Biology subjects

Caza, M.

Publications and source records attributed to Caza, M..

6 recordsLinked to original sources

bioRxiv2026

Chronic oral fentanyl consumption produces dynamic behavioral adaptations during abstinence in female mice

RationaleThe opioid epidemic continues to be driven by synthetic opioids, particularly fentanyl, yet the long-term behavioral manifestations of withdrawal remain poorly characterized. Women exhibit unique vulnerabilities to opioid use disorder, including greater susceptibility to withdrawal-related symptoms and relapse, suggesting that defining the behavioral adaptations that emerge during prolonged fentanyl abstinence may identify mechanisms underlying female-specific relapse vulnerability. Thus, we used an oral fentanyl drinking model to longitudinally examine nociceptive, affective, and exploratory behaviors across chronic exposure and abstinence in female mice. MethodsAdult female C57BL/6J mice underwent a modified 5-week Drinking-in-the-Dark paradigm with 4-hour daily access to fentanyl (30 {micro}g/mL) or water. Somatic withdrawal was assessed following naloxone-precipitated withdrawal. Thermal and mechanical nociception, sucrose preference, exploratory behavior, acoustic startle, nest building, and locomotor activity were evaluated during fentanyl exposure and throughout one month of abstinence. ResultsFemale mice exhibited robust naloxone-precipitated somatic withdrawal, confirming physiological effects of opioid drinking. Chronic fentanyl exposure produced transient thermal hyperalgesia during weeks 2-4 of drinking that resolved by the final week, but re-emerged after 30 days of abstinence. After 30 days of abstinence, we also found increased mechanical sensitivity. During early abstinence, fentanyl-exposed mice exhibited increased sucrose consumption and greater exploration of anxiogenic environments without alterations in general locomotion. Increased exploratory behavior persisted into prolonged abstinence and was partially rescued in mice who previously received naloxone. Additionally, fDID mice exhibited impaired nesting behavior, while prior naloxone-precipitated withdrawal improved nest-building performance. ConclusionsWe demonstrate that chronic opioid exposure, precipitated withdrawal trials, and duration of abstinence interact to shape protracted withdrawal phenotypes in female mice that extend well beyond drug exposure. These results provide insight into persistent withdrawal symptoms that may shape relapse vulnerability using a novel translationally relevant framework.

neuroscience↗

bioRxiv2026

AI-Driven Discovery and BSL-4 Validation of Cross-Filovirus Ebola-Marburg Inhibitors and their Synergistic Combinations

Filovirus outbreaks caused by Ebola virus (EBOV) and Marburg virus (MARV), pose severe global health threats characterized by high rates of fatal hemorrhagic fever. While species-specific vaccines and therapeutic monoclonal antibodies are approved for Zaire ebolavirus, broadly-active therapeutics remain unavailable, leaving populations vulnerable to MARV and other pathogenic Ebola species, such as Bundibugyo (BDBV) and Sudan (SUDV) ebolaviruses. Here we report a computationally guided, infectious virus validated screening platform for the rapid discovery of broad-spectrum filovirus antivirals. By leveraging quantitative structure-activity relationship (QSAR) models, we screened 142,382 compounds in silico to prioritize 125 high-potential candidates. Subsequent dose-response and viability profiling identified 23 compounds exhibiting potent, low-micromolar pan-filovirus activity and favorable cytotoxicity profiles. Molecular docking indicates these compounds target conserved structural and functional domains--primarily the VP35 and L proteins--which may disrupt essential viral replication and immune antagonism. Furthermore, systematic combinatorial screening revealed three highly synergistic compound pairs, notably NCGC00113249-01 and NCGC00118008-01, demonstrating robust cross-species efficacy. By targeting conserved vulnerabilities across the filovirus family, this integrated in silico and in vitro pipeline provides a scalable framework to rapidly nominate and optimize synergistic therapeutic regimens against both endemic and emerging viral threats including BDBV. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/737586v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@531939org.highwire.dtl.DTLVardef@19d1454org.highwire.dtl.DTLVardef@1a1c9e3org.highwire.dtl.DTLVardef@cd861b_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2024

Glutathione metabolism impacts fungal virulence by modulating the redox environment

Pathogens must overcome the hostile conditions of their hosts to survive, proliferate and cause disease. The fungal pathogen Cryptococcus neoformans is particularly adept at mitigating challenges in the host environment and has developed an arsenal of defense mechanisms to evade oxidative and nitrosative agents released by phagocytic cells during infection. Among these mechanisms, melanin production is crucially linked to both fungal virulence and defense against harmful free radicals that facilitate host innate immunity and clearance of invading pathogens. Here, we employed comparative global metabolomics to demonstrate that metabolism of the antioxidant glutathione (GSH) is inextricably linked to redox-active processes that facilitate melanin production, and that genetic perturbations in GSH biosynthesis affect fungal growth and virulence in a murine model of cryptococcosis. Furthermore, we show that disruption of GSH biosynthesis leads to overaccumulation of reducing and acidic compounds in the extracellular environment of mutant cells. These changes not only impacted melanin formation but also influenced titan cell and urease production as well as survival in macrophages. Overall, these findings highlight the importance of redox homeostasis and metabolic compensation in pathogen adaptation to the host environment and suggest new avenues for antifungal drug development.

microbiology↗

bioRxiv2018

The monothiol glutaredoxin Grx4 interacts with the Cryptococcus iron regulator Cir1 and regulates iron homeostasis and virulence in the Cryptococcus neoformans

The acquisition of iron and the maintenance of iron homeostasis are important aspects of the virulence in the pathogenic fungus Cryptococcus neoformans. In this study, we identified the monothiol glutaredoxin Grx4 as a binding partner of Cir1, a master regulator of iron-responsive genes and virulence factor elaboration in C. neoformans. Monothiol glutaredoxins are important regulators of iron homeostasis because of their conserved roles in [2Fe-2S] cluster sensing and trafficking. We confirmed that Grx4 binds Cir1 and demonstrated that iron repletion promotes the relocalization of Grx4 from the nucleus to the cytoplasm. Nuclear retention is partially dependent on Cir1 and also influenced by treatment with the proteasome inhibitor bortezomib. Cir1 remains in the nucleus in both iron replete and iron limiting conditions. We also found that a grx4{Delta} mutant displayed iron-related phenotypes similar to those of a cir1{Delta} mutant, including poor growth upon iron deprivation. Importantly, a grx4{Delta} mutant was avirulent in mice, a phenotype consistent with observed defects in the key virulence determinants, capsule and melanin, and poor growth at 37{degrees}C. A comparative transcriptome analysis of a grx4{Delta} mutant and the WT strain in low iron and iron-replete conditions confirmed a central role for Grx4 in iron homeostasis. Dysregulation of iron-related metabolism was consistent with grx4{Delta} mutant phenotypes related to oxidative stress, mitochondrial function, and DNA repair. Overall, the phenotypes of the grx4{Delta} mutant and the RNA-Seq analysis support the hypothesis that Grx4 functions as a sensor of iron levels, in part through an interaction with Cir1, to extensively regulate iron homeostasis and contribute to virulence.

microbiology↗

bioRxiv2018

The Sec1/Munc18 (SM) protein Vps45 is involved in iron uptake, mitochondrial function and virulence in the pathogenic fungus Cryptococcus neoformans

The battle for iron between invading microorganisms and mammalian hosts is a pivotal determinant of the outcome of infection. The pathogenic fungus, Cryptococcus neoformans, employs multiple mechanisms to compete for iron during cryptococcosis, a disease primarily of immunocompromised hosts. In this study, we examined the role of endocytic trafficking in iron uptake by characterizing a mutant defective in the Sec1/Munc18 (SM) protein Vps45. This protein is known to regulate the machinery for vesicle trafficking and fusion via interactions with SNARE proteins. As expected, a vps45 deletion mutant was impaired in endocytosis and showed sensitivity to trafficking inhibitors. The mutant also showed poor growth on iron-limited media and a defect in transporting the Cfo1 ferroxidase of the high-affinity iron uptake system from the plasma membrane to the vacuole. Remarkably, we made the novel observation that Vps45 also contributes to mitochondrial function in that a Vps45-Gfp fusion protein associated with mitotracker, and a vps45 mutant showed enhanced sensitivity to inhibitors of electron transport complexes as well as changes in mitochondrial membrane potential. Consistent with mitochondrial function, the vps45 mutant was impaired in calcium homeostasis. To assess the relevance of these defects for virulence, we examined cell surface properties of the vps45 mutant and found increased sensitivity to agents that challenge cell wall integrity and antifungal drugs. A change in cell wall properties was consistent with our observation of altered capsule polysaccharide attachment, and with attenuated virulence in a mouse model of cryptococcosis. Overall, our studies reveal a novel role for Vps45-mediated trafficking for iron uptake, mitochondrial function and virulence.

microbiology↗

bioRxiv2018

Role of clathrin-mediated endocytosis in the use of heme and hemoglobin by the fungal pathogen Cryptococcus neoformans

Heme is a major source of iron for pathogens of humans, and its use is critical in determining the outcome of infection and disease. Cryptococcus neoformans is an encapsulated fungal pathogen that causes life-threatening infections in immunocompromised individuals. C. neoformans effectively uses heme as an iron source but the underlying mechanisms are poorly defined. Non-iron metalloporphyrins (MPPs) are toxic analogues of heme and are thought to enter microbial cells via endogenous heme acquisition systems. We therefore carried out a mutant screen for susceptibility against manganese metalloporphyrin (Mn MPP) to identify new components for heme uptake in C. neoformans. We identified several genes involved in signaling, DNA repair, sugar metabolism and trafficking that play important roles in susceptibility to Mn MPP and in the use of heme as an iron source. We focused on investigating the role of clathrin-mediated endocytosis (CME) and found that several components of CME including Chc1, Las17, Rvs161 and Rvs167 are required for growth on heme and hemoglobin, and for endocytosis and intracellular trafficking of these molecules. We show that the hemoglobin uptake process in C. neoformans involves clathrin heavy chain, Chc1, which appears to co-localize with hemoglobin containing-vesicles and to potentially assist in proper delivery of hemoglobin to the vacuole. Additionally, C. neoformans strains lacking Chc1, Las17, Rvs161, or Rvs167 were defective in the elaboration of several key virulence factors and a las17 mutant was avirulent in a mouse model of cryptococcosis. Overall, this study unveils crucial functions of CME in the use of heme iron by C. neoformans and reveals a role for CME in fungal pathogenesis.

microbiology↗