Biology subjects

Guzman, A.

Publications and source records attributed to Guzman, A..

12 recordsLinked to original sources

bioRxiv2026

Negative affective states are not detected in rats following an intravenous self-administration regimen leading to incubation of oxycodone craving

In rats, cue-induced opioid craving intensifies (incubates) during abstinence from opioid self-administration and then remains high for a prolonged period. The prolonged plateau models persistent vulnerability to cue-induced craving and relapse in humans recovering from opioid use disorder. However, a very significant contributor to relapse vulnerability in these individuals is the presence of negative affective states that can persist for months to years, far beyond physical dependence. The goal of this study was to determine if the incubation of craving model recapitulates this aspect of relapse vulnerability. We began by comparing rats trained to self-administer oxycodone using a regimen leading to persistent elevation of cue-induced craving (6 h/d x 10 d) and rats trained to self-administer saline. We assessed somatic withdrawal signs in early abstinence and conducted behavioral tests modeling negative affect (open field, social preference, sucrose preference, and elevated plus maze) in late abstinence. Some somatic withdrawal signs were greater in oxycodone rats on abstinence day (AD)1, but cumulative scores did not differ between groups on AD1-3. On AD41-46, no group differences were found in behavioral tests modeling negative affect. To compare early and late abstinenceperiods, a second cohort of rats self-administered saline and oxycodoneand then received two cue-induced seeking tests (AD1 and AD40; oxycodone rats exhibited incubation of craving) and two series of negative affect tests (AD2-7 and AD41-48). While some time-dependent changes in affect were observed within each group, they were suggestive of reduced anxiety-like behavior in oxycodone rats. Finally, because rats are single-housed during our incubation studies, we compared drug-naive rats after 8-9 weeks of single vs pair housing and found no difference in behavioral tests modeling negative affect. We conclude that the persistence of elevated cue-induced craving observed after a standard opioid incubation regimen is not accompanied by negative affective states, probably due to lower drug intake during the intravenous regimen compared to non-contingent escalating dose regimens typically used to study withdrawal signs. This does not negate the utility of the incubation model for studying cue-induced opioid craving and its neurobiological basis.

neuroscience↗

bioRxiv2025

Higher glucose levels buffer against everyday stress load

Adaptive stress responses are dependent on the availability of energy and the bodys effectiveness in metabolizing glucose as fuel. However, it is not well understood if glucose levels contribute to the functional regulation of daily stress in healthy individuals. Here, we used 4,307 ratings of stress during a four-week period of ecological momentary assessment (EMA) with concurrent recordings from continuous glucose monitoring (CGM) in 91 participants (47 women). In accordance with the hypothesized metabolic link, we found that higher glucose levels were associated with lower self-reported stress (b = -.038, p = .035, controlling for mood). Crucially, dynamic changes in glucose levels also precipitated subsequent increases in stress levels (0-2h before EMA: p = .013; incremental effect 10-12h before: p < .001), with lower glucose levels associated with subsequent stress increases and elevated glucose with stress relief, highlighting the potential of monitoring physiological biosensor data to improve adaptive stress regulation. We conclude that metabolic signals, such as glucose, play an important role in channeling stress responsiveness in healthy adults, pointing to a pivotal avenue for improved interventions to avoid excessive stress experiences.

neuroscience↗

bioRxiv2025

An integrated workflow for structural virology with a 100 keV electron microscope

Cryo-EM has revolutionized structural biology, especially for flexible and heterogeneous samples, although access to high end microscopes that enable these studies remains a bottleneck. While 300 keV microscopes have been the go-to for high-resolution structural determination, they are expensive and restricted to institutional and national facilities needing specialized expertise, with access falling far short of the demand. Here, we present the user-managed operation of a cheaper 100 keV electron microscope within a structural biology laboratory enabling close integration with protein production, biochemical and biophysical studies. We provide details and considerations for the installation of the microscope, its day-to-day maintenance, and operations. Using virus surface glycoproteins as case studies, we illustrate the workflow from grid screening, data collection, and data processing, and provide examples of data quality. This user-administered setup provides a training platform for researchers at all levels, with beginners in cryo-EM achieving proficiency to independently operate the microscope within a month of regular use and training. We have demonstrated routine high-quality low-resolution reconstructions using a Ceta CMOS camera and high-resolution reconstructions enabling building of atomic models using a Falcon C direct detector. While there are several examples of facilities that manage cryo-EM and individual laboratories leveraging cryo-EM, we provide here the first demonstration of a modern group independently doing both successfully, something that has been talked about frequently but rarely seen.

biochemistry↗

bioRxiv2025

Nuclear Phase Separation Drives NPM1-mutant Acute Myeloid Leukemia

During cancer development, mutations promote gene expression changes that cause transformation. Leukemia is frequently associated with aberrant HOXA expression driven by translocations in nucleoporin genes or KMT2A, and mutations in NPM1. How disparate mutations converge on this regulatory pathway is not understood. Here we demonstrate that mutant NPM1 (NPM1c) forms nuclear condensates in multiple human cell lines, mouse models, and primary patient samples. We show NPM1c phase separation is necessary and sufficient to coordinate the recruitment of NUP98 and KMT2A to condensates. Through extensive mutagenesis and pharmacological destabilization of phase separation, we find that NPM1c condensates are necessary for regulating gene expression, promoting in vivo expansion, and maintaining the undifferentiated leukemic state. Finally, we show that nucleoporin and KMT2A fusion proteins form condensates that are biophysically indistinguishable from NPM1c condensates. Together, these data define a new condensate underlying leukemias that we term coordinating bodies (C-bodies), and propose C-bodies as a therapeutic vulnerability.

biophysics↗

bioRxiv2024

Mutant p53 regulates cancer cell invasion in complex three-dimensional environments through mevalonate pathway-dependent Rho/ROCK signaling.

Certain mutations can confer neomorphic gain of function (GOF) activities to the p53 protein that affect cancer progression. Yet the concept of mutant p53 GOF has been challenged. Here, using various strategies to alter the status of mutant versions of p53 in different cell lines, we demonstrate that mutant p53 stimulates cancer cell invasion in three-dimensional environments. Mechanistically, mutant p53 enhances RhoA/ROCK-dependent cell contractility and cell-mediated extracellular matrix (ECM) re-organization via increasing mevalonate pathway-dependent RhoA localization to the membrane. In line with this, RhoA-dependent pro-invasive activity is also mediated by IDI-1, a mevalonate pathway product. Further, the invasion-enhancing effect of mutant p53 is dictated by the biomechanical properties of the surrounding ECM, thereby adding a cell-independent layer of regulation to mutant p53 GOF activity that is mediated by dynamic reciprocal cell-ECM interactions. Together our findings link mutant p53 metabolic GOF activity with an invasive cellular phenotype in physiologically relevant and context-dependent settings. SignificanceThis study addresses the contribution of mutant p53 to the process of cancer cell dissemination in physiologically relevant three-dimensional environments - a key characteristic of metastatic disease. Several mutant p53 proteins display pro-oncogenic activity with respect to cancer cell invasion in 3D environments via mevalonate pathway-dependent Rho/ROCK signaling axis.

cancer biology↗

bioRxiv2024

Clonal dynamics and somatic evolution of haematopoiesis in mouse

Haematopoietic stem cells maintain blood production throughout life. While extensively characterised using the laboratory mouse, little is known about how the population is sustained and evolves with age. We isolated stem cells and progenitors from young and old mice, identifying 221,890 somatic mutations genome-wide in 1845 single cell-derived colonies, and used phylogenetic analysis to infer the ontogeny and population dynamics of the stem cell pool. Mouse stem cells and progenitors accrue [~]45 somatic mutations per year, a rate only about 2-fold greater than human progenitors despite the vastly different organismal sizes and lifespans. Phylogenetic patterns reveal that stem and multipotent progenitor cell pools are both established during embryogenesis, after which they independently self-renew in parallel over life. The stem cell pool grows steadily over the mouse lifespan to approximately 70,000 cells, self-renewing about every six weeks. Aged mice did not display the profound loss of stem cell clonal diversity characteristic of human haematopoietic ageing. However, targeted sequencing revealed small, expanded clones in the context of murine ageing, which were larger and more numerous following haematological perturbations and exhibited a selection landscape similar to humans. Our data illustrate both conserved features of population dynamics of blood and distinct patterns of age-associated somatic evolution in the short-lived mouse.

genomics↗

bioRxiv2024

Evolution and Diversification of the Aposematic Poison Frog, Oophaga pumilio, in Bocas del Toro

The aposematic strawberry poison frog, Oophaga pumilio, is an iconic model system for studying the evolution and maintenance of color variation. Through most of its range, this frog is red with blue limbs. However, frogs from the Bocas del Toro Province, Panama, show striking variance in color and pattern, both sympatrically and allopatrically. This observation contradicts standard models of the evolution of aposematism and has led to substantial speculation about its evolutionary and molecular causes. Since the enigma of O. pumilio phenotypic variation is partly unresolved because of its large, [~] 6.7 Gb genome, we here sequence exomes from 347 individuals from ten populations and map a number of genetic factors responsible for the color and pattern variation. The kit gene is the primary candidate underlying the blue-red polymorphism in Dolphin Bay, where an increase in melanosomes is correlated with blue coloration. Additionally, the ttc39b gene, a known enhancer of yellow-to-red carotenoid conversion in birds, is the primary factor behind the yellow-red polymorphism in the Bastimentos West area. The causal genetic regions show evidence of selective sweeps acting locally to spread the rare phenotype. Our analyses suggest an evolutionary model in which selection is driving the formation of new morphs in a dynamic system resulting from a trade-off between predation avoidance, intraspecific competition, and mate choice.

evolutionary biology↗

bioRxiv2023

Sprouty4 is required for Mdm2 regulation of invasion, focal adhesion formation and metastasis in cells lacking p53.

Although the E3 ligase Mdm2 and its homologue and binding partner MdmX are the major regulators of the p53 tumor suppressor protein, it is now evident that Mdm2 and MdmX have multiple functions that are independent of p53. For example, Mdm2 can regulate cell migration, although mechanistic insight into this function is still lacking. Here we show in cells lacking p53 expression that knockdown of Mdm2 or MdmX, as well as pharmacological inhibition of the Mdm2/MdmX complex, not only reduces cell migration and invasion, but also impairs cell spreading and focal adhesion formation. In addition, Mdm2 knockdown decreases metastasis in vivo. Remarkably, Mdm2 modulates the expression of Sprouty4, which is required for the Mdm2 mediated effects on cell migration, focal adhesion formation and metastasis. Our findings describe a molecular mechanism by which the Mdm2-X complex, through Sprouty4, regulates cellular processes leading to decreased metastatic capability independent of p53.

cancer biology↗

bioRxiv2021

Water Stress and Disruption of Mycorrhizae Induce Parallel Shifts in Phyllosphere Microbiome Composition

Water and nutrient limitation are key stressors that affect plant health and ecosystem function. These environmental factors impact both soil- and root-associated microbial communities, and systemically alter plant physiology, but it is less clear how they affect aboveground plant-microbe interactions. Through experimental manipulations in the field and growth chamber, we examined the interacting effects of irrigation, soil fertility, and root mycorrhizal associations on bacterial and fungal communities of the tomato phyllosphere (Solanum lycopersicum). Both water stress and mycorrhizal disruption reduced bacterial richness within plants, homogenized bacterial community diversity among plants, and reduced the relative abundance of dominant fungal taxa. We observed striking parallelism in the individual microbial taxa affected by irrigation and mycorrhizal associations. Given the increasingly understood role of the phyllosphere in shaping plant health and pathogen susceptibility, these results offer an additional mechanism by which belowground conditions shape plant fitness.

ecology↗

bioRxiv2020

Oncometabolism Drives Autophagy Activation in Skeletal Muscle

About 20-30% of cancer-associated deaths are due to complications from cachexia which is characterized by skeletal muscle atrophy. Metabolic reprogramming in cancer cells causes body-wide metabolic and proteomic remodeling, which remain poorly understood. Here, we present evidence that the oncometabolite D-2-hydroxylgutarate (D2-HG) impairs NAD+ redox homeostasis in skeletal myotubes, causing atrophy via deacetylation of LC3-II by the nuclear deacetylase Sirt1. Overexpression of p300 or silencing of Sirt1 abrogate its interaction with LC3, and subsequently reduced levels of LC3 lipidation. Using RNA-sequencing and mass spectrometry-based metabolomics and proteomics, we demonstrate that prolonged treatment with the oncometabolite D2-HG in mice promotes cachexia in vivo and increases the abundance of proteins and metabolites, which are involved in energy substrate metabolism, chromatin acetylation and autophagy regulation. We further show that D2-HG promotes a sex-dependent adaptation in skeletal muscle using network modeling and machine learning algorithms. Our multi-omics approach exposes new metabolic vulnerabilities in response to D2-HG in skeletal muscle and provides a conceptual framework for identifying therapeutic targets in cachexia.

systems biology↗

bioRxiv2020

Modeling IKZF1 lesions in B-ALL reveals distinct chemosensitivity patterns and potential therapeutic vulnerabilities

IKAROS family zinc finger 1 (IKZF1) alterations represent a diverse group of genetic lesions that are associated with an increased risk of relapse in B-lymphoblastic leukemia (B-ALL). Due to the heterogeneity of concomitant lesions it remains unclear how IKZF1 abnormalities directly affect cell function and therapy resistance and whether their consideration as a prognostic indicator is valuable in improving outcome. We used CRISPR/Cas9 to engineer multiple panels of isogeneic lymphoid leukemia cell lines with a spectrum of IKZF1 lesions in order to measure changes in chemosensitivity, gene expression, cell cycle, and in vivo engraftment dynamics that can be directly linked to loss of IKAROS protein. IKZF1 knockout and heterozygous null cells displayed relative resistance to a number of commonly employed therapies for B-ALL including dexamethasone, vincristine, asparaginase, and daunorubicin. Transcription profiling revealed a stem/myeloid cell-like phenotype and JAK/STAT upregulation after IKAROS loss. We also used a CRISPR homology-directed repair (HDR) strategy to knock-in the dominant-negative IK6 isoform tagged with GFP into the endogenous locus and observed a similar drug resistance profile with the exception of retained sensitivity to dexamethasone. Interestingly, IKZF1 knockout and IK6 knock-in cells both have significantly increased sensitivity to cytarabine, suggesting intensification of nucleoside analog therapy may be specifically effective for IKZF1-deleted B-ALL. Both types of IKZF1 lesions decreased survival time of xenograft mice, with higher numbers of circulating blasts and increased organ infiltration. Given these findings, exact specification of IKZF1 status in patients may be a beneficial addition to risk stratification and could inform therapy. Key pointsO_LIEngineered IKZF1 perturbations result in a stem-cell like expression signature, enhanced engraftment in vivo, and multi-drug resistance C_LIO_LILoss of IKAROS may result in new vulnerabilities due to increased sensitivity to cytarabine and upregulation of JAK/STAT and mAb targets C_LI

cancer biology↗

bioRxiv2019

A Systems Biology perspective of Dynamical Patterning Modules in the transition to multicellularity: lessons from an aggregative bacteria

Development and evolution are dynamical processes under the continuous control of organismic and environmental factors. Generic physical processes, associated with biological materials and certain genes or molecules, provide a morphological template for the evolution and development of organism forms. Generic dynamical behaviors, associated with recurring network motifs, provide a temporal template for the regulation and coordination of biological processes. The role of generic physical processes and their associated molecules in development is the topic of the Dynamical Patterning Module (DPM) framework. The role of generic dynamical behaviors in biological regulation is studied via the identification of the associated Network Motifs (NM). We propose a joint DPM-NM perspective on the emergence and regulation of multicellularity focusing on a multicellular aggregative bacterium, Myxococcus xanthus. Understanding M. xanthus development as a dynamical process embedded in a physical substrate provides novel insights into the interaction between developmental regulatory networks and generic physical processes in the evolutionary transition to multicellularity.

systems biology↗