Biology subjects

Portela, M.

Publications and source records attributed to Portela, M..

11 recordsLinked to original sources

bioRxiv2026

DNA replication is dispensable for developmental progression, but required for heterochromatin organization at mouse zygotic genome activation

Early mammalian development progresses through a small number of cleavage divisions with extensive transcriptional and epigenetic reprogramming that result in the very first lineage decisions, leading to blastocyst formation. Although classical embryological studies suggested that developmental progression can occur independently of normal cell division, the extent to which DNA replication contributes to these processes remains unclear. In particular, whether replication is required simply for proliferation or also for establishing the cellular states that regulate stage-specific gene expression is unknown. Here we show that mouse preimplantation development can proceed without DNA replication, but it is required to maintain proper heterochromatin organization at the 2-cell stage, the time of zygotic genome activation. The inhibition of DNA replication with aphidicolin blocked cleavage divisions without preventing zygotic genome activation, embryo morphogenesis, activation of lineage-specification factors, or the establishment of stage-specific transcriptional programs. However, replication arrest selectively altered the expression of developmental genes associated with late-replicating, Polycomb-enriched chromatin domains, and also reduced global levels of H3K27me3 and H3K9me3. These findings indicate that developmental timing in the early embryo is largely uncoupled from DNA replication and cell-cycle progression. Instead, DNA replication contributes to transcriptional fidelity by establishing or maintaining repressive chromatin states during the extensive genome reprogramming that accompanies zygotic genome activation.

developmental biology↗

bioRxiv2026

IFNγ at the crossroads of systemic protection and brain vulnerability to coronavirus infection

The COVID-19 pandemic, together with previous outbreaks caused by highly pathogenic coronaviruses, highlighted the importance of understanding the host determinants that influence pathogenesis, both to improve our understanding of disease mechanisms, and to strengthen preparedness for future outbreaks. In our previous work we demonstrated dysregulation of IFN{gamma} response genes in COVID-19 patients, associated with both age and viral burden, positioning IFN{gamma} as a key mediator of host defense against SARS-CoV-2. Here, we used a preclinical murine model of coronavirus infection based on murine hepatitis virus (MHV-A59), widely validated for studying SARS-CoV-2 pathogenesis, together with IFN{gamma} knockout (IFN{gamma}-KO) mice to investigate the role of this cytokine during infection. We performed a comprehensive morphological, biochemical, hematological and proteomic characterization of infected wild-type (WT) and IFN{gamma}-deficient mice. Plasma proteomics revealed impaired inflammatory and coagulation-related responses in IFN{gamma}-KO animals compared to WT controls. We found increased viral load and infectious particles in peripheral organs, including liver, spleen, heart and muscle in IFN{gamma}-KO mice, whereas these were significantly reduced in the brain. Region-specific analysis further demonstrated decreased viral load in the prefrontal cortex and hippocampus of IFN{gamma}-KO animals. Consistently, bioinformatics analysis of transcriptomics data of IFN{gamma}-treated primary neuron cultures, together with frontal cortex from human COVID-19 patients, revealed activation of neuroinflammatory and neurological disease-associated pathways, supporting a role for IFN{gamma} in driving brain inflammatory responses during coronavirus infection. Together, these findings reveal a dual role for IFN{gamma} during coronavirus infection: it is important for controlling systemic viral dissemination and limiting peripheral tissue damage, yet it may also promote viral susceptibility and exacerbated inflammatory responses in the brain. These results support IFN{gamma} modulation as a potential therapeutic strategy to prevent or mitigate neurological complications associated with COVID-19. Author SummaryViral infections are controlled by the immune system, but the same responses that protect the body can sometimes contribute to tissue damage and disease. We investigated how a key immune molecule, called Interferon Gamma, influences the outcome of Coronavirus infection in different parts of the body. Using a mouse model of Coronavirus infection with Interferon Gamma deficiency, we found that this molecule plays two opposing roles: it helps control the spread of the virus throughout the body, but it also increases vulnerability of the brain to infection and inflammation. When this immune signal was absent, animals showed higher amounts of virus in several peripheral organs but surprisingly had lower levels of virus in specific brain regions. We further found that this immune response was associated with changes in inflammation and neurological processes in human data. Our findings reveal that antiviral immunity is not always uniformly protective and that its effects depend on the tissue involved. Understanding this balance may help guide future approaches to reduce virus-associated complications while preserving the protective functions of the immune response.

microbiology↗

bioRxiv2026

Spores-on-a-Chip: a new sample-loading strategy for multiplexed single-cell monitoring of spore germination within a microfluidic platform

We demonstrate a new sample-loading strategy ("spot-loading") for Spores-on-a-Chip microfluidic sensors that enables multiplexed experimentation. The previous limitation of one biological sample per device is overcome through controlled sample loading at an intermediate step of the chip fabrication process. As a proof-of-concept, we use dual spore chips to compare the germination behaviours of two spore strains in two distinct microenvironments.

bioengineering↗

bioRxiv2025

Inherited Brain Fitness Deficit caused by Light Stress Promotes Glioblastoma Progression in Drosophila

Environmental and lifestyle factors can influence long-term human health outcomes, including the development of complex diseases such as cancer. Glioblastoma is the most aggressive and fatal form of brain cancer, marked by its invasive nature and resistance to standard therapies. Despite similar genetic alterations, patients diagnosed with glioblastoma often show highly variable survival outcomes, suggesting that non-genetic influences may play a critical role in tumor progression. Here we show that environmental conditions experienced by the parental line, such as permanent exposure to artificial light, can affect the brains resilience to tumor growth in the next generation. This effect, described as reduced Brain fitness, does not impair brain development but accelerates glioma progression and shortens survival in the offspring. These findings point to a novel concept, the Brain fitness hypothesis, in which inherited susceptibility to brain tumors may be shaped by parental exposures. Our results suggest that non-coding RNAs could mediate this intergenerational transmission, offering new insight into the complexity of inherited traits beyond direct genetic mutations. This work broadens current understanding of glioma biology and highlights the importance of environmental health across generations. AbstractEnvironmental exposure including modern lifestyle habits and occupational exposures describe a wide spectrum of risk factors for human health including carcinogenesis. Glioblastoma is the most common, aggressive, and lethal type of glioma. It is highly proliferative and invasive, infiltrates the surrounding brain parenchyma and is resistant to current treatments. Patients suffering from glioblastoma have a wide heterogeneity in life expectancy, even with similar mutations, that can vary from weeks to years. Many potential risk factors for glioma have been studied to date, but few provide significant correlations for tumor prognosis. Here, we provide a novel perspective on the interaction between the tumor and host depending on paternal environmental or lifestyle determinants, coined as Brain fitness hypothesis. Permanent light exposure induces genetic stress hallmarks in the progeny that do not alter central nervous system development, but compromise Brain fitness to an accelerated glioma growth, ultimately reducing life expectancy. Reduced survival in the progeny correlates with an increase in the number of activated brain genes involved in neuronal dynamics as well as those relevant for glioma progression such as presynaptic, circadian genes or genes required for the tumoral network. Among the inheritable mechanisms, ncRNAs emerge as potential vectors for the transmission of parental exposure to the subsequent generations and might explain the complexity of inheritable traits and phenotypes acquired as a pleiotropic effect. O_TEXTBOXBOX OF DEFINITIONSLight pollutome/exposome: The sum of environmental exposures or modern lifestyle habits as a source of artificial lighting or solar radiation that compromise fitness and disease progression from early stages (prenatal/postnatal) to adulthood. Luminic/Light stress (LL): Continuous light exposure (24h) at low luminance intensity (1,000 lux) of white light. Brain fitness hypothesis: Molecular factors that precondition brain susceptibility to pathological aggressions modulated by environmental or lifestyle determinants. Luminic stress effects (LSE): Genetic changes in unexposed F1 brains in response to pre-conceptional parental luminic exposure as a memory of parental experiences. Intergenerational Inheritance: Transmission of parental(F0) environmental effects to the descendants (F1or F2 depending on gestational state). C_TEXTBOX Graphical abstractMaternal pre-cexceptional exposure to light stress (1000 lux white light) induces intergenerational changes in the offspring brain fitness leading on a sex dependent light stress memory hallmark (LSE). Thus, F1 flies acquire a brain with the fitness tatus altered and preconditioned to GB development Consequently, GB grows faster and causes worse prognosis in affected animals. In this context miRNAs emerge as a potential epigenetic mechanism mediating this intergenerational effect, describing rriRNA-979 expression as a risk factor. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/672790v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@15be67borg.highwire.dtl.DTLVardef@78baddorg.highwire.dtl.DTLVardef@341c07org.highwire.dtl.DTLVardef@11c3393_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2025

Actin cytoskeletal deregulation, caused by RhoGEF2 overexpression, induces cell competition dependent on Ptp10D, Crumbs, and the Hippo signaling pathway.

In Drosophila larval epithelial tissues, cells containing mutations in the apico-basal polarity proteins, Scrib, Dlg or Lgl, are eliminated by cell competition when surrounded by wild-type cells. In these polarity-impaired cells, signaling mediated by the receptor-type tyrosine phosphatase Ptp10D upon engagement with its ligand Sas in the surrounding wild-type cells triggers cell competition via EGFR pathway inhibition and JNK pathway activation, which induces apoptosis of the mutant cells. Here, we investigate whether directly triggering cytoskeletal deregulation (which usually occurs downstream of cell polarity disruptions) is sufficient to trigger their elimination by cell competition via the Sas-Ptp10D signaling system. We show that actin cytoskeleton deregulated cells (as induced by RhoGEF2 overexpression (RhoGEF2OE)) are eliminated when surrounded by wild-type cells, and that Ptp10D knockdown increases RhoGEF2OE clone growth, revealing the importance of Ptp10D in the elimination of RhoGEF2OE cells. Mechanistically, in clones that are moderately overexpressing RhoGEF2OE, Ptp10D knockdown rescued cell elimination by reducing Hippo signaling. In this setting, JNK and EGFR-Ras signaling were not affected (in contrast to what occurs in apico-basal mutant cells), suggesting that Sas-Ptp10D may regulate the Hippo pathway directly in RhoGEF2OE cells. We also found that mutations in the apical cell polarity protein, Crb, partially rescued the elimination of RhoGEF2OE clones, showing that Crb normally plays a role in RhoGEF2OE clone elimination. In this setting, in which RhoGEF2OE is highly overexpressed, JNK and Hippo signaling were elevated while EGFR-Ras signaling was reduced, and crb loss normalized these pathways. crb mutant cells also showed reduced abundance of and apical membrane localization of Ptp10D, suggesting that Crb may play an important role in Sas-Ptp10D mediated cell competition. Thus, actin cytoskeleton deregulation, caused by RhoGEF2OE, results in clone elimination dependent on Crb, Ptp10D, and Hippo signaling. Altogether, our results reveal that Ptp10D is acting more broadly in cell competition to trigger the elimination of actin cytoskeleton deregulated loser cells, as well as polarity-impaired cells.

developmental biology↗

bioRxiv2024

Calcium signalling is required for anterior patterning in the mouse embryo

Anterior-Posterior axis formation in the mouse embryo requires the active migration of the DVE cell population at E5.5. While intracellular Ca2+ signalling has been shown to control cell migration in multiple cell contexts, it is unknown whether it is required for DVE migration. The pattern of Ca2+ activity in the mouse embryo at early peri-implantation stages is also unknown. Using the GCaMP6f Ca2+ reporter line we performed a detailed assessment of Ca2+ dynamics between E0.5 - E5.5 using live imaging. We find that prior to implantation, Ca2+ transients are rare, but at E5.5 widespread, periodic, Ca2+ transients in extra-embryonic tissues can be observed, including in the VE and ExE. In contrast, cells of the E5.5 epiblast remain relatively quiescent but show sporadic large-scale multi-cellular waves. Inhibition of SERCA at E5.5 abolishes Ca2+ transients and leads to DVE arrest, indicative that these transients are required for axial patterning. Together these results reveal the pattern of Ca2+ handling in the early mouse embryo and a novel requirement in anterior-posterior axis formation.

developmental biology↗

bioRxiv2023

The Drosophila Tumour Suppressor Lgl and Vap33 activate the Hippo pathway by a dual mechanism, involving RtGEF/Git/Arf79F and inhibition of the V-ATPase.

The tumour suppressor, Lethal (2) giant larvae (Lgl), is an evolutionarily conserved protein that was discovered in the vinegar fly, Drosophila, where its depletion results in tissue overgrowth and loss of cell polarity and tissue architecture. Our previous studies have revealed a new role for Lgl in linking cell polarity and tissue growth through regulation of the Notch (proliferation and differentiation) and the Hippo (negative tissue growth control) signalling pathways. Moreover, Lgl regulates vesicle acidification, via the Vacuolar ATPase (V-ATPase), and we showed that Lgl inhibits V-ATPase activity through Vap33 (a Vamp (v-SNARE)-associated protein, involved in endo-lysosomal trafficking) to regulate the Notch pathway. However, how Lgl acts to regulate the Hippo pathway was unclear. In this current study, we show that V-ATPase activity inhibits the Hippo pathway, whereas Vap33 acts to activate Hippo signalling. Using an in vivo affinity-purification approach we found that Vap33 binds to the actin cytoskeletal regulators RtGEF (Pix, a Rho-type guanine nucleotide exchange factor) and Git (G protein-coupled receptor kinase interacting ArfGAP), which also bind to the Hpo protein kinase, and are involved in the activation of the Hippo pathway. Vap33 genetically interacts with RtGEF and Git in Hippo pathway regulation. Additionally, we show that the ADP ribosylation factor Arf79F (Arf1), which is a Hpo interactor, is involved in the inhibition of the Hippo pathway. Altogether our data suggests that Lgl acts via Vap33 to activate the Hippo pathway by a dual mechanism, 1) through interaction with RtGEF/Git/Arf79F, and 2) through interaction and inhibition of the V-ATPase, thereby controlling epithelial tissue growth.

cell biology↗

bioRxiv2021

Establishment of 3D chromatin structure after fertilization and the metabolic switch at the morula-to-blastocyst transition require CTCF

The eukaryotic genome is tightly packed inside the nucleus, where it is organized in 3D at different scales. This structure is driven and maintained by different chromatin states and by architectural factors that bind DNA, such as the multi-zinc finger protein CTCF. Zygotic genome structure is established de novo after fertilization, but the impact of such structure on genome function during the first stages of mammalian development is still unclear. Here, we show that deletion of the Ctcf gene in mouse embryos impairs the correct establishment of chromatin structure, but initial lineage decisions take place and embryos are viable until the late blastocyst stage. Furthermore, we observe that maternal CTCF is not necessary for development. Transcriptomic analyses of mutant embryos show that the changes in metabolic and protein homeostasis programs that occur during the progression from the morula to the blastocyst depend on CTCF. Yet, these changes in gene expression do not correlate with disruption of chromatin structure, but mainly with proximal binding of CTCF to the promoter region of genes downregulated in mutants. Our results show that CTCF regulates both 3D genome organization and transcription during mouse preimplantation development, but mostly as independent processes.

developmental biology↗

bioRxiv2020

TNFα/Grnd mediate JNK/MMPs activation during glioma progression and neurodegeneration

Glioblastoma (GB) is the most aggressive and lethal tumour of the central nervous system (CNS). GB cells proliferate rapidly and display a network of ultra-long tumour microtubes (TMs) that mediate cell to cell communication. GB TMs infiltrate into the brain, enwrap neurons and facilitate the depletion of Wingless (Wg)/WNT from the neighbouring neurons. GB cells establish a positive feedback loop including Wg signalling upregulation that activates the JNK pathway and matrix metalloproteases (MMPs), in turn, these signals promote TMs infiltration, GB progression and neuronal synapse loss and degeneration. Thus, cellular and molecular signals other than primary mutations emerge as central players of GB. Here we describe the temporal organization of the events that occur in GB. We define the progressive activation of JNK pathway signalling mediated by Grindelwald (Grnd) receptor, is caused by the ligand Eiger (Egr)/TNF produced by the healthy tissue. We propose that cellular interactions of GB with the rest of the brain is an early event that precedes GB proliferation and expansion. We conclude that non-autonomous signals facilitate GB progression and contribute to the complexity and versatility of these incurable tumours.

cancer biology↗

bioRxiv2019

Proteomic profiling of Mycobacterium tuberculosis culture filtrate identifies novel O-glycosylated proteins

Despite being the subject of intensive research, tuberculosis, caused by Mycobacterium tuberculosis, remains at present the leading cause of death from an infectious agent. Secreted and cell wall proteins interact with the host and play important roles in pathogenicity. These proteins have been explored as candidate diagnostic markers, potential drug targets or vaccine antigens, and special attention has been given to the role of their post-translational modifications. With the purpose of contributing to the proteomic characterization of this important pathogen including an O-glycosylation profile analysis, we performed a shotgun analysis of culture filtrate proteins of M. tuberculosis based on a liquid nano-HPLC tandem mass spectrometry and a label-free spectral counting normalization approach for protein quantification. We identified 1314 M. tuberculosis proteins in culture filtrate and found that the most abundant proteins belong to the extracellular region or cell wall compartment, and that the functional categories with higher protein abundance factor were virulence, detoxification and adaptation, and cell wall and cell processes. In culture filtrate, 140 proteins were predicted to contain one of the three types of bacterial N-terminal signal peptides. Besides, various proteins belonging to the ESX secretion systems, and to the PE and PPE families, secreted by the type VII secretion system using nonclassical secretion signals, were also identified. O-glycosylation was identified as a frequent modification, being present in 108 proteins, principally lipoproteins and secreted immunogenic antigens. We could identify a group of proteins consistently detected in previous studies, most of which were highly abundant proteins. Interestingly, we also provide proteomic evidence for 62 novel O-glycosylated proteins, aiding to the glycoproteomic characterization of relevant antigenic membrane and exported proteins.

microbiology↗

bioRxiv2019

Wingless promotes JNK/MMPs positive feedback loop mediate tumour microtubes expansion, glioma progression and neurodegeneration

Glial cells display a network of projections (cytonemes) which mediate cell to cell communication. Under pathological conditions like glioblastoma (GB), cytonemes transform into ultra-long tumour microtubes (TMs). These filopodia infiltrate through the brain, enwrap neurons and deplete wingless (Wg)/WNT, as a consequence GB progress and neurons undergo synapse loss and degeneration. Thus TMs emerge as a central cellular feature of GB which correlates with a poor prognosis in patients and animal models. Here we describe in a Drosophila model for GB the molecular mechanisms behind TMs production, infiltration and maintenance. Glial cells are initially transformed into malignant GB upon EGFR and PI3K pathways constitutive activation, afterwards GB cells establish a positive feedback loop including Wg signalling, JNK and matrix metalloproteases (MMP). In order, Frizzled1 mediates Wg signalling upregulation which activates JNK in GB. As a consequence, MMPs are upregulated and facilitate TMs infiltration in the brain, hence GB TMs network expands and mediate further wingless depletion to close the loop.

cancer biology↗