Biology subjects

Pujol, M.

Publications and source records attributed to Pujol, M..

12 recordsLinked to original sources

bioRxiv2026

The VGLUT3-p.T8I Mutation Alters Striatal Acetylcholine Dynamics in Response to Social Stressors and Drives Selective Social Avoidance in Male Mice

The nucleus accumbens (NAc) is a key structure translating dopaminergic inputs into social avoidance following repeated social defeat. Within the NAc, dopamine (DA) release is modulated in part by cholinergic interneurons (CINs). CINs express the vesicular glutamate transporter type 3 (VGLUT3), which supports glutamate co-release and regulates vesicular acetylcholine (ACh) loading, thereby fine-tuning cholinergic signaling. A rare human VGLUT3-p.T8I missense variant is associated with compulsive behaviors toward food and drugs of abuse. We hypothesized that p.T8I disrupts NAc ACh dynamics and affects social avoidance following social defeat. Male and female mice expressing the VGLUT3-p.T8I variant were exposed to a 10-day chronic social defeat stress (CSDS) paradigm. Social avoidance toward the aggressor strain and a C57BL/6 (B6) conspecific was measured using the social interaction and 3-chamber tests, and anxiety-like behavior was evaluated in the elevated zero maze. Fiber photometry was used to monitor ACh and DA dynamics in the NAc medial shell (NAcMsh) during CSDS and social interaction. After CSDS, VGLUT3T8I/T8I males showed aggressor strain-specific social avoidance and increased anxiety-like behavior compared to WT mice. Mutants showed blunted CSDS-induced ACh increase during attacks but enhanced ACh release upon re-exposure to the CD1 aggressor strain correlating with avoidance severity. DA dynamics were minimally affected. Females, regardless of genotype, displayed a male-mutant-like phenotype. In males, the VGLUT3-p.T8I variant alters NAcMsh ACh signaling and promotes selective social avoidance toward aggressors and elevated anxiety after CSDS, highlighting a potential cholinergic mechanism underlying stress-induced social avoidance with broader relevance to psychiatric disorders.

neuroscience↗

bioRxiv2025

Environmental Enrichment Attenuates Fentanyl-Seeking Behavior and Protects Against Stress-Induced Reinstatement in Both Male and Female Rats

Environmental enrichment (EE) reduces vulnerability to multiple drugs of abuse, yet its impact on fentanyl use and relapse-like behavior remains unclear. Here, we tested whether long-term, non-social, object-based EE alters fentanyl self-administration, extinction, and stress-induced reinstatement in male and female rats. Rats were individually housed in either standard non-enriched (NE) conditions or in EE cages containing a rotating set of novel objects beginning at least three days prior to self-administration. EE did not impact acquisition of fentanyl self-administration but reduced fentanyl intake during maintenance of self-administration and reduced the persistence of drug-seeking in extinction. Following extinction, yohimbine robustly reinstated drug-seeking behavior in NE rats but reinstatement in EE rats was markedly attenuated, indicating reduced sensitivity to stress-induced relapse triggers. Circulating corticosterone levels were lower in EE rats across the experiment and were positively correlated with reinstatement responding, suggesting that enrichments protective effects may be mediated in part by reduced hypothalamic-pituitary-adrenal (HPA) axis activation. These findings demonstrate that object-based EE, even in the absence of social enrichment, is sufficient to blunt fentanyl use, facilitate extinction, and constrain stress-induced reinstatement. The results highlight the role of environmental context and stress regulation in fentanyl vulnerability and suggest that enrichment-inspired, non-social interventions may offer a scalable strategy to reduce opioid use and relapse risk. Significance StatementThis study demonstrates that long-term, object-based environmental enrichment in the absence of social peers is sufficient to reduce fentanyl intake and blunt stress-induced reinstatement in male and female rats. These findings identify a simple, scalable environmental intervention capable of dampening both drug use and vulnerability to relapse triggers. Given the prevalence of individuals exposed to periods of social isolation or with limited access to support, understanding how non-peer-based enrichment influences maladaptive opioid use and stress reactivity is of timely and translational importance. The ability of enrichment to reduce corticosterone, fentanyl consumption, and drug-cue reactivity highlights the importance of environmental context in shaping opioid misuse liability and implicates novel and practical avenues for relapse prevention

animal behavior and cognition↗

bioRxiv2025

STAT3 operates as an inflammation-dependent transcriptional switch

Signal transducer and activator of transcription 3 (STAT3) is a key regulator of immune cell function, but its role in lymphocytes remains incompletely understood. Here, we show that STAT3 has a context-dependent function in antiviral natural killer (NK) cells, either promoting or impairing adaptive NK cell responses dependent on the level of inflammation. STAT3 is recruited to distinct genomic sites under homeostatic versus inflammatory environments, where it drives different transcriptional programs. Through this re-localization, STAT3 regulates downstream transcription factors MYB and BLIMP-1 in an inflammation-dependent manner to shape NK cell differentiation under homeostasis and during infection. Thus, STAT3 acts as a transcriptional switch that integrates cytokine signals to control lymphocyte adaptation to different environments. This mechanism highlights how therapeutic interventions targeting STAT3 can result in different outcomes depending on the degree of inflammation. HIGHLIGHTS- STAT3 exerts a context-dependent role on adaptive NK cells during viral infection - STAT3 modulates IL-15 signaling by competing with STAT5 and regulating MYB - Homeostatic versus inflammatory cytokines relocate STAT3 to distinct genomic sites - Downstream transcription factors MYB and BLIMP-1 govern STAT3-dependent differentiation

immunology↗

bioRxiv2025

A melon (Cucumis melo) homologue of REPRESSOR OF PHOTOSYNTHETIC GENES prevents chloroplast differentiation in the fruit flesh

Fruit flesh color in melon can be orange, green or white, depending on the accumulation of the orange carotenoid {beta}-carotene or / and green chlorophylls. The dominant allele of Green flesh (Gf) causes orange melons, but in the absence of this allele the flesh of ripe melon can be white or green depending on the White flesh (Wf) locus, being white dominant over green. The identity of Wf has remained unclear despite several candidates have been proposed. Here we identified Wf by fine mapping of a segregating population derived from the white-fleshed variety Piel de Sapo (PS, gf gf / Wf Wf) and the orange-fleshed Vedrantais (VED, Gf Gf / wf wf). Wf corresponds to the gene MELO3C003098, herein referred to as CmRPGE1 as it encodes a fruit-specific homologue of REPRESSOR OF PHOTOSYNTHETIC GENES (RPGE) microproteins. Similar to RPGE homologues from other plants, overexpression of the PS allele (CmRPGE1PS) caused a pale green leaf phenotype in Nicotiana benthamiana and Arabidopsis thaliana. By contrast, a 10-nucleotide deletion in the VED allele (CmRPGE1VED) resulted in a loss of RPGE function. The active CmRPGE1PS microprotein interacts with a fruit-localized melon homologue of ARABIDOPSIS PSEUDO-RESPONSE REGULATOR2 (APRR2), a GARP family transcription factor. Binding of CmRPGE1PS retains the melon APRR2 homologue in the cytosol, hence preventing the regulation of target genes involved in chloroplast biogenesis. In green fruit cultivars, the non-functional CmRPGE1VED allele allows APRR2 to perform its function, leading to chloroplast development and consequently a green flesh phenotype. SIGNIFICANT STATEMENTFruit color is a critical quality trait that directly influences consumer preference and market value in melon. In this study, we demonstrate that the White flesh (Wf) gene, a major determinant of melon fruit flesh color, encodes a fruit-specific homologue of REPRESSOR OF PHOTOSYNTHETIC GENES (RPGE) microproteins. The encoded microprotein, CmRPGE1, interacts with melon homologues of ARABIDOPSIS PSEUDO-RESPONSE REGULATOR2 (APRR2), a GARP family transcription factor. In white flesh melons, the CmAPRR2.2 paralog is sequestered in the cytoplasm upon interaction with CmRPGE1, thereby preventing the expression of target genes involved in chlorophyll accumulation. Conversely, the truncated version of CmRPGE1 present in green-fleshed melons fails to interact with CmAPRR2.2 and to prevent its nuclear localization and function, allowing chloroplasts differentiation.

plant biology↗

bioRxiv2025

Outer membrane vesicles from Bacteroides fragilis contain coding and non-coding small RNA species that modulate inflammatory signaling in intestinal epithelial cells.

Alterations to the community structure and function of the microbiome are associated with changes to host physiology, including immune responses. However, the contribution of microbe-derived RNAs carried by outer membrane vesicles (OMVs) to host immune responses remain unclear. This study investigated the role of OMVs and OMV-associated small RNA (sRNA) species from pathogenic and commensal Bacteroides fragilis (ETBF and NTBF respectively) in eliciting different immune responses from intestinal epithelial cells. To distinguish the differences in the sRNA profiles of the two strains and their OMVs, RNA-seq, qRT-PCR, and northern blotting were conducted to identify enrichment of discrete sRNA species in OMVs, which were also differentially expressed between the two strains. Specifically, both coding and non-coding RNAs were enriched in OMVs from NTBF and ETBF, with BF9343_RS22680 and BF9343_RS17870 being significantly enriched in ETBF OMVs compared to NTBF. To understand the effects of OMVs on pattern recognition receptors, reporter cells of Toll-like receptor (TLR) activation were treated with OMVs, demonstrating activation of TLRs 2, 3, and 7. Treatment of Caco-2 and HT29-MTX cells with OMVs demonstrated increased expression of IL-8. Surprisingly, we discovered that degradation of RNase-accessible RNAs within ETBF OMVs, but not NTBF OMVs, resulted in vesicles with enhanced capacity to stimulate IL-8 expression, indicating that these extravesicular RNAs exert an immunosuppressive effect. This suggests a dual role for OMV-associated RNAs in modulating host immune responses, with implications for both bacterial pathogenesis and therapeutic applications. Graphical Abstract

molecular biology↗

bioRxiv2024

TILLCANN: A TILLING platform in Cannabis sativa for mutation discovery and crop improvement

Cultivation of Cannabis sativa is increasing because of its therapeutic value and recognition as a multi-purpose and sustainable crop. Targeting Induced Local Lesions in Genomes (TILLING) is a versatile reverse genetics approach that unlocks induced variation through mutagenesis to accelerate the development of new cultivars and contribute to the functional validation of genes. Increasingly efficient next-generation sequencing technologies and genomic resources have combined to make TILLING by sequencing (TbyS) an attractive technique that can be applied in cannabis. Here we describe the development of a mutagenesis protocol for C. sativa and the development of the TILLCANN platform composed of 1,633 M2 families. As a demonstration of the functionality of the platform, we used TbyS to perform a high throughput screening of novel mutations in amplicons for genes associated with important agronomic and biochemical traits in a set of 512 M2 families. We confirmed 14 of the identified mutations and calculated an average mutation frequency range of 1/263 to 1/320 kb. We found that heterozygous mutants in the cannabis homologue of Class II TEOSINTE BRANCHED 1/CYCLOIDEA/PCF (TCP4) are linked to alterations in leaf number and morphology. We expect that the novel genetic variability unlocked in the TILLCANN platform for performing forward or reverse genetic screens can significantly boost breeding programs geared toward both medicinal cannabis and industrial hemp. Gene & accession numbersRaw sequencing data generated in this study was deposited at the European Nucleotide Archive with accession numbers xxx and xxx. Genes and accession numbers discussed within this manuscript correspond to the Cannabis sativa cs10 genome assembly deposited in National Center for Biotechnology Information with accession number GCA_900626175.2.

plant biology↗

bioRxiv2024

Dominant induction of the inflammasome by the SARS-CoV-2 accessory protein ORF9b, abrogated by small-molecule ORF9b homodimerization inhibitors

Viral accessory proteins play critical roles in viral escape form host innate immune responses and in viral inflammatory pathogenesis. Here we show that the SARS-CoV-2 accessory protein, ORF9b, but not other SARS-CoV-2 accessory proteins (ORF3a, ORF3b, ORF6, ORF7, ORF8, ORF9c, ORF10), strongly activates inflammasome-dependent caspase-1 in A549 lung carcinoma cells and THP-1 monocyte-macrophage cells. Exposure to lipopolysaccharide (LPS) and ATP additively enhanced the activation of caspase-1 by ORF9b, suggesting that ORF9b and LPS follow parallel pathways in the activation of the inflammasome and caspase-1. Following rational in silico approaches, we have designed small molecules capable of inhibiting the homodimerization of ORF9b, which experimentally inhibited ORF9b-ORF9b homotypic interactions, caused mitochondrial eviction of ORF9b, inhibited ORF9b-induced activation of caspase-1 in A549 and THP-1 cells, cytokine release in THP-1 cells, and restored type I interferon (IFN-I) signaling suppressed by ORF9b in both cell models. These small molecules are first-in-class compounds targeting a viral accessory protein critical for viral-induced exacerbated inflammation and escape from innate immune responses, with the potential of mitigating the severe immunopathogenic damage induced by highly pathogenic coronaviruses and restoring antiviral innate immune responses curtailed by viral infection.

microbiology↗

bioRxiv2022

Microglial TNFα controls GABAAR plasticity, slow waves and memory consolidation during sleep

Microglia sense the changes in their environment. How microglia actively translate these changes into suitable cues to adapt brain physiology is unknown. We reveal an activity-dependent regulation of cortical inhibitory synapses by microglia, driven by purinergic signaling acting on P2RX7 and mediated by microglia-derived TNF. We demonstrate that sleep induces microglia-dependent synaptic enrichment of GABAARs in a manner dependent on microglial TNF and P2RX7. We further show that microglia-specific depletion of TNF alters slow waves during NREM sleep and blunt memory consolidation in sleep-dependent learning tasks. Together, our results reveal that microglia orchestrate sleep-intrinsic plasticity of synaptic GABAARs, sculpt sleep slow waves and support memory consolidation.

neuroscience↗

bioRxiv2022

Knock-out of CmNAC-NOR affects melon climacteric fruit ripening

Fruit ripening is an important process that affects fruit quality. In melon, ETHQV6.3, a QTL involved in climacteric ripening regulation, was previously found to be encoded by CmNAC-NOR, a homologue of the tomato NOR gene. To further investigate CmNAC-NOR function we have obtained two CRISPR/Cas9 mediated mutants (nor-3 and nor-1) in the climacteric Vedrantais background. nor-3, containing a 3-bp deletion altering the NAC domain A, resulted in ~8 days delay of ripening without affecting fruit quality. In contrast, nor-1 contained a 1-bp deletion resulting in a fully disrupted NAC domain, which completely blocked climacteric ripening. nor-1 fruits did not produce ethylene, abscission layer was not formed and there was no external color change. Additionally, volatile components were dramatically altered, seeds were not well developed and flesh firmness was also altered. nor-1 allele in heterozygosis showed ~20 days delay of fruit ripening. Our results provide new information regarding the function of CmNAC-NOR in melon fruit ripening, suggesting that it as a potential target to modulate shelf life in climacteric melon commercial varieties.

plant biology↗

bioRxiv2022

CRISPR/Cas9 gene editing uncovers the role of CTR1 and ROS1 in melon fruit ripening and epigenetic regulation

Melon (Cucumis melo L.) has emerged as an alternative model to study fruit ripening due to the coexistence of climacteric and non-climacteric varieties. The previous characterization of a major QTL ETHQV8.1 sufficient to trigger climacteric ripening in a non-climacteric background allowed the identification within the QTL interval of a negative regulator of ripening CmCTR1-like (MELO3C024518), and a putative DNA demethylase CmROS1 (MELO3C024516), the orthologue of DML2, a DNA demethylase regulating fruit ripening in tomato. To understand the role of these genes in climacteric ripening, we generated homozygous CRISPR knockout mutants of CmCTR1-like and CmROS1 in a climacteric genetic background. The climacteric behavior was altered in both loss-of-function mutants in two summer seasons with an advanced ethylene production profile compared to the climacteric wild type, suggesting a role of both genes in climacteric ripening in melon. Single cytosine methylome analyses of the CmROS1 knockout mutant revealed DNA methylation changes in the promoter regions of key ripening genes as ACS1, ETR1 and ACO1, and ripening associated-transcription factors as NAC-NOR, RIN and CNR, suggesting the importance of CmROS1-mediated DNA demethylation for triggering fruit ripening in melon.

plant biology↗

bioRxiv2020

GUN1-independent retrograde signaling targets the ethylene pathway to repress photomorphogenesis

When germinating in the light, Arabidopsis seedlings undergo photomorphogenic development, characterized by short hypocotyls, greening and expanded cotyledons. Stressed chloroplasts emit retrograde signals to the nucleus that induce developmental responses and repress photomorphogenesis. The nuclear targets of these retrograde signals are not yet fully known. Here, we show that lincomycin-treated seedlings (which lack developed chloroplasts) show strong phenotypic similarities to seedlings treated with ethylene (ET) precursor 1-aminocyclopropane-1-carboxylic acid (ACC), as both signals inhibit cotyledon separation in the light. We show that the lincomycin-induced phenotype partly requires a functioning ET signaling pathway, but could not detect increased ET emissions in response to lincomycin treatment. The two treatments show overlap in up-regulated gene transcripts, downstream of transcription factors ETHYLENE INSENSITIVE3 (EIN3) and EIN3-LIKE1 (EIL1). The induction of the ethylene signaling pathway is triggered by an unknown retrograde signal acting independently of GENOMES UNCOUPLED1 (GUN1). Our data show how two apparently different stress responses converge to optimize photomorphogenesis. One Sentence SummaryChloroplast retrograde signaling targets the ethylene-regulated gene network to repress photomorphogenesis in Arabidopsis

plant biology↗

bioRxiv2020

Peripheral Natural Killer cells from chronic hepatitis B patients display molecular hallmarks of T cell exhaustion

A significant proportion of individuals infected by HBV develops chronic infection. Antiviral effectors such as Natural Killer (NK) cells have impaired functions in these patients, but the molecular mechanism responsible for this dysfunction remains poorly characterized. Here, we show that peripheral NK cells from chronic hepatitis B (CHB) patients have a defective capacity to produce IFN-{gamma}, MIP1-{beta} and TNF- but retain an intact killing capacity. This functional phenotype was associated with a decrease in the expression of NKp30 and CD16, combined with defects in IL-15 stimulation of the mTOR pathway. Transcriptome analysis of NK cells in CHB patients further revealed a strong enrichment for transcripts typically expressed in exhausted T cells suggesting that NK cell dysfunction and T cell exhaustion rely on common molecular mechanisms. In particular, the transcription factor thymocyte selection-associated HMG box protein (TOX) and several of its targets, including immune checkpoints, were over-expressed in NK cells of CHB patients. This T cell exhaustion signature was predicted to be dependent on the calcium (Ca2+)-associated transcription factor NFAT. In line with this, when stimulating the Ca2+-dependent pathway in isolation, we recapitulated the dysfunctional phenotype. Thus, deregulated Ca2+ signalling could be a central event in both T cell exhaustion and NK cell dysfunction that occur during chronic infections.

immunology↗