Biology subjects

Bianchi, M. E.

Publications and source records attributed to Bianchi, M. E..

7 recordsLinked to original sources

bioRxiv2026

NF-κB transcriptionally enhances p53 accumulation dynamics hampering DNA repair

Cells integrate multiple, often concurrent signals through intertwined genetic circuits whose dynamics shape transcriptional programs and cell fate decisions. Among these, the tumor suppressor p53 and the inflammatory transcription factor NF-{kappa}B are central regulators of stress responses in normal and cancer cells, yet their dynamic crosstalk under co-activation remains poorly characterized. Here, we combine genetic approaches, live cell imaging, transcriptomic analysis and mathematical modeling to dissect their dynamic interplay. We find that co-activation of NF-{kappa}B by the inflammatory cytokines TNF- and IL-1{beta} significantly enhance p53 nuclear accumulation upon genotoxic stress or Nutlin3a, and this effect is absent in NF-{kappa}B-deficient cells. Mechanistically, we show that cytokines induce an NF-{kappa}B-mediated increase of TP53 transcription, and mathematical modeling indicates that it is sufficient to account for the observed increased p53 accumulation. Functionally, NF-{kappa}B co-activation rewires p53-dependent transcriptional programs and impairs p53-mediated DNA repair following genotoxic stress, due to a shift of p53 dynamics from oscillatory to more sustained accumulation; p53 oscillatory dynamics and DNA repair remain largely unaltered in absence of NF-{kappa}B. Our results uncover an amplification of p53 response in presence of inflammatory cues that is transcriptionally mediated by NF-{kappa}B and that results, counterintuitively, in functional antagonism. SIGNIFICANCE STATEMENTp53 dynamics have been shown to correlate with the cellular responses of cancer cells to genotoxic insults such as those delivered by chemo- and radiotherapies. However, these dynamics have been mostly studied in settings that do not account for the pro-inflammatory cues that cancer cells might receive from the microenvironment. By addressing this gap, our study uncovers a previously unappreciated mechanism by which inflammatory signals induce an increased p53 accumulation, leading to reduced DNA repair capacity. Our results provide mechanistic understanding on the origin of p53-NF-{kappa}B antagonism and show how dynamically interconnected signalling pathways can produce counterintuitive functional outcomes. Translationally, the contribution of inflammation to the sensitivity of normal and cancer cells to DNA damage might be exploited in the management of chemo- and radiotherapies.

systems biology↗

bioRxiv2025

Neutrophil Reprogramming Underlie Vasculopathy and Lung Disease in Systemic Sclerosis

The role of neutrophils in systemic sclerosis (SSc) remains incompletely understood. To address this, blood samples from 39 SSc patients, 39 healthy controls, and 22 systemic lupus erythematosus (SLE) patients were analyzed. In SSc, neutrophils exhibited substantial activation, evidenced by granule mobilization, elevated plasma levels of Neutrophil Extracellular Trap (NET) byproducts, and upregulated TIE2 expression. In parallel, they underwent metabolic reprogramming, characterized by increased autophagy, likely to support the heightened energy demands of activation. By contrast, neutrophils from SLE patients displayed minimal autophagy, lacked TIE2 expression, and shifted toward low-density granulocytes. Neutrophil reprogramming in SSc correlated with plasma levels of HMGB1+ EVs. Mechanistically, EVs purified from the plasma of patients with SSc adhered to neutrophils when injected in immunodeficient NSG mice, inducing autophagy, TIE2 expression, and promoting lung inflammation and fibrosis. These effects were abrogated by HMGB1 inhibitors and required the HMGB1 receptor, RAGE. Recombinant HMGB1 recapitulated EV-induced effects, while neutrophil targeting by liposome-encapsulated clodronate prevented them. In summary, neutrophils in SSc exhibit a dual phenotype of autophagy and activation driven by HMGB1+ EVs, representing a pathogenic mechanism and a potential therapeutic target in SSc. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=139 HEIGHT=200 SRC="FIGDIR/small/637866v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@dbf3dcorg.highwire.dtl.DTLVardef@67243forg.highwire.dtl.DTLVardef@17cb6c4org.highwire.dtl.DTLVardef@c3370a_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2023

The acidic intrinsically disordered region of the inflammatory mediator HMGB1 mediates fuzzy interactions with chemokine CXCL12

Chemokines engage in heterodimeric interactions to activate or dampen their cognate receptors in inflammatory conditions. The chemokine CXCL12 forms with the alarmin HMGB1 a patho-physiologically relevant heterocomplex (HMGB1[bullet]CXCL12), whose formation synergically promotes the inflammatory response elicited by the G-protein coupled receptor CXCR4. However, the molecular details of complex formation were still elusive. Through an integrative structural approach (NMR, AUC, ITC, MST, SAXS) we show that HMGB1[bullet]CXCL12 represents the first fuzzy chemokines heterocomplex reported so far. HMGB1 and CXCL12 form a dynamic equimolar assembly, rather than involving one HMGB1 and two CXCL12 molecules as previously assumed, with structured and unstructured HMGB1 regions recognizing the dimerization surface of CXCL12. We uncover an unexpected role of the acidic intrinsically disordered region (IDR) in heterocomplex formation and provide the first evidence that the acidic IDR facilitates the ternary HMGB1*CXCL12*CXCR4 interaction on the cell surface. Thus, the interaction of HMGB1 with CXCL12 diverges radically from the classical rigid heterophilic chemokine-chemokine dimerization. Simultaneous interference with the multiple interactions within HMGB1[bullet]CXCL12 complex formation might offer novel pharmacological strategies to inhibit its detrimental activity in inflammatory conditions.

biochemistry↗

bioRxiv2021

Small transcriptional differences lead to distinct NF-κB dynamics in quasi-identical cells

Transcription factor dynamics is fundamental to determine the activation of accurate transcriptional programs and yet is heterogeneous at single-cell level, even between very similar cells. We asked how such heterogeneity emerges for the nuclear factor {kappa}B (NF-{kappa}B), whose dynamics have been reported to cover a wide spectrum of behaviors, including persistent, oscillatory and weak activation. We found that clonal populations of immortalized fibroblasts derived from a single mouse embryo display robustly distinct dynamics upon tumor necrosis factor (TNF-) stimulation, which give rise to differences in the transcription of NF-{kappa}B targets. Notably, standard transcriptomic analyses indicate that the clones differ mostly in transcriptional programs related with development, but not in TNF- signaling. However, by combining transcriptomics data and simulations we show how the expression levels of genes coding for proteins of the signaling cascade determine the differences in early NF-{kappa}B activation; differences in the expression of I{kappa}B determine differences in its persistence and oscillatory behavior. The same analysis predicts inter-clonal differences in the NF-{kappa}B response to IL-1{beta}. We propose that small (less than twofold) differences at transcript level can lead to distinct transcription factor dynamics in cells within homogeneous cell populations, and all the more so among different cell types.

systems biology↗

bioRxiv2021

Heparin protects human neural progenitor cells from Zika Virus-induced cell death and preserves their differentiation into mature neural-glia cells

Zika virus (ZIKV) is an arbovirus member of the Flaviviridae family that causes severe congenital brain anomalies in infected fetuses. Human neural progenitor cells (hNPCs) are highly permissive to ZIKV infection, causing inhibition of cell proliferation concomitant with an induction of cell death. We previously demonstrated that pharmaceutical-grade heparin inhibited virus-induced cell death with minor effects on in vitro virus replication in ZIKV-infected hNPCs. Here we show that heparin prevented ZIKV-induced intracellular vacuoles, a signature characteristic of paraptosis, but also inhibited necrosis and apoptosis of hNPCs when grown as neurospheres (NS). Furthermore, heparin preserved the differentiation of both ZIKV-infected human-induced pluripotent stem cells (hiPSC) derived-NPCs and fetus-derived NPCs into neural-glial cells. Collectively, these results highlight the potential neuroprotective effect of heparin that could be re-purposed and exploited to drive the development of novel agents for preventing ZIKV damage.

microbiology↗

bioRxiv2020

Discovery of 5,5'-methylenedi-2,3-cresotic acid as a potent inhibitor of the HMGB1/CXCL12 heterocomplex using Virtual Screening and NMR validation

HMGB1 is a key molecule that both triggers and sustains inflammation following infection or injury, and is involved in a large number of pathologies, including cancer. HMGB1 participates to the recruitment of inflammatory cells forming a heterocomplex with the chemokine CXCL12 (HMGB1*CXCL12), herewith activating the G-protein coupled receptor CXCR4. Thus, identification of molecules that disrupt this heterocomplex can offer novel pharmacological opportunities to treat inflammation related diseases. To identify new HMGB1*CXCL12 inhibitors we have performed a study on the ligandability of the single HMG boxes of HMGB1 followed by a virtual screening campaign on both HMG boxes using Zbc Drugs and three different docking programs (Glide, AutoDock Vina, AutoDock 4.2.6). The best poses in terms of scoring functions, visual inspection and predicted ADME properties were further filtered according to a pharmacophore model based on known HMGB1 binders and clustered according to their structures. Eight compounds representative of the clusters were tested for HMGB1 binding by NMR. We identified 5,5-methylenedi-2,3-cresotic acid (2a) as binder of both HMGB1 and CXCL12; 2a also targets the HMGB1*CXCL12 heterocomplex. In cell migration assays 2a inhibited the chemotactic activity of HMGB1*CXCL12 with IC50 in the subnanomolar range, the best documented up to now. These results pave the way for future structure activity relationship studies to optimize the pharmacological targeting of HMGB1*CXCL12 for anti-inflammatory purposes.

biochemistry↗

bioRxiv2020

First responders shape a prompt and sharp NF-Bκ-mediated transcriptional response to TNF-α

SummaryNF-κB acts as the master regulator of the transcriptional response to inflammatory signals by translocating into the nucleus upon stimuli, but we lack a single-cell characterization of the resulting transcription dynamics. Here we show that transcription of NF-κB target genes is strongly heterogeneous in individual cells but dynamically coordinated at the population level, since the average nascent transcription is prompt (i.e. occurs almost immediately) and sharp (i.e. increases and decreases rapidly) compared to NF-κB nuclear localization. Using an NF-κB-controlled MS2 reporter we confirm that the population-level transcriptional activity emerges from a strongly heterogeneous response in single cells as compared to NF-κB translocation dynamics, including the presence of a fraction of “first responders”. Mathematical models show that a combination of NF-κB mediated gene activation and a gene activity module including a gene refractory state is enough to produce sharp and prompt transcriptional responses. Our data and models show how the expression of the target genes of a paradigmatic inducible transcription activator upon stimuli can be time-resolved at population level and yet heterogeneous across single cells.Competing Interest StatementThe authors have declared no competing interest.View Full Text

systems biology↗