Biology subjects

Fernandez-Antoran, D.

Publications and source records attributed to Fernandez-Antoran, D..

9 recordsLinked to original sources

bioRxiv2026

Human 3D epithelioids enable continuous long-term clonal evolution studies across multiple epithelial tissues

Modeling human epithelia in vitro remains challenging because current systems do not fully preserve the combination of architecture, heterogeneity, clonal composition, and long-term dynamics shown in vivo. While 3D approaches such as organoids and organotypic cultures capture important aspects of lineage differentiation and niche signaling, they often lose stable organization over time, limiting studies of long-lasting processes such as clonal evolution and cell competition. Here, we present human epithelioids as continuous long-term, 3D epithelial cultures efficiently derived from eight adult human epithelia, including trachea, skin, buccal mucosa, esophagus, blader, urethra, submandibular gland and endometrium. Using immunostaining, electron microscopy, single-cell RNA sequencing, functional assays and somatic mutation analyses, we deeply characterized human epithelioids and confirmed that they recapitulate native architecture and cell diversity, sustain regenerative capacity, and preserve donor-specific mutational landscapes, establishing a robust and versatile platform for longitudinal interrogation of clonal evolution, tissue dynamics and responses to clinically relevant perturbations, including radiotherapy and chemotherapy, over extended timescales.

cancer biology↗

bioRxiv2026

YBX3 overexpression in mesothelioma drives aberrant cell proliferation

Malignant pleural mesothelioma (MpM) is a lethal tumour closely linked to asbestos exposure and is a cancer of unmet clinical need with no known oncogenic drivers. Recent advancements in technologies to identify RNA-binding proteins (RBPs) has uncovered an emerging role for RBP-RNA interactions in cancer progression and we therefore assessed changes in the RBPome of patient-derived MpM cell lines. We identify over 350 RBPs showing altered RNA binding, with functions consistent with key cancer hallmarks, and discovered YBX3 as a potential oncoprotein driving cell proliferation in MpM. Mechanistically we show the impact of YBX3 on cell growth is achieved through its control of the expression of the amino acid transporter SLC7A5/LAT1, with increased amino acid uptake increasing protein synthesis rates. Notably, we show the inhibition of cell growth by YBX3 deletion is recapitulated by the clinically-relevant SLC7A5/LAT1 inhibitor JPH203. Finally, we demonstrate that JPH203 sensitizes MpM cells to radiotherapy, which could provide a promising therapeutic strategy for MpM. TeaserHigher levels of YBX3 expression in mesothelioma increases cell proliferation and protein synthesis rates by upregulation of amino acid uptake.

molecular biology↗

bioRxiv2023

Epithelioids: Self-sustaining 3D epithelial cultures to study long-term processes

Studying long-term biological processes such as the colonization of aging epithelia by somatic mutant clones has been slowed by the lack of suitable culture systems. Here we describe epithelioids, a facile, cost-effective method of culturing multiple mouse and human epithelia. Esophageal epithelioids self-maintain without passaging for at least a year, recapitulating the 3D structure, cell dynamics, transcriptome, and genomic stability of the esophagus. Live imaging over 5 months showed epithelioids replicate in vivo cell dynamics. Epithelioids enable the study of cell competition and mutant selection in 3D epithelia, and how anti-cancer treatments modulate the competition between transformed and wild type cells. Epithelioids are a novel method with a wide range of applications in epithelial tissues, particularly the study of long term processes, that cannot be accessed using other culture models.

cell biology↗

bioRxiv2023

Early infection response of the first trimester human placenta at single-cell scale

Placental infections are a major worldwide burden, particularly in developing countries. The placenta is a transient tissue located at the interface between the mother and the fetus. Some pathogens can access the placental barrier resulting in pathological transmission from mother to fetus, which may have a profound impact on the health of the developing fetus. Limited tissue accessibility, critical differences between humans and mice, and, until recently, lack of proper in vitro models, have hampered our understanding of the early placental response to pathogens. Here we use single-cell transcriptomics to describe the placental primary defence mechanisms against three pathogens that are known to cause fetal and maternal complications during pregnancy - Plasmodium falciparum, Listeria monocytogenes and Toxoplasma gondii. We optimise ex vivo placental explants of the first-trimester human placenta and show that trophoblasts (the epithelial-like cells of the placenta), and Hofbauer cells (placental macrophages) orchestrate a coordinated inflammatory response after 24 hours of infection. We show that hormone biosynthesis and transport are downregulated in the trophoblasts, suggesting that protective responses are promoted at the expense of decreasing other critical functions of the placenta, such as the endocrine production and the nourishment of the fetus. In addition, we pinpoint pathogen-specific effects in some placental lineages, including a strong mitochondrial alteration in the Hofbauer cells in response to T. gondii. Finally, we identify adaptive strategies and validate nutrient acquisition employed by the P. falciparum during placental malaria infection. This study provides the first detailed cellular map of the first-trimester placenta upon infection and describes the early events that may lead to fetal and placental disorders if left unchecked.

genomics↗

bioRxiv2022

p53 mutation in normal esophagus promotes multiple stages of carcinogenesis but is constrained by clonal competition

Aging normal human epithelia, such as the esophagus, accumulate a substantial burden of TP53 mutant clones. These are the origin of most esophageal squamous carcinomas, in which biallelic TP53 disruption is almost frequent. However, the cellular mechanisms by which p53 mutants colonize the esophagus and participate in the subsequent stages of transformation are unclear. Here we show that inducing the p53R245W mutant in single esophageal progenitor cells in transgenic mice confers a proliferative advantage that drives clonal expansion but does not disrupt normal epithelial structure or function. Loss of the remaining p53 allele in mutant cells does not increase their competitive fitness, creating a bottleneck to the development of chromosomally unstable p53R245W/null epithelium. In carcinogenesis, p53 mutation does not initiate tumor formation, but tumors developing from areas with p53 mutation and LOH are larger and show extensive chromosomal instability compared to lesions arising in wild type epithelium. We conclude that p53 has distinct functions at different stages of carcinogenesis and that LOH within p53 mutant clones in normal epithelium is a critical step in malignant transformation.

cancer biology↗

bioRxiv2021

Precancer: Mutant clones in normal epithelium outcompete and eliminate esophageal micro-tumors

Human epithelial tissues accumulate cancer-driver mutations with age1-7, yet tumor formation remains rare. The positive selection of these mutations argues they alter the behavior and fitness of proliferating cells8-10. Hence, normal adult tissues become a patchwork of mutant clones competing for space and survival, with the fittest clones expanding by eliminating their less-competitive neighbors9-12. However, little is known about how such dynamic competition in normal epithelia impacts early tumorigenesis. Here we show that the majority of newly formed esophageal tumors are eliminated through competition with mutant clones in the surrounding normal epithelium. We followed the fate of microscopic tumors in a mouse model of esophageal carcinogenesis. Most neoplasms are rapidly lost despite no indication of tumor cell death, decreased proliferation, or an anti-tumor immune response. Deep-sequencing of 10-day and 1-year-old tumors shows evidence of genetic selection on the surviving neoplasms. Induction of highly competitive clones in transgenic mice increased tumor removal, while pharmacologically inhibiting clonal competition reduced tumor loss. The results are consistent with a model where survival of early neoplasms depends on their competitive fitness relative to that of mutant clones in the adjacent normal tissue. We have identified an unexpected anti-tumorigenic role for mutant clones in normal epithelium by purging early neoplasms through cell competition, thereby preserving tissue integrity.

cancer biology↗

bioRxiv2021

Levelling out differences in aerobic glycolysis neutralizes the competitive advantage of oncogenic PIK3CA mutant progenitors in the esophagus

Normal human tissues progressively accumulate cells carrying mutations. Activating mutations in PIK3CA generate large clones in the aging human esophagus, but the underlying cellular mechanisms are unclear. Here, we tracked mutant PIK3CA esophageal progenitor cells in transgenic mice by lineage tracing. Expression of an activating heterozygous Pik3caH1047R mutation in single progenitor cells tilts cell fate towards proliferation, generating mutant clones that outcompete their wild type neighbors. The mutation leads to increased aerobic glycolysis through the activation of Hif1 transcriptional targets compared with wild type cells. We found that interventions that level out the difference in activation of the PI3K/HIF1/aerobic glycolysis axis between wild type and mutant cells attenuate the competitive advantage of Pik3caH1047R mutant cells in vitro and in vivo. Our results suggest that clinically feasible interventions that even out signaling imbalances between wild type and mutant cells may limit the expansion of oncogenic mutants in normal epithelia.

cancer biology↗

bioRxiv2020

Illuminating chromatin compaction in live cells and fixed tissues using SiR-DNA fluorescence lifetime

The global compaction state of chromatin in a nucleus is an important component of cell identity that has been difficult to measure. We have developed a quantitative method to measure the chromatin compaction state in both live and fixed cells, without the need for genetic modification, using the fluorescence lifetime of SiR-DNA dye. After optimising this method using live cancer cell lines treated to induce chromatin compaction or decompaction, we observed chromatin compaction in differentiating epithelial cells in fixed tissue sections, as well as local decompaction foci that may represent transcription factories. In addition, we shed new light on chromatin decompaction during embryonic stem cell transition out of their naive pluripotent state. This method will be useful to studies of nuclear architecture, and may be easy, cheap, and accessible enough to serve as a general assay of stem-ness.

cell biology↗

bioRxiv2019

The single-progenitor model as the unifying paradigm of squamous epithelial maintenance

Adult tissues such as the epidermis of the skin and the epithelium lining the esophagus are continuously turned over throughout life. Cells are shed from the tissue surface and replaced by cell division. Yet, the cellular mechanisms that underpin these tissues homeostasis remain poorly established, having important implications for wound healing and carcinogenesis. Lineage tracing, in which of a cohort of proliferating cells and their descendants are genetically labelled in transgenic mice, has been used to study the fate behavior of the proliferating cells that maintain these tissues. However, based on this technique, distinct mutually irreconcilable models, differing in the implored number and hierarchy of proliferating cell types, have been proposed to explain homeostasis. To elucidate which of these conflicting scenarios should prevail, here we performed cell proliferation assays across multiple body sites in transgenic H2BGFP mouse epidermis and esophagus. Cell-cycle properties were then extracted from the H2BGFP dilution kinetics and adopted in a common analytic approach for a refined analysis of a new lineage-tracing experiment and eight published clonal data sets from esophagus and different skin territories. Our results show H2BGFP dilution profiles remained unimodal over time, indicating the absence of slow-cycling stem cells across all tissues analyzed. We find that despite using diverse genetic labelling approaches, all lineage-tracing data sets are consistent with tissues maintenance by a single population of proliferating cells. The outcome of a given division is unpredictable but, on average the likelihood of producing proliferating and differentiating cells is balanced, ensuring tissue homeostasis. The fate outcomes of sister cells are anticorrelated. We conclude a single cell population maintains squamous epithelial homeostasis.

cell biology↗