Biology subjects

Casas, J.

Publications and source records attributed to Casas, J..

14 recordsLinked to original sources

bioRxiv2025

Spatial Regulation of Lck Activation at the Immune Synapse Revealed by a FRET-Based Biosensor

T cell receptor (TCR) signaling is critically dependent on the Src-family kinase Lck, whose activation is tightly regulated both spatially and conformationally during antigen recognition. Here, we employ an improved second-generation FRET-based biosensor (TqLckV2.3) to visualize Lck conformational dynamics in live T cells with high spatial resolution. Upon TCR engagement, we observe a paradoxical increase in whole-cell FRET signal, which immunolabeling reveals to be due to selective internalization of inactive Lck (pY505), while active Lck (pY394) remains membrane-associated and enriched at the immune synapse (IS). Using CD8 mutants that disrupt Lck binding, we demonstrate that free Lck undergoes more pronounced conformational activation than CD8-bound Lck. Furthermore, we show that C-terminal Src kinase (Csk) preferentially phosphorylates free Lck at Y505, while CD45 suppresses its activation via dephosphorylation of Y394, suggesting a dual regulatory mechanism that maintains free Lck in an inactive state under resting conditions. High-resolution imaging confirms sustained activation of Lck at the IS and transient inactivation at the periphery, revealing a spatially confined signaling architecture. These findings uncover a novel regulatory mechanism involving selective internalization and spatial segregation of Lck conformations and establish TqLckV2.3 as a powerful tool for dissecting TCR signaling dynamics.

immunology↗

bioRxiv2025

Olfactory performance explains duality of antennal architectural designs in Lepidoptera

Male attraction by females through sex pheromones is widespread among Lepidoptera, and antennae are key olfactory organs during male orientation. Broadly speaking, two designs of antennae coexist in Lepidoptera: complex (pectinate) or stick-like (filiform) ones. Pectinate antennae have attracted attention because of their multiscale geometry, assumed to outperform filiform. Yet, the filiform design is by far more common. We compare the olfactory performance of the two designs using modeling, particle image velocimetry on 3D-printed scaled-up models, and computational simulations. In terms of absolute odor capture, pectinate antennae perform better at nearly all flying speeds. However, when considering drag, filiform designs are more energy-efficient than pectinate ones at low flight speeds, while the reverse holds at high speeds. This is due to the differential scaling of drag and molecule capture with flight speed. According to our results, small and slow moths would bear filiform antennae whereas big and fast moths would have pectinate ones, which is the general trend observed in Nature. We discuss exceptions to this general pattern and how species could evolve from one design to the other by investigating the influence of the antennal structural elements.

physiology↗

bioRxiv2022

Pharmacological elevation of cellular dihydrosphingomyelin provides a novel antiviral strategy against West Nile virus infection

Flavivirus life cycle is strictly dependent on cellular lipid metabolism. Polyphenols like gallic acid and its derivatives are promising lead compounds for new therapeutic agents as they can exert multiple pharmacological activities, including the alteration of lipid metabolism. The evaluation of our own collection of polyphenols against West Nile virus, a representative medically relevant flavivirus, led to the identification of N,N'-(dodecane-1,12-diyl)bis(3,4,5-trihydroxybenzamide) and its 2,3,4-trihydroxybenzamide regioisomer as selective antivirals with low cytotoxicity and high antiviral activity (EC50 of 2.2 and 0.24 M, respectively in Vero cells; EC50 of 2.2 and 1.9 M, respectively in SH-SY5Y cells). These polyphenols also inhibited the multiplication of other flaviviruses, namely Usutu, dengue, and Zika viruses, exhibiting lower antiviral or negligible antiviral activity against other RNA viruses. The mechanism underlying their antiviral activity against WNV involved the alteration of sphingolipid metabolism. These compounds inhibited ceramide desaturase (Des1) promoting the accumulation of dihydrosphingomyelin (dhSM), a minor component of cellular sphingolipids with important roles on membrane properties. Addition of exogenous dhSM, or Des1 blockage by using the reference inhibitor GT-11, confirmed the involvement of this pathway in WNV infection. These results unveil the potential of novel antiviral strategies based on the modulation of the cellular levels of dhSM and Des1 activity for the control of flavivirus infection.

microbiology↗

bioRxiv2022

Beta-Cyclodextrins as affordable antivirals to treat coronavirus infection

The SARS-CoV-2 pandemic made evident that we count with few coronavirus-fighting drugs. Here we aimed to identify a cost-effective antiviral with broad spectrum activity and high safety and tolerability profiles. We began elaborating a list of 116 drugs previously used to treat other pathologies or characterized in pre-clinical studies with potential to treat coronavirus infections. We next employed molecular modelling tools to rank the 44 most promising inhibitors and tested their efficacy as antivirals against a panel of and {beta} coronavirus, e.g., the HCoV-229E and SARS-CoV-2 viruses. Four drugs, OSW-1, U18666A, hydroxypropyl-{beta}-cyclodextrin (H{beta}CD) and phytol, showed antiviral activity against both HCoV-229E (in MRC5 cells) and SARS-CoV-2 (in Vero E6 cells). The mechanism of action of these compounds was studied by transmission electron microscopy (TEM) and by testing their capacity to inhibit the entry of SARS-CoV-2 pseudoviruses in ACE2-expressing HEK-293T cells. The entry was inhibited by H{beta}CD and U18666A, yet only H{beta}CD could inhibit SARS-CoV-2 replication in the pulmonary cells Calu-3. With these results and given that cyclodextrins are widely used for drug encapsulation and can be safely administered to humans, we further tested 6 native and modified cyclodextrins, which confirmed {beta}-cyclodextrins as the most potent inhibitors of SARS-CoV-2 replication in Calu-3 cells. All accumulated data points to {beta}-cyclodextrins as promising candidates to be used in the therapeutic treatments for SARS-CoV-2 and possibly other respiratory viruses.

microbiology↗

bioRxiv2022

Obesity promotes Fumonisin B1 toxicity and induces hepatitis

Background and aimObesity is a major public health issue worldwide. Obesity is associated with chronic inflammation that contribute to long-term complications, including insulin resistance, type 2 diabetes and non-alcoholic fatty liver disease. We hypothesized that obesity may also influence the sensitivity to food contaminants, such as fumonisin B1 (FB1), a mycotoxin produced mainly by the Fusarium verticillioides. FB1, a common contaminant of corn, is the most abundant and best characterized member of the fumonisins family. This toxin provokes severe mycotoxicosis in animals, which leads to hepatotoxicity and alterations in the immune response and intestinal barrier permeability. We investigated here whether diet-induced obesity could modulate the sensitivity to oral FB1 exposure, with emphasis on gut health and hepatotoxicity. MethodsThe metabolic effects of FB1 were assessed in obese and non-obese male C57BL/6J mice. For 15 weeks, mice received a high-fat diet (HFD) or normal chow diet (CHOW). During the last three weeks, mice were exposed or not to FB1 (10 mg/kg body weight/day) through drinking water. ResultsAs expected, HFD feeding induced significant body weight gain, glucose intolerance, and hepatic steatosis. FB1-exposed mice displayed a higher sphinganine/sphingosine ratio, a well-known FB1 biomarker of exposure, due to inhibition of ceramide synthases activity by FB1. Combined exposure to HFD and FB1 resulted in body weight loss and a decrease in fasting blood glucose level. This co-exposition also induces gut dysbiosis, an increase in plasma FB1 level, a decrease in liver weight and hepatic steatosis. Moreover, plasma transaminase levels were significantly increased and associated with liver inflammation in HFD/FB1-treated mice. Liver gene expression analysis revealed that the combined exposure to HFD and FB1 was associated with reduced expression of genes involved in lipogenesis and increased expression of immune response and cell cycle-associated genes. ConclusionThese results suggest that, in the context of obesity, FB1 exposure promotes gut dysbiosis and severe liver inflammation. To our knowledge, this study provides the first example of obesity-induced hepatitis in response to a food contaminant.

pharmacology and toxicology↗

bioRxiv2021

Active sensing in bees through antennal movements is independent of odor molecule

When sampling odors, many insects are moving their antennae in a complex but repeatable fashion. Previous works with bees have tracked antennal movements in only two dimensions, with a low sampling rate and with relatively few odorants. A detailed characterization of the multimodal antennal movement patterns as function of olfactory stimuli is thus wanting. The aim of this study is to test for a relationship between the scanning movements and the properties of the odor molecule. We tracked several key locations on the antennae of 21 bumblebees at high frequency (up to 1200 fps) and in three dimensions while submitting them to puffs of 11 common odorants released in a low-speed continuous flow. To cover the range of diffusivity and molecule size of most odors sampled by bees, compounds as different as butanol and farnesene were chosen, with variations of 200% in molar masses. Water and paraffin were used as negative controls. Movement analysis was done on the tip, the scape and the base of the antennae tracked with the neural network Deeplabcut. Bees use a stereotypical motion of their antennae when smelling odors, similar across all bees, independently of the identity of the odors and hence their diffusivity. The variability in the movement amplitude among odors is as large as between individuals. The first oscillation mode at low frequencies and large amplitude (ca. 1-3 Hz, ca. 100{degrees}) is triggered by the presence of an odor and is in line with previous work, as is the speed of movement. The second oscillation mode at higher frequencies and smaller amplitude (40 Hz, ca. 0.1{degrees}) is constantly present. Antennae are quickly deployed when a stimulus is perceived, decorrelate their movement trajectories rapidly and oscillate vertically with a large amplitude and laterally with a smaller one. The cone of air space thus sampled was identified through the 3D understanding of the motion patterns. The amplitude and speed of antennal scanning movements seem to be function of the internal state of the animal, rather than determined by the odorant. Still, bees display an active olfaction strategy. First, they deploy their antennae when perceiving an odor rather than let them passively encounter it. Second, fast vertical scanning movements further increase the flow speed experienced by an antenna and hence the odorant capture rate. Finally, lateral movements might enhance the likelihood to locate the source of odor, similarly to the lateral scanning movement of insects at odor plume boundaries. Definitive proofs of this function will require the simultaneous 3D recordings of antennal movements with both the air flow and odor fields.

animal behavior and cognition↗

bioRxiv2021

Intracellular pools of DAG-activated TRPC3 channels are essential for TLR4 activation

Toll-like receptor 4, the receptor for bacterial lipopolysaccharide (LPS), drives inflammatory responses that protect against pathogens and boost the adaptive immunity. LPS responses are known to depend on calcium fluxes, but the molecular mechanisms involved are poorly understood. Here we present evidence that the transient receptor potential canonical channel 3 (TRPC3) is activated intracellularly during macrophage exposure to LPS and is essential for Ca2+ release from internal stores. In this way TRPC3 participates in cytosolic Ca2+ elevations, TLR4 endocytosis, activation of inflammatory transcription factors and cytokine upregulation. We also report that TRPC3 is activated by diacylglycerol (DAG) generated by the phosphatidic acid phosphatase lipin-1. In accord with this, lipin-1-deficient cells show reduced Ca2+ responses to LPS challenge. A cameleon indicator directed to the endoplasmic reticulum shows that this is the organelle from which TRPC3 mediates the Ca2+ release. Finally, pharmacological inhibition of TRPC3 reduces systemic inflammation induced by LPS in mice. Collectively, our study unveils a central component of LPS-triggered Ca2+ signaling that involves intracellular sensing of lipin-1-derived DAG by TRPC3.

immunology↗

bioRxiv2021

Catabolism of lysosome-related organelles in color-changing spiders supports intracellular turnover of pigments

Pigment organelles of vertebrates belong to the lysosome-related organelle (LRO) family, of which melanin-producing melanosomes are the prototypes. While their anabolism has been extensively unraveled through the study of melanosomes in skin melanocytes, their catabolism remains poorly known. Here, we tap into the unique ability of crab spiders to reversibly change body coloration to examine the catabolism of their pigment organelles. By combining ultrastructural and metal analyses on high-pressure frozen integuments, we first assess whether pigment organelles of crab spiders belong to the LRO family and, second, how their catabolism is intracellularly processed. Using scanning-transmission electron microscopy, electron tomography and nanoscale Synchrotron-based scanning X-ray fluorescence, we show that pigment organelles possess ultrastructural and chemical hallmarks of LROs, including intraluminal vesicles and metal deposits, similar to melanosomes. Monitoring ultrastructural changes during bleaching suggests that the catabolism of pigment organelles involves the degradation and removal of their intraluminal content, possibly through lysosomal mechanisms. In contrast to skin melanosomes, anabolism and catabolism of pigments proceed within the same cell without requiring either cell death or secretion/phagocytosis. Our work hence provides support for the hypothesis that the endolysosomal system is fully functionalized for within-cell turnover of pigments, leading to functional maintenance under adverse conditions and phenotypic plasticity. First formulated for eye melanosomes in the context of human vision, the hypothesis of intracellular turnover of pigments gets unprecedented strong support from pigment organelles of spiders.

cell biology↗

bioRxiv2020

CCR5 deficiency impairs CD4+ T cell memory responses and antigenic sensitivity through increased ceramide synthesis

In CD4+ T cells, CCR5 is not only a coreceptor for HIV-1 infection, but also contributes to their functional fitness. Here we show that by limiting GATA-1-induced transcription of specific ceramide synthases, CCR5 signaling reduces ceramide levels and thereby increases T cell antigen receptor (TCR) nanoclustering in antigen-experienced mouse and human CD4+ T cells. This activity is CCR5-specific and independent of CCR5 costimulatory activity. CCR5-deficient mice showed reduced production of high affinity class-switched antibodies, but only after antigen rechallenge, which implies an impaired memory CD4+ T cell response. This study identifies a CCR5 function in the generation of CD4+ T cell memory responses, and establishes an antigen-independent mechanism that regulates TCR nanoclustering by altering specific lipid species.

immunology↗

bioRxiv2019

Field application of the geometric framework reveals a multistep strategy of nutrient regulation in a leaf-miner

Animals have evolved a vast array of behavioral and physiological strategies that allow them to achieve a nutritionally balanced diet. Plants as food for herbivores are often considered suboptimal, but phytophagous insects can employ pre- and post-ingestive mechanisms and/or symbiotic associations to help overcome food nutritional imbalances. This is particularly crucial for permanent multivoltine leaf-miner insects such as the caterpillar Phyllonorycter blancardella which completes development within a restricted area of a single leaf and use deciduous leaves to fuel growth and reproduction even under senescing autumnal conditions. Using the geometric framework for nutrition under natural field conditions, we show that this insect has multiple strategies to deal with inadequate food supply from the plant. First, larvae manipulate the protein-sugar content of both normal, photosynthetically active, and senescing, photosynthetically inactive, leaf tissues. Control of nutritional homeostasis of mined tissues is however higher for late instars, which differ from younger larval instars in their feeding mode (fluid-vs. tissue-feeder). Second, slight differences in the protein-sugar environment remain between mined tissues on green and yellow leaves despite this manipulation of the leaf physiology. This insect uses post-ingestive mechanisms to achieve similar body protein, sugar and lipid composition. This study demonstrates, for the first time under natural conditions, the ability of an insect herbivore to practice a combination of pre- and post-ingestive compensatory mechanisms to attain similar growth and metabolic outcomes in fundamentally different nutritional environments. Additionally, a comparison of larval nutritional requirements of 117 species from various insect groups further reinforces the hypothesis of a close association between P. blancardella and endosymbiotic bacteria for nutritional purposes.

ecology↗

bioRxiv2019

Manipulation of plant primary metabolism by leaf-mining larvae in the race against leaf senescence

Plants are often considered as suboptimal food for phytophagous insects, requiring them to employ various adaptive mechanisms to overcome food nutritional imbalances. This could include host-plant manipulation and/or symbiotic associations. The extensive reconfiguration of plant primary metabolism upon herbivory, as well as its impact on herbivores, have been largely overlooked, while studies investigating secondary metabolites is extensive. Here, we document how the apple leaf-mining caterpillar Phyllonorycter blancardella, a highly-specialized insect which completes development within a restricted area of a single Malus domestica leaf over successive different larval feeding modes, maintains nutrient-rich green tissues in its feeding area on green and senescent leaves. For this purpose, we quantified a large number of compounds involved in plant primary metabolism: starch, total soluble sugars, five individual sugars, twenty protein-bound amino acids and twenty free amino acids. Plant alteration can be observed not only on senescing (photosynthetically inactive) but also normal (photosynthetically active) leaf tissues of its host-plant to compensate for detrimental environmental variations. Our results show a differential control of the primary metabolism depending on the larva developmental stage, itself correlated to the fluid-feeding and tissue-feeding modes. Our results also suggest that leaf amino acid alterations favor a faster insect development. Finally, chemical scores indicate that the most growth-limiting essential amino acids are also common to other phytophagous insects and large herbivores, suggesting that these limitations are a general consequence of using plants as food source. We discuss the possible mechanisms responsible for these different manipulative capacities, as well as their ecological implications.

plant biology↗

bioRxiv2019

Biological identification and localization of uncyclized xanthommatin, a key intermediate in ommochrome biosynthesis: an in vitro-in vivo study

Ommochromes are widespread pigments that mediate multiple functions in invertebrates. The two main families of ommochromes are ommatins and ommins, which both originate from the kynurenine pathway but differ in their backbone, thereby in their coloration and function. Despite its broad significance, how the structural diversity of ommochromes arises in vivo has remained an open question since their first description. In this study, we combined organic synthesis, analytical chemistry and organelle purification to address this issue. From a set of synthesized ommatins, we derived a fragmentation pattern that helped elucidating the structure of new ommochromes. We identified uncyclized xanthommatin as the elusive biological intermediate that links the kynurenine pathway to the ommatin pathway within ommochromasomes, the ommochrome-producing organelles. Due to its unique structure, we propose that uncyclized xanthommatin functions as a key branching metabolite in the biosynthesis and structural diversification of ommatins and ommins, from insects to cephalopods.

biochemistry↗

bioRxiv2018

Phenome-wide association analysis of LDL-cholesterol lowering genetic variants in PCSK9

BackgroundWe characterised the phenotypic consequence of genetic variation at the PCSK9 locus and compared findings with recent trials of pharmacological inhibitors of PCSK9.\n\nMethodsPublished and individual participant level data (300,000+ participants) were combined to construct a weighted PCSK9 gene-centric score (GS). Fourteen randomized placebo controlled PCSK9 inhibitor trials were included, providing data on 79,578 participants. Results were scaled to a one mmol/L lower LDL-C concentration\n\nResultsThe PCSK9 GS (comprising 4 SNPs) associations with plasma lipid and apolipoprotein levels were consistent in direction with treatment effects. The GS odds ratio (OR) for myocardial infarction (MI) was 0.53 (95%CI 0.42; 0.68), compared to a PCSK9 inhibitor effect of 0.90 (95%CI 0.86; 0.93). For ischemic stroke ORs were 0.84 (95%CI 0.57; 1.22) for the GS, compared to 0.85 (95%CI 0.78; 0.93) in the drug trials. ORs with type 2 diabetes mellitus (T2DM) were 1.29 (95% CI 1.11; 1.50) for the GS, as compared to 1.00 (95%CI 0.96; 1.04) for incident T2DM in PCSK9 inhibitor trials. No genetic associations were observed for cancer, heart failure, atrial fibrillation, chronic obstructive pulmonary disease, or Alzheimers disease - outcomes for which large-scale trial data were unavailable.\n\nConclusionsGenetic variation at the PCSK9 locus recapitulates the effects of therapeutic inhibition of PCSK9 on major blood lipid fractions and MI. Apparent discordance between genetic associations and trial outcome for T2DM might be explained lack by a of statistical precision, or differences in the nature and duration of genetic versus pharmacological perturbation of PCSK9.\n\nFundingThis research was funded by the British Heart Foundation (SP/13/6/30554, RG/10/12/28456, FS/18/23/33512), UCL Hospitals NIHR Biomedical Research Centre, by the Rosetrees and Stoneygate Trusts.\n\nCondensed abstractEvidence on the long-term efficacy and safety of therapeutic inhibition of PCSK9 is lacking. To explore potential long-term effects of PCSK9 inhibition, we characterised the phenotypic consequence of LDL-cholesterol lowering variants at the PCSK9 locus. A PCSK9 gene score comprising 4 SNPs recapitulated the effects of therapeutic inhibition of PCSK9 on major blood lipid fractions and risk of myocardial infarction, and was associated with an increased risk of type 2 diabetes. No associations with safety outcomes such as cancer, COPD, Alzheimers disease or atrial fibrillation were identified. Our findings suggest PCSK9 inhibition may be safe and effective during prolonged use.

genetics↗

bioRxiv2018

Underestimation of carbohydrates by sugar alcohols in classical anthrone-based colorimetric techniques compromises insect metabolic and energetic studies

Physiologically based metabolic studies usually search for easy, sensitive, and cheap techniques to rapidly estimate biological parameters such as nutrient content. Colorimetric methods to estimate carbohydrates have been extensively used (over 120,000 references). However, sugar alcohols are underestimated under conventionally used analytical conditions, in particular if using the popular van Handel method. This may lead to misinterpretations of sugar implications in biological systems. We determined the anthrone reaction with various sugar alcohols and non-alcohols individually under standard conditions (Van Handel 1985). We then manipulated the proportion of either sugar alcohols or non-alcohols in three different sugar mixtures in order to estimate the impact on the total sugar estimation. In the case of a mixture with over 50% of sugar alcohols, total sugars are underestimated by 50% when using glucose as standard.

ecology↗