Biology subjects

Pinto, C.

Publications and source records attributed to Pinto, C..

10 recordsLinked to original sources

bioRxiv2026

State-selective nanobodies to probe Guanylate-binding protein coat assembly and function

Cell-autonomous immunity against gram-negative bacterial pathogens relies on Guanylate-binding proteins (GBPs). GBPs are interferon-inducible guanosine triphosphate hydrolases that coat cytosolic bacteria and promote non-canonical inflammasome activation. GBP coat assembly is initiated by GBP1, which binds bacterial lipopolysaccharide and recruits further paralogs in a hierarchical and nucleotide-dependent manner. Dissecting the contributions of individual GBPs has remained challenging because the seven human paralogs are structurally similar and undergo nucleotide-driven conformational changes that are difficult to resolve in cellular and biochemical experiments. Here, we developed a nanobody toolbox for the state-selective probing of human GBP coat assembly. We identify eight nanobodies with distinct paralog, domain, and conformational specificities and define their binding profiles and epitopes using complementary biophysical assays and cryogenic electron microscopy. We further demonstrate the utility of selected nanobodies by demonstrating their functionality in membrane reconstitution assays and in infected cells. A dimer-disrupting nanobody blocks progression of GBP-coat assembly from soluble GBP1 assemblies to LPS-associated coats, suggesting a stage-specific structural transition during coat formation. In contrast, a dimer selective nanobody binds assembled coats without measurably perturbing their organisation and is robustly recruited to GBP1-positive bacteria in infected cells, indicating that this dimer interface remains intact and accessible in coat-associated GBP1. Together, these nanobodies establish a set of state-selective reagents to distinguish, perturb, and probe GBP assemblies in structural and cellular studies.

biophysics↗

bioRxiv2026

CircZBTB46, a promising therapeutic target in crizotinib resistant ALK-positive T lymphomas

Circular RNAs (circRNAs) are increasingly recognized as functional non-coding transcripts with oncogenic potential. Here, a comprehensive analysis of circRNA expression in primary ALK(+) anaplastic large-cell lymphoma (ALK(+) ALCL) is presented. Integrated transcriptomic profiling revealed that aberrant expression of circZBTB46 and of its linear host transcript, normally restricted to dendritic cells, is exclusive to ALK(+) lymphoma cells and driven by the oncogenic NPM1-ALK/STAT3 axis. Functional studies showed that circZBTB46, unlike its protein-coding counterpart, promotes resistance to the ALK inhibitor crizotinib. Silencing circZBTB46 restored crizotinib sensitivity in resistant ALCL cells both in vitro and in vivo. Transcriptomic analyses identified PIP5K1C as a downstream effector regulated through a competitive endogenous RNA mechanism in which circZBTB46 acts as a sponge to miR-25-3p, alleviating its repression of PIP5K1C. These findings uncover a previously unrecognized mechanism of drug resistance in ALK(+) ALCL and establish circZBTB46 as a promising therapeutic target.

cancer biology↗

bioRxiv2025

From berries to brain: Assessing the impact of (poly)phenols in the MPTP mouse model of Parkinson's disease

The growing burden of chronic neurodegenerative diseases (NDs), particularly Parkinsons disease (PD), prompts the need for effective preventive strategies and treatments. Dietary (poly)phenols have emerged for their neuroprotective potential. This study investigated a (poly)phenol-enriched diet, comprising a berry mixture, to counteract key PD hallmarks in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-intoxicated mice and elucidate the phenolic metabolic fingerprint underlying these effects. The berry-enriched diet prevented motor deficits in the MPTP mice model, preserved dopaminergic neurons in the midbrain, reduced glial activation and gene expression of inflammatory cytokines. Notably, berries also attenuated macrophage infiltration observed in the substantia nigra 7 days post-MPTP. Metabolomic analysis revealed distinct phenolic signatures in plasma and brain tissue between standard- and berry-fed mice. Overall, this pioneering study provides compelling evidence that a (poly)phenol-enriched diet may play a protective role in neurodegenerative disorders, highlighting its potential for future strategies to prevent or slow PD progression.

neuroscience↗

bioRxiv2025

Lactucopicrin: A sesquiterpene lactone with anti-inflammatory activity modulates the crosstalk between NF-kB and AHR pathways

Chronic inflammatory diseases, including inflammatory bowel diseases and cardiovascular diseases, share inflammation as a central pathological feature. The transcription factors NF-kB and AHR are key regulators of inflammatory responses, yet their interplay remains underexplored in the context of natural anti-inflammatory compounds. This study investigates the anti-inflammatory properties of sesquiterpene lactones derived from Cichorium intybus L. (chicory), focusing on their modulation of NF-kB and AHR signaling pathways. Using a TNF-induced inflammation model in macrophages, endothelial, and intestinal epithelial cells, we identified lactucopicrin as a potent NF-kB antagonist. Notably, in silico docking and functional assays revealed lactucopicrin as a novel AHR modulator. Crucially, silencing AHR expression attenuated lactucopicrin-mediated NF-kB inhibition, uncovering a previously unrecognized AHR-NF-kB crosstalk mechanism. These findings not only position lactucopicrin as a dual-pathway modulator but also highlight the therapeutic potential of chicory-derived sesquiterpene lactones in treating inflammation-driven diseases and opening new avenues for leveraging natural products in targeted anti-inflammatory strategies.

molecular biology↗

bioRxiv2024

Intensive Chemotherapy Induces Cardiotoxicity via Reverse Electron Transport

Chemotherapy-induced cardiotoxicity has emerged as an important focus in oncology, driven by the growing number of cancer survivors. Intensive chemotherapies (iCT) used in the treatment of acute myeloid leukemia (AML) often lead to significant adverse cardiac events, which can reduce therapeutic benefits, limit treatment options, or even necessitate discontinuation--particularly for patients with pre-existing cardiac conditions, resulting in a loss of therapeutic opportunity. This study shows that the iCT triggers severe mitochondrial dysfunction in cardiac tissue, mirroring effects seen in ischemia-reperfusion models. Specifically, iCT results in succinate accumulation and elevated reactive oxygen species production, consistent with reverse electron transport phenomenon. Importantly, these effects were entirely prevented with RET inhibitors such as malonate or S1QEL1.1. In vivo, we demonstrate that malonate administration successfully prevents iCT-induced cardiotoxicity, maintaining left ventricular ejection fraction and fibrosis levels comparable to controls. Additionally, in an MLL-AF9-driven AML model, malonate sensitized leukemic cells to iCT. These findings support the dual potential of malonate: as an OXPHOS metabolism inhibitor to overcome chemoresistance in AML, while also reducing cardiotoxic risk for already vulnerable patients. One Sentence Summary: This study demonstrates that malonate prevents chemotherapy-induced cardiotoxicity by inhibiting mitochondrial reverse electron transport (RET), preserving cardiac function, and simultaneously sensitizing AML cells to intensive chemotherapy.

pathology↗

bioRxiv2024

Comprehensive evaluation of lifespan-extending molecules in C. elegans

The nematode C. elegans has long served as a gold-standard model organism in aging research, particularly since the discovery of long-lived mutants in conserved aging pathways including daf-2 (IGF1) and age-1 (PI3K). Its short lifespan and small size make it highly suitable for high throughput experiments. While numerous molecules have been tested for their effects on C. elegans lifespan, consensus is still lacking regarding the most effective and reproducible compounds. Confounding effects, especially those related to drug-bacteria interactions, remain a contentious issue in the literature. In this study, we evaluated 16 of the most frequently reported lifespan-extending molecules in C. elegans, examining their effects on lifespan with two different diets (live and UV-killed OP50). In addition, we assessed the compounds impact on bacterial growth, their effects on various nematode strains, and the impact of the starting age of treatment. Our findings first confirmed robust lifespan extension with many, but not all, of the 16 tested compounds from the literature, and revealed that some of them could be combined to get synergistic effects. Additionally, we showed that some of these compounds also extend lifespan in the fly D. melanogaster, demonstrating a conserved effect across species. Finally, by expanding our screen to a broader pool of molecules, we identified novel lifespan-extending compounds in C. elegans.

pharmacology and toxicology↗

bioRxiv2021

Altered interactions between circulating and tissue-resident CD8 T cells with the colonic mucosa define colitis associated with immune checkpoint inhibitors

Therapeutic blockade of co-inhibitory immune receptors PD-1 and CTLA-4 has revolutionized oncology, but treatments are limited by immune-related adverse events (IRAEs). IRAE Colitis (irColitis) is the most common, severe IRAE affecting up to 25% of patients on dual PD-1 and CTLA-4 inhibition. Here, we present a systems biology approach to define the cell populations and transcriptional programs driving irColitis. We collected paired colon mucosal biopsy and blood specimens from 13 patients with irColitis, 8 healthy individuals, and 8 controls on immune checkpoint inhibitors (ICIs), and analyzed them with single-cell/nuclei RNA sequencing with paired TCR and BCR sequencing, multispectral fluorescence microscopy, and secreted factor analysis (Luminex). We profiled 299,407 cells from tissue and blood and identified 105 cell subsets that revealed significant tissue remodeling in active disease. Colon mucosal immune populations were dominated by tissue-resident memory (TRM) ITGAE-expressing CD8 T cells representing a phenotypic spectrum defined by gene programs associated with T cell activation, cytotoxicity, cycling, and exhaustion. CD8 TRM and effector CD4 T cells upregulated type 17 immune programs (IL17A, IL26) and Tfh-like programs (CXCL13, PDCD1). We also identified for the first time an increased abundance of two KLRG1 and ITGB2-expressing CD8 T cell populations with circulatory cell markers, including a GZMK TRM-like population and a CX3CR1 population that is predicted to be intravascular. These two populations were more abundant in irColitis patients treated with dual PD-1/CTLA-4 inhibition than those receiving anti-PD-1 monotherapy. They also had significant TCR sharing with PBMCs, suggesting a circulatory origin. In irColitis we observed significant epithelial turnover marked by fewer LGR5-expressing stem cells, more transit amplifying cells, and upregulation of apoptotic and DNA-sensing programs such as the cGAS-STING pathway. Mature epithelial cells with top crypt genes upregulated interferon-stimulated pathways, CD274 (PD-L1), anti-microbial genes, and MHC-class II genes, and downregulated aquaporin and solute-carrier gene families, likely contributing to epithelial cell damage and absorptive dysfunction. Mesenchymal remodeling was defined by increased endothelial cells, both in irColitis patients and specifically in patients on dual PD-1/CTLA-4 blockade. Cell-cell communication analysis identified putative receptor-ligand pairs that recruit CD8 T cells from blood to inflamed endothelium and positive feedback loops such as the CXCR3 chemokine system that retain cells in tissue. This study highlights the cellular and molecular drivers underlying irColitis and provides new insights into the role of CTLA-4 and PD-1 signaling in maintaining CD8 TRM homeostasis, regulating CD8 T recruitment from blood, and promoting epithelial-immune crosstalk critical to gastrointestinal immune tolerance and intestinal barrier function.

immunology↗

bioRxiv2021

Characterization of an Agarophyton chilense oleoresin containing PPARγ natural ligands with insulin-sensitizing effects in a C57BL/6J mouse model of diet-induced obesity and antioxidant activity in Caenorhabditis elegans.

The biomedical potential of the edible red seaweed Agarophyton chilense (former Gracilaria chilensis) has not been explored. Red seaweeds are enriched in polyunsaturated fatty acids and eicosanoids, which are known natural ligands of the PPAR{gamma} nuclear receptor. PPAR{gamma} is the molecular target of thiazolidinediones (TZDs), drugs used as insulin sensitizers to treat type 2 diabetes mellitus. TZDs medical use is limited due to undesired side effects, the fact that it has triggered the search for partial agonists without the TZDs side effects. We produced A.chilense oleoresin (Gracilex(R)) that induces the activation of PPAR{gamma} without inducing adipocyte differentiation, similar to PPAR{gamma} partial agonists. In a model of high-fat diet-induced obesity in male mice, we showed that treatment with Gracilex(R) improves insulin sensitivity, normalizing altered glucose and insulin parameters. Gracilex(R) was enriched in palmitic acid, arachidonic acid, oleic acid, and lipophilic antioxidants such as {beta}-carotene and tocopherols. Gracilex(R) possesses antioxidant activity in vitro (CUPRAC) and increased the antioxidant capacity in vivo in Caenorhabditis elegans. These findings support the idea that Gracilex(R) is an excellent source of healthy lipids targeting PPAR{gamma} with antioxidant activity and a high nutraceutical value.

physiology↗

bioRxiv2019

CtIP promotes the motor activity of DNA2 to accelerate long-range DNA end resection

BLM or WRN helicases function with the DNA2 helicase-nuclease to resect DNA doublestrand breaks and initiate homologous recombination. Upon DNA unwinding by BLM/WRN, RPA directs the DNA2 nuclease to degrade the 5-strand, revealing the 3 overhang needed for recombination. RPA bound to ssDNA also represents a barrier, explaining the need for the motor activity of DNA2 to displace RPA prior to resection. Using ensemble and single molecule biochemistry, we show that phosphorylated CtIP dramatically stimulates the ATP hydrolysis driven motor activity of DNA2. This activation in turn strongly promotes the degradation of RPA-coated ssDNA by DNA2. The domains of CtIP required to stimulate DNA2 are separable from those that regulate the MRN complex. These results establish that CtIP couples both MRE11-dependent short and DNA2-dependent long-range resection, and show how the motor activity of DNA2 promotes resection. Our data explain the less severe resection defects of MRE11 nuclease-deficient cells compared to those lacking CtIP.\n\nHighlightsO_LIPhosphorylated CtIP stimulates the motor activity of DNA2\nC_LIO_LIThe activated DNA2 translocase facilitates degradation of RPA-coated ssDNA\nC_LIO_LICtIP promotes both MRN and DNA2 nucleases coupling short and long-range resection\nC_LIO_LIThe CtIP domains required to promote DNA2 and MRN are distinct and fully separable\nC_LI

biochemistry↗

bioRxiv2018

Spider mites escape bacterial infection by avoiding contaminated food

To fight infection, arthropods rely on the deployment of an innate immune response but also upon physical/chemical barriers and avoidance behaviours. However, most studies focus on immunity, with other defensive mechanisms being relatively overlooked.\n\nWe have previously shown that the spider mite Tetranychus urticae does not mount an induced immune response towards systemic bacterial infections, entailing very high mortality rates. Therefore, we hypothesized that other defence mechanisms may be operating to minimize infection risk. Here, we test (a) if spider mites are also highly susceptible to other infection routes - spraying and feeding - and (b) if they display avoidance behaviours towards infected food. Individuals sprayed with or fed on Escherichia coli or Pseudomonas putida survived less than the control, pointing to a deficient capacity of the gut epithelium, and possibly of the cuticle, to contain bacteria. Additionally, we found that spider mites prefer uninfected food to food contaminated with bacteria, a choice that probably does not rely on olfactory cues.\n\nOur results suggest that spider mites may rely mostly on avoidance behaviours to minimize bacterial infection and highlight the multi-layered nature of immune strategies present in arthropods.

ecology↗