Biology subjects

Lambert, F.

Publications and source records attributed to Lambert, F..

4 recordsLinked to original sources

bioRxiv2025

Quantification of H3.1-nucleosomes using a chemiluminescent immunoassay: a reliable method for neutrophil extracellular trap detection

Neutrophil extracellular traps (NETs) are chromatin-based web-like structures released by activated neutrophils in response to infectious agents. Overproduction or insufficient clearance of NETs contributes to dysfunction of immune response and disease pathogenesis, underlying the importance of early detection and monitoring of NET levels in clinical samples. While existing methods for NETs detection and quantification face limitations, there is a pressing need for a reliable, sensitive, and clinically applicable assay. Since NETs consist of long strains of decondensed chromatin, with nucleosomes as their basic units, we propose circulating H3.1-nucleosomes as biomarkers for NETs detection in clinical plasma samples. In the initial phase of our study, we confirmed the presence of H3.1-nucleosomes by immunofluorescence and immunoprecipitation experiments in two in vitro NET models: neutrophil-like cells differentiated from the HL-60 cell line and primary neutrophils isolated from whole blood, both treated with phorbol 12-myristate 13-acetate to induce NET formation. Subsequently, we developed and analytically validated a chemiluminescent immunoassay for the quantification of circulating H3.1-nucleosomes in plasma. This fully automated assay demonstrates impressive analytical performance in parameters including sensitivity, precision, linearity and reproducibility. Overall, by measuring the H3.1-nucleosome levels in plasma samples from patients suffering from NETs-related diseases compared to healthy donors, we demonstrated the assays potential as a groundbreaking diagnostic tool for disease management.

molecular biology↗

bioRxiv2023

Translational control of furina by an RNA regulon is essential for heart morphogenesis and cardiac valve function

Heart development is a complex process, starting from specification of cardiac precursors and formation of a linear tube to gradual progression to a functional beating organ. For normal heart development, many processes, including asymmetric positioning of the heart along the left-right (L/R) axis, cardiac growth, and cardiac valve morphogenesis must be completed successfully. Although heart development has been studied extensively, the mechanisms that control heart morphogenesis and valve formation are not fully understood. The pro-convertase FurinA is a key protein that functions in heart development in many vertebrates including zebrafish. How FurinA activity is regulated during heart development is not known. Through computational analysis of the zebrafish transcriptome, we identified a short sequence and structure RNA motif in a variant transcript of FurinA harbouring a long 3untranslated region (3UTR). The alternative 3UTR furina isoform is expressed at embryonic stages preceding organ positioning. Reporter localization and RNA-binding assays show that the furina 3UTR forms complexes with the conserved RNA-binding protein and translational repressor Ybx1. Conditional mutant zebrafish embryos affecting ybx1 show premature and increased Furin reporter protein expression, abnormal cardiac morphogenesis and heart looping defects. Our mutant ybx1 hearts have an expanded atrioventricular canal, abnormal sino-atrial valves and many mutant embryos show retrograde blood flow from the ventricle to the atrium. This is similar to human heart valve regurgitation patients. Our findings show an essential function for the 3UTR element/Ybx1 regulon in translational repression of FurinA, revealing a new upstream regulatory mechanism that controls embryonic heart development, and demonstrates the ybx1 mutant as a model to study cardiac valve development and function.

developmental biology↗

bioRxiv2022

Pytri: A multi-weight detection system for biological entities

The enumeration of biological entities is a critical part of experimental assays and usually requires large lengths of time. The standard method is to count the entities by hand or with OpenCV-based software, which can lead to inaccurate results. Here, we propose an online platform for biologists consisting of a system with multiple trained machine learning weights to detect various biological entities such as yeast colonies, bacterial colonies, and melanoma clusters. The Pytri model achieved a median relative error rate of 7.56% for bacterial and yeast colonies on Petri dishes, 6.58% for colonies on 96-well plates and 10.28% for melanoma cluster microscopy images. We showcase the application of state-of-the-art deep learning tools in bacterial entity detection, achieving significantly higher accuracy than traditional methods when compared to our base standard manual count.

bioinformatics↗

bioRxiv2021

Time-resolved secretome analysis of three Colletotrichum species identifies copper radical alcohol oxidases for the production of fatty aldehydes

Copper Radical Alcohol Oxidases (CRO-AlcOx), which have been recently discovered among fungal phytopathogens are attractive for the production of fragrant fatty aldehydes. To investigate the secretion of CRO-AlcOx by natural fungal strains, we undertook time-course analyses of the secretomes of three Colletotrichum species (C. graminicola, C. tabacum and C. destructivum) using proteomics. The addition of a copper-manganese-ethanol mixture to Colletotrichum cultures unexpectedly induced the secretion of up to 400 proteins, 29-52% of which were carbohydrate-active enzymes (CAZymes), including a wide diversity of copper-containing oxidoreductases from the auxiliary activities (AA) class (AA1, AA3, AA5, AA7, AA9, AA11-AA13, AA16). Under these specific conditions, while a CRO-glyoxal oxidase from the AA5_1 subfamily was among the most abundantly secreted proteins, the targeted AA5_2 CRO-AlcOx were secreted at lower levels, suggesting heterologous expression as a more promising strategy for CRO-AlcOx production and utilization. C. tabacum and C. destructivum CRO-AlcOx were expressed in Pichia pastoris and their preference toward both aromatic and aliphatic primary alcohols was assessed. The CRO-AlcOx from C. destructivum was further investigated in applied settings, revealing a full conversion of C6 and C8 alcohols into their corresponding fragrant aldehydes. IMPORTANCEIn the context of the industrial shift toward greener processes, the biocatalytic production of aldehydes is of utmost interest owing to their importance as intermediates in preparative chemistry and for their use as flavors and fragrances ingredients. In the search for new biocatalysts, CRO-AlcOx have the potential to become platform enzymes for the oxidation of alcohols to aldehydes. The use of crude fungal secretomes is often seen has an appealing approach by industries since alleviating various costs pertaining to biocatalysts production. However, the secretion of CRO-AlcOx by natural fungal strains has never been explored. This study showed that Colletotrichum species can secrete a broad diversity of copper-containing enzymes, but only little amount of CRO-AlcOx. Thus, recombinant expression remains the most promising approach. The potential of CRO-AlcOx as biocatalyst for flavor and fragrance applications was confirmed through the production of two new enzymes with activity on fatty alcohols.

biochemistry↗