Biology subjects

Goncalves, J.

Publications and source records attributed to Goncalves, J..

17 recordsLinked to original sources

bioRxiv2026

Dissecting Complex Interactions Between Ferroptosis and the Proteasome

The proteasome is an essential multiprotein complex whose inhibition can lead to apoptosis. Ferroptosis is a non-apoptotic cell death mechanism whose fundamental regulation continues to be elucidated. How proteasome function regulates ferroptosis sensitivity is poorly understood and difficult to study given the essential nature of the proteasome. Here, we isolated the effects of proteasome inhibition on ferroptosis by combining direct cell death imaging, cell death pathway-specific inhibitors, and mathematical modeling. We find that proteasome inhibition enhances sensitivity to ferroptosis induced by glutathione peroxidase 4 (GPX4) inhibition while simultaneously promoting resistance to ferroptosis induced by system x - inhibition. Sensitization to GPX4 inhibition requires protein synthesis but not the apoptosis execution machinery and is opposed by the activating transcription factor 4 (ATF4) stress response pathway. This work demonstrates a complex role for proteasome function in ferroptosis regulation and establishes new methods to dissect cross-talk between ferroptosis and essential cellular processes.

cell biology↗

bioRxiv2026

Combined impacts of invasive alien species and fire on ecosystems are complex, mostly negative, and understudied: a global review

Invasion-fire interactions can negatively impact ecosystems by driving biodiversity loss, altering ecological processes, modifying habitat structure, and compromising ecosystem functioning. Understanding how this interaction operates is essential to design effective management strategies that are successful in controlling both invasive alien species (IAS) and fire. Therefore, the present literature review aims to synthesize the current knowledge on invasion-fire interaction and its impacts on ecosystems, as well as identify knowledge gaps in the field. The review included 464 studies, from which information on context, species and fire characteristics, interaction outcomes, and research approaches was extracted. Fire generally promotes IAS, although studies on invasive animals are limited and no research has examined the effects of fire on fungi. Management through prescribed fire showed significantly better outcomes than wildfires in suppressing IAS, yet positive impacts still outnumbered the negative ones. In turn, IAS can change fire regimes causing regime shifts, but this direction of interaction is much less studied. Combined impacts of fire and IAS on ecosystems are predominantly negative, although interactions are complex and not always synergistic. Key knowledge gaps include geographic regions with known Invasion-fire interactions that remain underreported, a lack of broad-scale studies, limited management interventions, understudied taxa, and limited understanding of the combined effects on ecosystems. Remote sensing and laboratory experiments, which have been rarely used, could address some of these gaps.

ecology↗

bioRxiv2025

Integrative Spatial Omics for Systems-Level Mapping of Pathological Niches

Spatial omics technologies are a powerful tool for mapping the relationship between cellular organization and molecular distributions in healthy and diseased tissue microenvironments. Here, we describe a novel multimodal pipeline that represents experimental and computational advances for spatiomolecular analysis of tissue samples across molecular classes. This adaptable method integrates matrix-assisted laser desorption/ionization imaging mass spectrometry spatial lipidomics, spatial transcriptomics, protein imaging via multiplexed immunofluorescence microscopy, and histopathological staining to uncover spatiomolecular profiles associated with unique cellular niches and pathological features. We demonstrate the power of this approach using two different complex human disease systems: Alzheimers disease in human brain tissue and type 2 diabetes mellitus in the human pancreas. This work establishes and demonstrates a generalizable framework for multimodal spatial integration, enabling precise mapping of molecular mechanisms that underlie complex tissue pathologies.

systems biology↗

bioRxiv2024

Gene regulatory network inference from single-cell data using optimal transport

Modelling gene expression is a central problem in systems biology. Single-cell technologies have revolutionised the field by enabling sequencing at the resolution of individual cells. This results in a much richer data compared to what is obtained by bulk technologies, offering new possibilities and challenges for gene regulatory network inference. Here we introduce GRIT -- a method to fit a differential equation model and to infer gene regulatory networks from single-cell data using the theory of optimal transport. The idea consists in tracking the evolution of the cell distribution over time and finding the system that minimises the transport cost between consecutive time points.

bioinformatics↗

bioRxiv2023

Kisspeptin modulates sex-like phenotypes in the neonatal period

In recent years, our understanding of the physiological role of kisspeptin (Kiss1) has evolved significantly, revealing its pivotal role in shaping sexual differentiation within the brain. Predominantly expressed in the hypothalamus, Kiss1 plays a crucial role in regulating release of the gonadotrophin-releasing hormone (GnRH). Kisspeptin 1 receptor (Kiss1R) exhibits sex-dependent expression in various brain regions, including the medial amygdala and preoptic area (POA) of the hypothalamus, regions underlying sexual development. Importantly, in the critical period for the sexual development in mammals occurs a synchronized surge of Kiss1 and testosterone, highlighting its importance in sexual dimorphism. It has been suggested that the neonatal Kiss1 surge is associated with the buildup of gender-linked social development. We investigated this hypothesis by testing the effect of neonatal blockage of Kiss1 signaling on the sex-like phenotype in rats. We postulate that Kiss1 actions are both time and region-dependent, particularly during the neonatal period, having lasting impact on social interactions. For that we injected intracerebroventricular (ICV) a Kiss1R antagonist, Kp234, in the first 24h of life. Our data showed, that neonatal Kiss1 is determinant in modulating physiological responses, such as weight and testis development in males (N: males Veh = 10; males Kp234 = 10). Also, we document, for the first time, that Kiss1 modulates the testosterone surge in males and inhibits it in females (N: males Vehicle = 3; males Kp234 = 4; females Vehicle = 3; females Kp234 = 4). Furthermore, neonatal Kiss1 blockade modulated the sexual drive in, while the sexual performance in both sexes is maintained males (N: males Vehicle = 8; males Kp234 = 9; females Vehicle = 7; females Kp234 = 8). In conclusion, this study underscores the significance of Kiss1 during the neonatal period, a critical phase for sexual differentiation and neuroplasticity. Neonatal Kiss1 emerges as a key player in the development of a sex-like phenotype in rats. Additionally, our findings show dimorphic patterns in Kiss1 influence. This work raises the intriguing possibility that the early influence of Kiss1 in brain circuitry is pivotal in the modulation of gender and plays a role in the social construction of the bimodality of gender identity.

neuroscience↗

bioRxiv2023

The maintenance of centriole appendages and motile cilia basal body anchoring relies on TBCCD1

Centrosomes are organelles consisting of two structurally and functionally distinct centrioles, with the mother centriole having complex distal (DA) and subdistal appendages (SDA). Despite their importance, how appendages are assembled and maintained remains unclear. This study investigated human TBCCD1, a centrosomal protein essential for centrosome positioning, to uncover its localization and role at centrioles. We found that TBCCD1 localizes at both proximal and distal regions of the two centrioles, forming a complex structure spanning from SDA to DA and extending inside and outside the centriole lumen. TBCCD1 depletion caused centrosome mispositioning, which was partially rescued by taxol, and the loss of microtubules (MTs) anchored to centrosomes. TBCCD1 depletion also reduced levels of SDA proteins involved in MT anchoring such as Centriolin/CEP110, Ninein, and CEP170. Additionally, TBCCD1 was essential for the correct positioning of motile cilia basal bodies and associated structures in Paramecium. This study reveals that TBCCD1 is an evolutionarily conserved protein essential for centriole and basal body localization and appendage assembly and maintenance. A BioID screening also linked TBCCD1 to ciliopathy-associated protein networks.

cell biology↗

bioRxiv2022

Systematic analysis and optimization of early warning signals for critical transitions

Abrupt shifts between alternative regimes occur in complex systems, from cell regulation to brain functions to ecosystems. Several model-free Early Warning Signals (EWS) have been proposed to detect impending transitions, but failure or poor performance in some systems have called for better investigation of their generic applicability. In particular, there are still ongoing debates whether such signals can be successfully extracted from data. In this work, we systematically investigate properties and performance of dynamical EWS in different deteriorating conditions, and we propose an optimised combination to trigger warnings as early as possible, eventually verified on experimental data. Our results explain discrepancies observed in the literature between warning signs extracted from simulated models and from real data, provide guidance for EWS selection based on desired systems and suggest an optimised composite indicator to alert for impending critical transitions. HighlightsO_LIHow to extract early warning signals (EWS) against critical transitions from data is still poorly understood C_LIO_LIA mathematical framework assesses and explains the performance of EWS in noisy deteriorating conditions C_LIO_LIComposite indicators are optimised to alert for impending shifts C_LIO_LIThe results are applicable to wide classes of systems, as shown with models and on empirical data. C_LI

systems biology↗

bioRxiv2021

Amino acids 484 and 494 of SARS-CoV-2 spike are hotspots of immune evasion affecting antibody but not ACE2 binding

Understanding SARS-CoV-2 evolution and host immunity is critical to control COVID-19 pandemics. At the core is an arms-race between SARS-CoV-2 antibody and angiotensin-converting enzyme 2 (ACE2) recognition, a function of the viral protein spike. Mutations in spike impacting antibody and/or ACE2 binding are appearing worldwide, with the effect of mutation synergy still incompletely understood. We engineered 25 spike-pseudotyped lentiviruses containing individual and combined mutations, and confirmed that E484K evades antibody neutralization elicited by infection or vaccination, a capacity augmented when complemented by K417N and N501Y mutations. In silico analysis provided an explanation for E484K immune evasion. E484 frequently engages in interactions with antibodies but not with ACE2. Importantly, we identified a novel amino acid of concern, S494, which shares a similar pattern. Using the already circulating mutation S494P, we found that it reduces antibody neutralization of convalescent and post-immunization sera, particularly when combined with E484K and N501Y. Our analysis of synergic mutations provides a landscape for hotspots for immune evasion and for targets for therapies, vaccines and diagnostics. One-Sentence SummaryAmino acids in SARS-CoV-2 spike protein implicated in immune evasion are biased for binding to neutralizing antibodies but dispensable for binding the host receptor angiotensin-converting enzyme

immunology↗

bioRxiv2021

Discovery of Dynamical Network Models for Genetic Circuits from Time-Series Data with Incomplete Measurements

Synthetic gene networks are frequently conceptualized and visualized as static graphs. This view of biological programming stands in stark contrast to the transient nature of biomolecular interaction, which is frequently enacted by labile molecules that are often unmeasured. Thus, the network topology and dynamics of synthetic gene networks can be difficult to verify in vivo or in vitro, due to the presence of unmeasured biological states. Here we introduce the dynamical structure function as a new mesoscopic, data-driven class of models to describe gene networks with incomplete measurements. We introduce a network reconstruction algorithm and a code base for reconstructing the dynamical structure function from data, to enable discovery and visualization of graphical relationships in a genetic circuit diagram as time-dependent functions rather than static, unknown weights. We prove a theorem, showing that dynamical structure functions can provide a data-driven estimate of the size of crosstalk fluctuations from an idealized model. We illustrate this idea with numerical examples. Finally, we show how data-driven estimation of dynamical structure functions can explain failure modes in two experimentally implemented genetic circuits, a historical genetic circuit and a new E. coli based transcriptional event detector.

synthetic biology↗

bioRxiv2020

Transcriptome-Wide Off-Target Effects of Steric-Blocking Oligonucleotides

Steric-blocking oligonucleotides (SBOs) are short, single-stranded nucleic acids designed to modulate gene expression by binding to mRNA and blocking access from cellular machinery such as splicing factors. SBOs have the potential to bind to near-complementary sites in the transcriptome, causing off-target effects. In this study, we used RNA-seq to evaluate the off-target differential splicing events of 81 SBOs and differential expression events of 46 SBOs. Our results suggest that differential splicing events are predominantly hybridization-driven, while differential expression events are more common and driven by other mechanisms. We further evaluated the performance of in silico screens for off-target events, and found an edit distance cutoff of three to result in a sensitivity of 14% and false discovery rate of 99%. A machine learning model incorporating splicing predictions substantially improved the ability to prioritize low edit distance hits, increasing sensitivity from 4% to 26% at a fixed FDR. Despite these large improvements in performance, the approach does not detect the majority of events at a false discovery rate below 99%. Our results suggest that in silico methods are currently of limited use for predicting the off-target effects of SBOs.

molecular biology↗

bioRxiv2020

FastField: An Open-Source Toolbox for Efficient Approximation of Deep Brain Stimulation Electric Fields

Deep brain stimulation (DBS) is a surgical therapy to alleviate symptoms of certain brain disorders by electrically modulating neural tissues. Computational models predicting electric fields and volumes of tissue activated are key for efficient parameter tuning and network analysis. Currently, we lack efficient and flexible software implementations supporting complex electrode geometries and stimulation settings. Available tools are either too slow (e.g. finite element method-FEM), or too simple, with limited applicability to basic use-cases. This paper introduces FastField, an efficient open-source toolbox for DBS electric field and VTA approximations. It computes scalable e-field approximations based on the principle of superposition, and VTA activation models from pulse width and axon diameter. In benchmarks and case studies, FastField is solved in about 0.2s, ~ 1000 times faster than using FEM. Moreover, it is almost as accurate as using FEM: average Dice overlap of 92%, which is around typical noise levels found in clinical data. Hence, FastField has the potential to foster efficient optimization studies and to support clinical applications.

neuroscience↗

bioRxiv2020

Causal dynamical modelling predicts novel regulatory genes of FOXP3 in human regulatory T cells

Regulatory T cells (Tregs), characterized as a CD4+CD25+FOXP3+ subset of T cells, are vital to the induction of immune tolerance and the maintenance of immune homeostasis. While target genes of Treg master regulator FOXP3 have been identified, the upstream regulatory machinery of FOXP3 still remains largely unknown. Here we dynamically model causal relationships among genes from available time-series genome-scale datasets, to predict direct or indirect regulatory genes of FOXP3 in human primary Tregs. From the whole genome, we selected five top ranked candidates for further experimental validation. Following knockdown, three out of the five candidates indeed showed significant effects on the mRNA expression of FOXP3. Further experiments showed that one out of these three predicted candidates, namely nuclear receptor binding factor 2 (NRBF2), also affected FOXP3 protein expression. These results open new doors to identify potential new mechanisms of immune related diseases.

systems biology↗

bioRxiv2019

Differential effects of day-night cues and the circadian clock on the barley transcriptome

The circadian clock is a complex transcriptional network that regulates gene expression in anticipation of the day-night cycle and controls agronomic traits in plants. However, in crops, information on the effects of the internal clock and day-night cues on the transcriptome is limited. We analysed the diel and circadian leaf transcriptomes in the barley cultivar Bowman and derived introgression lines carrying mutations in EARLY FLOWERING 3 (ELF3), LUX1, and EARLY MATURITY 7 (EAM7). Mutations in ELF3 and LUX1 abolished circadian transcriptome oscillations under constant conditions, whereas eam7 maintained oscillations of {approx}30% of the circadian transcriptome. However, day-night cues fully restored transcript oscillations in all three mutants and thus compensated for a disrupted oscillator in the arrhythmic barley clock mutants elf3 and lux1. Nevertheless, elf3 but not lux1 affected the phase of the diel oscillating transcriptome and thus the integration of external cues into the clock. Using dynamical modelling, we predicted a structure of the barley circadian oscillator and interactions of its individual components with day-night cues. Our findings provide a valuable resource for exploring the function and output targets of the circadian clock and for further investigations into the diel and circadian control of the barley transcriptome.

plant biology↗

bioRxiv2019

Biallelic mutations in M1AP are a frequent cause of meiotic arrest leading to male infertility

Male infertility affects [~]7% of men in Western societies, but its causes remain poorly understood. The most clinically severe form of male infertility is non-obstructive azoospermia (NOA), which is, in part, caused by an arrest at meiosis, but so far only few genes have been reported to cause germ cell arrest in males. To address this gap, whole exome sequencing was performed in 60 German men with complete meiotic arrest, and we identified in three unrelated men the same homozygous frameshift variant c.676dup (p.Trp226LeufsTer4) in M1AP, encoding meiosis 1 arresting protein. Then, with collaborators from the International Male Infertility Genomics Consortium (IMIGC), we screened a Dutch cohort comprising 99 infertile men and detected the same homozygous variant c.676dup in a man with hypospermatogenesis predominantly displaying meiotic arrest. We also identified two Portuguese men with NOA carrying likely biallelic loss-of-function (LoF) and missense variants in M1AP among men screened by the Genetics of Male Infertility Initiative (GEMINI). Moreover, we discovered a homozygous missense variant p.(Pro389Leu) in M1AP in a consanguineous Turkish family comprising five infertile men. M1AP is predominantly expressed in human and mouse spermatogonia up to secondary spermatocytes and previous studies have shown that knockout male mice are infertile due to meiotic arrest. Collectively, these findings demonstrate that both LoF and missense M1AP variants that impair its protein cause autosomal-recessive meiotic arrest, non-obstructive azoospermia and male infertility. In view of the evidence from several independent groups and populations, M1AP should be included in the growing list of validated NOA genes.

genetics↗

bioRxiv2019

LUZP1 and the tumour suppressor EPLIN are negative regulators of primary cilia formation

Cilia/flagella are microtubule-based cellular projections with important sensory and motility functions. Their absence or malfunction is associated with a growing number of human diseases collectively referred to as ciliopathies. However, the fundamental mechanisms underpinning cilia biogenesis and functions remain only partly understood. Here, we show that LUZP1, and its interacting protein EPLIN, are negative regulators of primary cilia formation. LUZP1 is an actin-associated protein that localizes to both actin filaments and the centrosome/basal body. Like EPLIN, LUZP1 is an actin-stabilizing protein and likely regulates actin dynamics at the centrosome. Both proteins interact with ciliogenesis and cilia length regulators, and are potential players in the actin-dependent processes involved in centrosome to basal body conversion. Ciliogenesis deregulation caused by LUZP1 or EPLIN loss may contribute to the pathology of their associated diseases.\n\nSUMMARYGoncalves et al. show that LUZP1 and its interactor EPLIN are negative regulators of ciliogenesis. LUZP1 is a novel actin-stabilizing protein localizing at the centrosome and basal body where it may regulate actin-associated processes.

cell biology↗

bioRxiv2019

Long-term single-cell passaging of human iPSCs fully supports pluripotency and high-efficient trilineage differentiation capacity

ObjectivesTo validate a straightforward single-cell passaging cultivation method that enables high-quality maintenance of hiPSC without the appearance of karyotypic abnormalities or loss of pluripotency. MethodsCells were kept in culture for over 50 passages, following a structured chronogram of passage and monitoring cell growth by population doubling time (PDT) calculation and cell confluence. Standard procedures for iPSC monitoring as embryonic body (EB) formation, karyotyping and pluripotency markers expression were evaluated in order to monitor the cellular state in the long-term culture. Cells that underwent these tests were then subjected to differentiation into keratinocytes and cardiomyocytes to evaluate its differentiation capacity. ResultshiPSC clones maintained its pluripotent capability as well as chromosomal integrity and were able to generate derivatives from the three germ layers at high passages by embryoid body formation and high-efficient direct differentiation into keratinocytes and cardiomyocytes. ConclusionOur findings support the routine of hiPSC single-cell passaging as a reliable procedure even after long-term cultivation, providing healthy PSCs to be used in drugs discovery, toxicity and disease modeling as well as for therapeutic approaches.

cell biology↗

bioRxiv2019

Development of Triazoles based on AZT and their Anti-Viral Activity Against HIV-1

We report herein a set of 3-azido-3-deoxythymidine (AZT) derivatives based on triazoles and triazolium salts for HIV1 infection. Compounds were tested with {square}using HIV1 pre-exposure prophylaxis experimental model. All compounds were able to decrease infection and two of them were able to clear almost all the infection, suggesting that these drugs could play an important role in pre-exposure prophylaxis therapies.

biochemistry↗