormal SG dynamics characteristic of the disease. This study supports the hypothesis that restoring correct m6A levels could serve as a potential therapeutic strategy for ALS.

molecular biology↗

bioRxiv2025

A Short 63-Nucleotide Element Promotes Efficient circRNA Translation

Circular RNAs (circRNAs) are a class of RNA with multiple functions, including the ability to be translated. Several intrinsic features of circRNAs, such as high stability, confer them advantages over linear RNAs; therefore, circRNA-based drugs have recently received increasing attention. However, the inefficiency of their cap-independent translation and the difficulties in the large-scale production of long circRNAs negatively impact on their use in therapy. Some efforts have been done to solve these issues related to circRNA adoption, but, to date, circRNA translation still relies on long IRESs (600-800) and chemical group addition. In this study, identified a 63-nt element able to drive circRNA translation comparably to the most commonly used IRESs. This element includes a a 13-nt sequence previously reported to enhance linear RNA translation and a segment of the UTR of the endogenously translated circRNA circZNF609. Notably, this element combines a comparable IRES-like efficiency to a considerably shorter length, expanding the landscape of ORFs potentially suitable for being translated from circRNAs and enhancing their potential as therapeutic agents in therapy.

molecular biology↗

bioRxiv2025

Understanding the Role of Toggle Genes in Chronic Lymphocytic Leukemia Proliferation

Cancer cell populations, such as chronic lymphocytic leukemia (CLL), are characterized by aberrant proliferation and plasticity: cells may switch between states so increasing population heterogeneity. Previous works have shown that gene expression noise can impact cell-state transition. To gain better insights into transcriptome-wide expression dynamics and the effect of noise on state transition, here we investigate RNA-Seq data of proliferative (PC) and non-proliferative (NPC) CLL cells. Various data analytics were applied to the whole transcriptome, switch-like toggle (ON/OFF) genes, temporal differentially expression (DE) genes, and randomly selected genes. Collectively, we identified 2713 temporal DE genes (DESeq2 with 4-fold, p < 0.05) and 1704 toggle genes shaping the differentiation process over a period of 96h, with 604 overlapping genes between them. Despite their lower numbers compared to DE, toggle genes contributed significantly to gene expression noise in both cell types. Toggle gene analyses revealed the enrichment of genes involved in processes such as G-alpha signaling and muscle contraction as proliferation related RHO-GTPase, interleukin and chemokine signaling, and lymphoid cell communication. Thus, toggle genes, although being random (at single cell level) ON/OFF genes, can show (at population level) a symmetry breaking favoring one of the two ON/OFF states so contributing to gene expression functional variability. These results suggest that toggle genes play an important role in shaping cell population plasticity.

genomics↗

bioRxiv2024

Genomic-Thermodynamic Phase Synchronization: Maxwell's Demon-Like Regulation of Cell Fate Transition

Dynamic criticality - the balance between order and chaos - is fundamental to genome regulation and cellular transitions. In this study, we investigate the distinct behaviors of gene expression dynamics in MCF-7 breast cancer cells under two stimuli: Heregulin (HRG), which promotes cell-fate transitions, and Epidermal Growth Factor (EGF), which binds to the same receptor but fails to induce cell-fate changes. We model the system as an open, non-equilibrium thermodynamic system and introduce a convergence-based framework for robust estimation of information-thermodynamic metrics. Our analysis reveals that the Shannon entropy of the critical point (CP) dynamically synchronizes with the entropy of the rest of the whole expression system (WES), reflecting coordinated transitions between ordered and disordered phases. This phase synchronization is driven by net mutual information scaling with CP entropy dynamics, demonstrating how the CP governs genome-wide coherence. Furthermore, higher-order mutual information emerges as a defining feature of the nonlinear gene expression network, capturing collective effects beyond simple pairwise interactions. By achieving thermodynamic phase synchronization, the CP orchestrates the entire expression system. Under HRG stimulation, the CP becomes active, functioning as a Maxwells demon with dynamic, rewritable chromatin memory to guide a critical transition in cell fate. In contrast, under EGF stimulation, the CP remains passive during non-critical expression dynamics. These findings establish a biophysical framework for cell-fate determination, paving the way for innovative approaches in cancer research and stem cell therapy.

genomics↗

bioRxiv2023

M6A reduction relieves FUS-associated ALS granules

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease due to gradual motorneurons (MN) degeneration1. Among the processes associated to ALS pathogenesis, there is the formation of cytoplasmic inclusions produced by mutant protein aggregation, among which the RNA binding protein FUS2. In this work we show that such inclusions are significantly reduced in number and dissolve faster when the RNA m6A content is diminished as a consequence of the m6A writer METTL3 knock-down. These effects were observed both in neuronal cell lines and in iPSC-derived human motor neurons expressing mutant FUS. Importantly, stress granules formed in ALS condition showed a distinctive transcriptome with respect to control cells; interestingly, after METTL3 downregulation, it reverted to similar to control. Finally, we show that FUS inclusions are reduced also in patient-derived fibroblasts treated with STM-2457, a well characterized inhibitor of METTL3 activity, paving the way for its possible use for counteracting aggregate formation in ALS.

molecular biology↗

bioRxiv2023

ALS-associated FUS mutation reshapes the RNA and protein composition and dynamic of Stress Granules.

Stress Granules (SG) formation is a cellular protection mechanism, constituting a storage for untranslated mRNAs and RNA-binding proteins (RBPs); however, these condensates can turn into pathological aggregates, related to the onset of neurodegenerative diseases like Amyotrophic Lateral Sclerosis (ALS). This transition towards cytotoxic inclusions is triggered by ALS-causative mutations in the RBP FUS, which lead to its cytoplasmic mis-localization and accumulation in SG. Here, we describe the SG transcriptome in a neural context and describe several features for RNA recruitment in SG. We demonstrate that SG dynamics and RNA content are strongly modified by the incorporation of mutant FUS, switching to a more unstructured, AU-rich SG transcriptome. Moreover, we show that mutant FUS, together with its protein interactors and their target RNAs, are responsible for the reshaping of the mutant SG transcriptome with alterations that can be linked to neurodegeneration. Therefore, our data give a comprehensive view of the molecular differences between physiological and pathological SG in ALS conditions, showing how FUS mutations impact the RNA and protein population of these condensates.

molecular biology↗

bioRxiv2023

Dbx2, an aging-related homeobox gene, inhibits the proliferation of adult neural progenitors

The subventricular zone (SVZ) of the adult mouse brain contains quiescent neural stem cells, which can be activated (aNSCs) to generate transit amplifying progenitors (TAPs), neuroblasts (NBs) and newborn neurons. Neurogenesis declines during aging, as the aged SVZ niche causes transcriptomic changes that promote NSC quiescence and decrease proliferating neural/stem progenitor cells (NSPCs). The transcription factors mediating these changes, however, remain unclear. We previously found that the homeobox gene Dbx2 is upregulated in aged SVZ NSPCs and inhibits NSPC culture growth. Here, we report that Dbx2 is repressed by Epidermal Growth Factor Receptor signaling, which promotes NSPC proliferation and decreases in the aged SVZ. We show that Dbx2 inhibits NSPC proliferation by hindering the G2/M transition and elucidate the transcriptomic networks modulated by Dbx2, highlighting its role in the downregulation of the cell cycle molecular pathways. Accordingly, Dbx2 function is negatively correlated with the transcriptional signatures of proliferative NSPCs (aNSCs, TAPs and early NBs). These results point to Dbx2 as a molecular node relaying the anti-neurogenic input of the aged niche to the NSPC transcriptome.

developmental biology↗

bioRxiv2021

A Unified Genomic Mechanism of Cell-Fate Change

The purpose of our studies is to elucidate the nature of massive control of whole genome expression with a particular emphasis on cell-fate change. Whole genome expression is coordinated by the emergence of a critical point (CP: a peculiar set of bi-phasic genes) through the genome-engine. In response to stimuli, the genome expression self-organizes three critical states, each exhibiting distinct collective behaviors with its center of mass acting as a local attractor, coexisting with whole genome attractor (GA). Genome-engine mechanism accounts for local attractors interaction in phase space. The CP acts as the organizing center of cell-fate change, and its activation makes local perturbation spread over the genome affecting GA. The activation of CP is in turn elicited by hot-spots, genes with elevated temporal variance, normally in charge to keep genome expression at pace with microenvironment fluctuations. When hot-spots oscillation exceeds a given threshold, the CP synchronizes with the GA driving genome expression state transition. The expression synchronization wave invading the entire genome depends on the power law fusion-bursting dynamics of silencing pericentromere-associated heterochromatin domains and the consequent folding-unfolding status of transcribing euchromatin domains. The proposed mechanism is a unified step toward a time-evolutional transition theory of biological regulation.

cell biology↗

bioRxiv2019

Self-Organization of Whole Gene Expression through Coordinated Chromatin Structural Transition: Validation of Self-Organized Critical Control of Genome Expression

Through our studies on whole genome regulation, we have demonstrated the existence of self-organized critical control (SOC) of whole gene expression - genomic self-organization mechanism through the emergence of a critical point (CP) at both the cell population and single cell level. In this paper, based on HRG and EGF-stimulated MCF-7 breast-cancer cell line, we shed light on the origin of critical transitions stemming from coordinated chromatin remodeling. In so doing, we validated the core of the SOC control mechanism through the application of a non-linear signal analysis technique (Recurrence Quantification Analysis: RQA), and of Principal Component Analysis (PCA). The main findings were: O_LITranscriptional co-regulation follows a strong and invariant exponential decay as between gene spacing along the chromosome is increased. This shows that the co-regulation occurs on a mainly positional basis reflecting local chromatin organization. C_LIO_LIThere are two main fluctuation modes on the top of the cell-kind specific gene expression values spanning the entire genome expression. These modes establish an autonomous genomic critical control system (genome-engine) through the activation of the CP for cell-fate guiding critical transitions revealed by SOC analysis. C_LI The elucidation of the link between spatial position on chromosome and co-regulation together with the identification of specific locations on the genome devoted to the generalization of perturbation stimuli, give a molecular basis to the self-organization dynamics of genome expression and cell-fate decision.

genomics↗

bioRxiv2019

Underlying Genomic Mechanism for Cell-Fate Change from Embryo to Cancer Development

The elucidation of the how and when of a cell-fate change asks for a physically reasonable mechanism allowing to achieve a coordinated switching of thousands of genes within a small and highly packed cell nucleus. We previously demonstrated that whole genome expression is dynamically self-organized through the emergence of a critical point. Furthermore, it has been confirmed that this happens at both the cell-population and single-cell level through the physical principle of self-organized criticality. In this paper, we further examine the genomic mechanism which determines cell-fate changes from embryo to cancer development. The state of the critical point, acting as the organizing center of cell-fate, determines whether the genome resides in a super- or sub-critical state. In the super-critical state, a specific stochastic perturbation can spread over the entire system through the genome engine - an autonomous critical-control genomic system, whereas in the sub-critical state, the perturbation remains at a local level. We provide a consistent framework to develop a biological regulation transition theory demonstrating the cell-fate change.

cell biology↗

bioRxiv2018

Self-Organized Critical Control of Genome Expression: Novel Scenario on Cell-Fate Decision

In our current studies on whole genome expression in several biological processes, we have demonstrated the actual existence of self-organized critical control (SOC) of gene expression at both population and single cell level. SOC allows for cell-fate change by critical transition encompassing the entire genome expression that, in turn, is partitioned into distinct response domains (critical states).\n\nIn this paper, we go more in depth into the elucidation of SOC control of genome expression focusing on the determination of critical point (CP) and associated distinct critical states in single-cell genome expression. This leads us to the proposal of a potential universal model with genome-engine mechanism for cell-fate change. Our findings suggest that the CP is fixed point in terms of temporal expression variance, where the CP (set of critical genes) becomes active (ON) for cell-fate change ( super-critical in genome-state) or else inactive (OFF) state ( sub-critical in genome-state); this may lead to a novel scenario of the cell-fate control through activating or inactivating CP.

genomics↗

bioRxiv2017

Single-Cell Reprogramming Of Mouse Embryo Development Through A Critical Transition State

Our work dealing with the temporal development of the genome-expression profile in single-cell mouse early embryo indicated that reprogramming occurs via a critical transition state, where the critical-regulation pattern of the zygote state disappears. In this report, we unveil the detailed mechanism of how the dynamic interaction of thermodynamic states (critical states) enables the genome system to pass through the critical transition state to achieve genome reprogramming.\n\nSelf-organized criticality (SOC) control of overall expression provides a snapshot of self-organization and explains the coexistence of critical states at a certain experimental time point. The time-development of self-organization is dynamically modulated by exchanges in expression flux between critical states through the cell nucleus milieu, where sequential global perturbations involving activation-inhibition of multiple critical states occur from the early state to the late 2-cell state. Two cyclic fluxes act as feedback flow and generate critical-state coherent oscillatory dynamics. Dynamic perturbation of these cyclic flows due to vivid activation of the ensemble of low-variance expression (sub-critical state) genes allows the genome system to overcome a transition state during reprogramming.\n\nOur findings imply that a universal mechanism of long-term global RNA oscillation underlies autonomous SOC control, and the critical gene ensemble at a critical point (CP) drives genome reprogramming. Unveiling the corresponding molecular players will be essential to understand single-cell reprogramming.

developmental biology↗

bioRxiv2017

Single-Cell Statistical Thermodynamics In Early Embryo Development

A statistical thermodynamics approach to the temporal development of biological regulation provides a phenomenological description of the dynamical behavior of genome expression in terms of autonomous self-organization with a critical transition (Self-Organized Criticality: SOC). In early mouse embryo development, the dynamical change in the self-organization of overall expression determines how and when reprogramming of the genome-expression state occurs. Reprogramming occurs via a transition state (climbing over an epigenetic landscape), where the critical-regulation pattern of the zygote state disappears. A critical transition is well captured in terms of the bimodality of expression ensembles, which reflects distinct thermodynamic states (critical states). These critical states exhibit a genome avalanche pattern: competition between order (scaling) and disorder (divergence) around a critical point. The genome avalanche in mouse embryo development, which is committed to erase a previous ordered state, reveals that the direction of early embryo single-cell development traverses the same steps as in differentiation, but in the opposite order of self-organization.

genomics↗