Biology subjects

Bravo, P.

Publications and source records attributed to Bravo, P..

8 recordsLinked to original sources

bioRxiv2026

Single-cell profiling reveals disruption of cell-cycle homeostasis following JDP2 depletion in colorectal epithelial cells

Intestinal epithelial cells (IECs) undergo rapid and continuous renewal to maintain gut epithelial homeostasis and barrier integrity at the interface with luminal microbiota, dietary antigens, and enteric pathogens. IEC proliferation is tightly regulated by a complex machinery of cell cycle regulators and effectors, and genetic or transcriptional dysregulation of cell-cycle regulatory pathways is a hallmark of colorectal cancer development and progression. Previously, JDP2 (Jun dimerization protein 2) has been shown to be involved in cell cycle control and is associated with an array of cancers. JDP2 functions as a transcription factor or epigenetic regulator depending on cellular context, disease, and cancer types. However, the role of JDP2 in CRC, cell cycle homeostasis of colon cancer cells, and their fate remains poorly defined. In this study, we found that patients with higher JDP2 expression have significantly worse survival than patients with lower JDP2 expression. Our functional assays showed that JDP2 depletion increased EdU incorporation during S phase and PHH3 abundance, which is consistent with increased proliferative and mitotic activity. Our single-cell RNA sequencing and transcriptomic profiling further revealed reprogramming of cell-cycle-associated transcriptional states following JDP2 depletion. Differential-expression and pathway analysis identified changes in mitotic and cell-cycle regulatory genes, including programs involving cyclins, cyclin-dependent kinases, chromosome segregation, and mitotic progression. We also found that lower residual JDP2 expression was associated with escalated G2/M representation and a change toward later states along a G1 - S - G2/M-associated transcriptional trajectory. Furthermore, low-JDP2 cells showed significantly greater late-state occupancy than High-JDP2 cells (33.6% versus 24.0%; paired P=0.048). Among the JDP2-siRNA-treated cells, analysis of residual JDP2-expressing cells also revealed heterogeneous associations of JDP2 with cell-cycle, survival, apoptotic, and stress-response programs, consistent with JDP2's context-dependent transcriptional functions. Together, these data identify JDP2 as an important component of intestinal epithelial cell-cycle homeostasis and transcriptional landscape and suggest that JDP2 depletion perturbs the balance of proliferative cell states.

molecular biology↗

bioRxiv2026

Campylobacter jejuni Infection Is Associated With Cell-Cycle, Redox, and Metabolic Remodeling of Human Intestinal Epithelial Cells at Single-Cell Resolution

Campylobacter jejuni is the leading cause of bacterial enteric infections worldwide, including in the US. It is also a zoonotic pathogen that is transmitted by food and water, causing diarrhea and intestinal inflammation, and is responsible for high morbidity and mortality in young children, the elderly, and immunocompromised patients. During infection, C. jejuni profoundly perturbs intestinal epithelial physiology, leading to widespread mucosal damage and inflammatory response. However, the heterogeneity and coordination of host transcriptional responses remain incompletely defined. We performed single-cell RNA sequencing of Caco-2 intestinal epithelial cells from uninfected Control and C. jejuni-infected biological samples using 10x Genomics GEM-X Flex chemistry. Our dataset comprised 159,088 cells and 18,142 measured features. A Seurat workflow resolved 19 transcriptional states and revealed a highly reproducible condition-associated redistribution of clusters across biological replicates. Our study demonstrated that C. jejuni infection was associated with increased representation of G2/M-classified cells and coordinated induction of checkpoint and mitotic-spindle genes, including CDKN1A, WEE1, MAD2L1, BUB1B, PLK1, CDC20, CDK1, and UBE2C. Furthermore, ranked enrichment independently identified a C. jejuni-enriched Reactome mitotic spindle checkpoint program (NES ~1.67, FDR ~0.013), while Hallmark analysis identified strong TNF/NF-kB, hypoxia, G2M checkpoint, TGF-beta, glycolysis, apoptosis, p53, and mTORC1-associated programs. Infection was also associated with significantly decreased JDP2 and higher NOX1 and DUOX2 gene expression. Furthermore, scMetabolism/AUCell identified 20 KEGG metabolic pathways at FDR <0.05 in a balanced 10,000-cell analysis, with 12 lower and 8 higher in infection. Together, these data support a model in which C. jejuni dramatically restructures epithelial transcriptional states and is accompanied by mitotic-checkpoint, inflammatory/stress, redox, and metabolic remodeling.

microbiology↗

bioRxiv2025

Microglia and Chek2 contribute to sex-specific organization of the adult zebrafish brain

Sex specific differences in size and distribution of cell types have been observed in mammalian brains. How sex-specific differences in the brain are established and to what extent sexual dimorphism contributes to sex-biased neurodevelopment and neurological disorders is not well understood. Microglia are the resident immune cells of the nervous system and have been implicated in masculinizing the mammalian brain and refining neural connections to promote remodeling of neural circuitry, yet their contributions to developmental brain patterning and plasticity in zebrafish remains unclear. Here, we report anatomical and cellular differences between juvenile brains and adult female and male brains. Leveraging the plasticity of the zebrafish female brain and genetic models lacking microglia and tumor suppressor factors, we provide insight into the mechanisms that establish sex-specific brain dimorphism in zebrafish. Specifically, we identified sexually dimorphic features in the adult zebrafish brain that depend on microglia and Chek2, which may have broader implications and represent therapeutic targets for sex-biased neurological disorders. Plain language summaryMales and females of species can have significant differences in appearance, including differences in size, color, or sex specific anatomical structures. In addition to overt morphological differences, sex specific differences in size and distribution of cell types have been observed in mammalian brains. How these sex-specific differences in the brain are established and to what extent these differences contribute to sex-specific neurodevelopment and neurological disorders that differentially impact males and females is not well understood. Despite an incomplete picture of the mechanisms regulating sex-specific development, some of the cell types involved include microglia. Microglia are the resident immune cells of the nervous system and have been implicated in promoting features that are typical in the male mammalian brain. Specifically, microglia may refine neural connections and promote remodeling of neural circuitry and influence sex-specific behaviors. The contributions of microglia to developmental brain patterning and plasticity in zebrafish remain unclear. Here, we report anatomical and cellular differences between juvenile brains and adult female and male brains. Leveraging zebrafish genetic models lacking microglia and tumor suppressor factors, and the unique plasticity of the zebrafish female brain, we investigated and provide insight into the mechanisms that establish sex-specific brain differences in zebrafish. Specifically, we identified sexually distinct features in the adult zebrafish brain that depend on microglia and the tumor suppressor Chek2. If these or similar mechanisms operate in other species, our findings may have broader implications for sex-specific brain development and represent therapeutic targets for sex-biased neurological disorders. HighlightsO_LITissue clearing and immunostaining of juvenile and adult whole-mount zebrafish brains allows analysis of sex differences. C_LIO_LIAnatomical and cellular sexual dimorphism in the adult vertebrate brain appears after gonadal sex differentiation. C_LIO_LISexual dimorphism in the adult brain is driven by differences in cell death regulation. C_LIO_LIMicroglia colonization of brain areas involved in courtship is sexually dimorphic. C_LIO_LIMicroglia involvement in establishing sex-specific differences in the adult brain. C_LI

developmental biology↗

bioRxiv2024

Microbial Metabolite Trimethylamine N-Oxide Promotes Campylobacter jejuni Infection by Escalating Intestinal Inflammation, Epithelial Damage, and Barrier Disruption

The interactions between Campylobacter jejuni, a critical foodborne cause of gastroenteritis, and the intestinal microbiota during infection are not completely understood. The crosstalk between C. jejuni and its host is impacted by the gut microbiota through mechanisms of competitive exclusion, microbial metabolites, or immune response. To investigate the role of gut microbiota on C. jejuni pathogenesis, we examined campylobacteriosis in the IL10KO mouse model, which was characterized by an increase in the relative abundance of intestinal proteobacteria, E. coli, and inflammatory cytokines during C. jejuni infection. We also found a significantly increased abundance of microbial metabolite Trimethylamine N-Oxide (TMAO) in the colonic lumens of IL10KO mice. We further investigated the effects of TMAO on C. jejuni pathogenesis. We determined that C. jejuni senses TMAO as a chemoattractant and the administration of TMAO promotes C. jejuni invasion into Caco-2 monolayers. TMAO also increased the transmigration of C. jejuni across polarized monolayers of Caco-2 cells, decreased TEER, and increased C. jejuni-mediated intestinal barrier damage. Interestingly, TMAO treatment and presence during C. jejuni infection of Caco-2 cells synergistically caused an increased inflammatory cytokine expression, specifically IL-1{beta} and IL-8. These results establish that C. jejuni utilizes microbial metabolite TMAO for increased virulence during infection.

microbiology↗

bioRxiv2023

Examining the role of oxygen-binding proteins on the early evolution of multicellularity

Oxygen availability is a key factor in the evolution of multicellularity, as larger and more sophisticated organisms often require mechanisms allowing efficient oxygen delivery to their tissues. One such mechanism is the presence of oxygen-binding proteins, such as globins and hemerythrins, which arose in the ancestor of bilaterian animals. Despite their importance, the precise mechanisms by which oxygen-binding proteins influenced the early stages of multicellular evolution under varying environmental oxygen levels are not yet clear. We addressed this knowledge gap by heterologously expressing the oxygen binding proteins myoglobin and myohemerythrin in snowflake yeast, a model system of simple, undifferentiated multicellularity. These proteins increased the depth and rate of oxygen diffusion, increasing the fitness of snowflake yeast growing aerobically. Experiments show that, paradoxically, oxygen-binding proteins confer a greater fitness benefit for larger organisms under high, not low, O2 conditions. We show via biophysical modeling that this is because facilitated diffusion is more efficient when oxygen is abundant, transporting a greater quantity of O2 which can be used for metabolism. By alleviating anatomical diffusion limitations to oxygen consumption, the evolution of O2-binding proteins in the oxygen-rich Neoproterozoic may have been a key breakthrough enabling the evolution of increasingly large, complex multicellular metazoan lineages.

evolutionary biology↗

bioRxiv2023

Emergence and maintenance of stable coexistence during a long-term multicellular evolution experiment

The evolution of multicellular life spurred evolutionary radiations, fundamentally changing many of Earths ecosystems. Yet little is known about how early steps in the evolution of multicellularity transform eco-evolutionary dynamics, e.g., via niche expansion processes that may facilitate coexistence. Using long-term experimental evolution in the snowflake yeast model system, we show that the evolution of multicellularity drove niche partitioning and the adaptive divergence of two distinct, specialized lineages from a single multicellular ancestor. Over 715 daily transfers, snowflake yeast were subject to selection for rapid growth in rich media, followed by selection favoring larger group size. Both small and large cluster-forming lineages evolved from a monomorphic ancestor, coexisting for over ~4,300 generations. These small and large sized snowflake yeast lineages specialized on divergent aspects of a trade-off between growth rate and survival, mirroring predictions from ecological theory. Through modeling and experimentation, we demonstrate that coexistence is maintained by a trade-off between organismal size and competitiveness for dissolved oxygen. Taken together, this work shows how the evolution of a new level of biological individuality can rapidly drive adaptive diversification and the expansion of a nascent multicellular niche, one of the most historically-impactful emergent properties of this evolutionary transition.

evolutionary biology↗

bioRxiv2023

Macrophage activation drives ovarian failure and masculization

In humans, premature ovarian insufficiency (POI) is caused by autoimmunity and genetic factors, such as mutation of BMP15, a key ovarian determining gene. The cellular mechanisms associated with ovarian failure caused by BMP15 mutation and immune contributions to the disorder are not understood. BMP15s role in ovarian follicle development is conserved in vertebrates, including zebrafish. Using zebrafish, we established a causal link between macrophage activation and ovarian failure. We identified a germline-somatic gonadal cell-macrophage axis underlying ovarian atresia. Germline loss of Bmp15 triggers this axis that single-cell RNA sequencing and genetic analyses indicate involves activation of ovarian somatic cells that express conserved macrophage-activating ligands. Genetic ablation of macrophages blocks premature oocyte loss. Thus, the axis identified here represents potential therapeutic targets to preserve female fertility. One-Sentence SummarySex reversal due to Bmp15 deficiency requires macrophage activation by Csf1a, which is expressed by specialized pre-follicle cells in zebrafish.

developmental biology↗

bioRxiv2022

Vertical growth dynamics of biofilms

During the biofilm life cycle, bacteria attach to a surface then reproduce, forming crowded, growing communities. Many theoretical models of biofilm growth dynamics have been proposed; however, difficulties in measuring biofilm height accurately across relevant time and length scales have prevented testing these models or their biophysical underpinnings empirically. Using white light interferometry, we measure the heights of microbial colonies with nanometer precision from inoculation to their final equilibrium height, producing a novel and detailed empirical characterization of vertical growth dynamics. We show that models relying on logistic growth or nutrient depletion fail to capture biofilm height dynamics on short and long time scales. Our empirical results support a simple model inspired by the fact that biofilms only interact with the environment through their interfaces. This interface model captures the growth dynamics from short to long time scales (10 minutes to 14 days) of diverse microorganisms, including prokaryotes like gram-negative and gram-positive bacteria and eukaryotes like aerobic and anaerobic yeast. This model provides heuristic value, highlighting the biophysical constraints that limit vertical growth as well as establishing a quantitative model for biofilm development.

biophysics↗