Biology subjects

Pereira, L.

Publications and source records attributed to Pereira, L..

35 records · Page 2Linked to original sources

bioRxiv2022

Resistance to EGFR inhibitors in lung cancer occurs through horizontal transfer and is associated with increased caveolins expression

Resistance to treatment is a major clinical problem and a major cause of cancer-related deaths. Understanding the biological basis of resistance acquisition is of utmost importance to improve the clinical management of cancer patients. NGS analysis of human lung cancer (LC) tumors from patients that relapsed after treatment with EGFR-tyrosine kinase inhibitors (TKI), revealed that the p.T790M resistance mutation is not present in all the relapsing tumor cells, suggesting that LC cells can become resistant even if not carrying the p.T790M mutation. Using in vitro treatments with conditioned medium (CM) and in vivo co-inoculation experiments, we show that LC cells sensitive to EGFR-TKIs (S cells) acquire resistance faster when treated with CM from LC cells resistant to EGFR-TKIs (R cells) or when co-inoculated with R cells in opposite flanks of the same animal. Importantly, we show that acquisition of resistance is not due to the emergence of subpopulations of cancer cells with new resistance mutations. Using transcriptomics, we show that acquisition of resistance is associated with upregulation of genes involved in endocytosis, namely caveolins CAV1 and CAV2. These findings were validated in human clinical samples, where an increase in CAV1 and CAV2 expression was associated with tumor relapse after treatment with EGFR-TKIs. Our results suggest that acquisition of resistance to targeted therapies results from the combined effect of selection of cells harboring specific resistance mutations and horizontal transfer of the resistance phenotype. These findings may pave the way to bring intercellular communication into the realm of cancer treatment. One Sentence SummaryResistance to EGFR inhibitors is transferred horizontally between lung cancer cells and is associated with gain of expression of caveolins.

cancer biology↗

bioRxiv2022

Dynamic changes in gas solubility of xylem sap reiterate the enigma of plant water transport under negative pressure

Despite a long research history, we do not fully understand why plants are able to transport xylem sap under negative pressure without constant failure. Microbubble formation via direct gas entry is assumed to cause hydraulic failure, while the concentration of gas dissolved in xylem sap is traditionally supposed to be constant, following Henrys law. Here, the concentration of soluble gas in xylem sap was estimated in vivo using well-watered Citrus plants under varying levels of air temperature and photoperiodic exposure, and compared to modelled data. The gas concentration in xylem sap showed non-equilibrium curves, with a minimum over- or undersaturation of 5% compared to gas solubility based on Henrys law. A similar diurnal pattern was obtained from the gas concentration in the cut-open conduits and discharge tube, and oversolubility was strongly associated with decreasing xylem water potentials during transpiration. Although our model did not explain the daily changes in gas solubility for an anisobaric situation, oversolubility characterises nanoconfined liquids, such as sap inside cell walls. Thus, plants are able to transport sap under negative pressure with relatively high amounts of dissolved gas, providing them with a buffering capacity to prevent hydraulic failure, despite diurnal changes in pressure and temperature.

plant biology↗

bioRxiv2021

Selective whole genome amplification as a tool to enrich specimens with low Treponema pallidum genomic DNA copies for whole genome sequencing

Downstream next generation sequencing (NGS) of the syphilis spirochete Treponema pallidum subspecies pallidum (T. pallidum) is hindered by low bacterial loads and the overwhelming presence of background metagenomic DNA in clinical specimens. In this study, we investigated selective whole genome amplification (SWGA) utilizing multiple displacement amplification (MDA) in conjunction with custom oligonucleotides with an increased specificity for the T. pallidum genome, and the capture and removal of CpG-methylated host DNA using the NEBNext(R) Microbiome DNA Enrichment Kit followed by MDA with the REPLI-g Single Cell Kit as enrichment methods to improve the yields of T. pallidum DNA in isolates and lesion specimens from syphilis patients. Sequencing was performed using the Illumina MiSeq v2 500 cycle or NovaSeq 6000 SP platform. These two enrichment methods led to 93-98% genome coverage at 5 reads/site in 5 clinical specimens from the United States and rabbit propagated isolates, containing >14 T. pallidum genomic copies/l of sample for SWGA and >129 genomic copies/l for CpG methylation capture with MDA. Variant analysis using sequencing data derived from SWGA-enriched specimens, showed that all 5 clinical strains had the A2058G mutation associated with azithromycin resistance. SWGA is a robust method that allows direct whole genome sequencing (WGS) of specimens containing very low numbers of T. pallidum, which have been challenging until now. ImportanceSyphilis is a sexually transmitted, disseminated acute and chronic infection caused by the bacterial pathogen Treponema pallidum subspecies pallidum. Primary syphilis typically presents as single or multiple mucocutaneous lesions, and if left untreated, can progress through multiple stages with varied clinical manifestations. Molecular studies often rely on direct amplification of DNA sequences from clinical specimens; however, this can be impacted by inadequate samples due to disease progression or timing of patients seeking clinical care. While genotyping has provided important data on circulating strains over the past two decades, WGS data is needed to better understand strain diversity, perform evolutionary tracing, and monitor antimicrobial resistance markers. The significance of our research is the development of a SWGA DNA enrichment method that expands the range of clinical specimens that can be directly sequenced to include samples with low numbers of T. pallidum.

genomics↗

bioRxiv2021

Projecting ancient ancestry in modern-day Arabians and Iranians: a key role of the past exposed Arabo-Persian Gulf on human migrations

Arabian Peninsula is strategic for investigations centred on the structuring of the modern human population in the three main groups, in the awake of the out-of-Africa migration. Despite the poor climatic conditions for recovery of ancient DNA human evidence in Arabia, the availability of genomic data from neighbouring ancient specimens and of informative statistical tools allow better modelling the ancestry of these populations. We applied this approach to a dataset of 741,000 variants screened in 291 Arabians and 78 Iranians, and obtained insightful evidence. The west-east axis was a strong forcer of population structure in the Peninsula, and, more importantly, there were clear continuums throughout time linking west Arabia with Levant, and east Arabia with Iran and Caucasus. East Arabians also displayed the highest levels of the basal Eurasian lineage of all tested modern-day populations, a signal that was maintained even after correcting for possible bias due to recent sub-Saharan African input in their genomes. Not surprisingly, east Arabians were also the ones with higher similarity with Iberomaurusians, who were so far the best proxy for the basal Eurasians amongst the known ancient specimens. The basal Eurasian lineage is the signature of ancient non-Africans that diverged from the common European-East Asian pool before 50 thousand years ago, and before the later interbred with Neanderthals. Our results are strong evidence to include the exposed basin of the Arabo-Persian Gulf as possible home of basal Eurasians, to be investigated further on namely by searching ancient Arabian human specimens.

genetics↗

bioRxiv2021

Accuracy of the pneumatic method for estimating xylem vulnerability to embolism in temperate diffuse-porous tree species

O_LIThe increasing frequency of global change-type droughts has created a need for fast, accurate and widely applicable techniques for estimating xylem embolism resistance to improve forecasts of future forest changes. C_LIO_LIWe used data from 12 diffuse-porous temperate tree species covering a wide range of xylem safety to compare the pneumatic and flow-centrifuge method for constructing xylem vulnerability curves. We evaluated the agreement between parameters estimated with both methods and the sensitivity of pneumatic measurements to the measurement duration. C_LIO_LIThe agreement between xylem water potentials at 50% air discharged (PAD) estimated with the Pneumatron and 50% loss of hydraulic conductivity (PLC) estimated with the flow-centrifuge method was high (mean signed deviation: 0.12 MPa, Pearson correlation: 0.96 after 15 sec of gas extraction). However, the relation between the estimated slopes was more variable, resulting in lower agreement in xylem water potential at 12% and 88% PAD/PLC. All parameters were sensitive to the duration of the pneumatic measurement, with highest overall agreement between methods after 16 sec. C_LIO_LIWe conclude that, if applied correctly, the pneumatic method enables fast and inexpensive estimations of embolism resistance for a wide range of temperate, diffuse-porous species, which makes it attractive for predicting plant performance under climate change. C_LI

plant biology↗

bioRxiv2021

A user manual to measure gas diffusion kinetics in plants: Pneumatron construction, operation and data analysis

The Pneumatron device presented measures gas diffusion kinetics in the xylem of plants. The device provides an easy, low-cost, and powerful tool for research on plant water relations. Here, we describe in detail how to construct and operate this device to estimate xylem vulnerability to embolism, and how to analyse pneumatic data. Simple and more elaborated ways of constructing a Pneumatron are shown, either using wires, a breadboard, or a printed circuit board. The instrument is based on an open-source hardware and software system, which allows users to operate it in an automated or semi-automated way. A step-by-step manual and a troubleshooting section are provided. An excel spreadsheet and an R-script are also presented for fast and easy data analysis. This manual should help new users to avoid common mistakes, especially regarding stable measurements of the minimum and maximum amount of gas that can be discharged from xylem tissue.

plant biology↗

bioRxiv2021

A Unit Pipe Pneumatic model to simulate gas kinetics during measurements of embolism in excised angiosperm xylem

O_LIThe Pneumatic method has been introduced to quantify embolism resistance in plant xylem of various organs. Despite striking similarity in vulnerability curves between the Pneumatic and hydraulic methods, a modeling approach is highly needed to demonstrate that xylem embolism resistance can be accurately quantified based on gas diffusion kinetics. C_LIO_LIA Unit Pipe Pneumatic (UPPn) model was developed to estimate gas diffusion from intact conduits, which were axially interconnected by interconduit pit membranes. The physical laws used included Ficks law for diffusion, Henrys law for gas concentration partitioning between liquid and gas phases at equilibrium, and the ideal gas law. C_LIO_LIThe UPPn model showed that 91% of the extracted gas came from the first two series of embolized, intact conduits, and only 9% from the aqueous phase after 15 s of simulation. Embolism resistance measured with a Pneumatic apparatus was systematically overestimated by 2 to 17%, corresponding to a typical measuring error of 0.11 MPa for P50 (the water potential equivalent to 50% of the maximum amount of gas extracted). C_LIO_LIBecause results from the UPPn model are supported by experimental evidence, there is a good theoretical and experimental basis for applying the pneumatic method to research on embolism resistance of angiosperms. C_LI

plant biology↗

bioRxiv2020

Pore constrictions in intervessel pit membranes reduce the risk of embolism spreading in angiosperm xylem

O_LIEmbolism spreading in angiosperm xylem occurs via mesoporous pit membranes between vessels. Here, we investigate how the size of pore constrictions in pit membranes is related to pit membrane thickness and embolism resistance. C_LIO_LIIn three models, pit membranes are modelled as multiple layers to investigate how pit membrane thickness and the number of intervessel pits per vessel determine pore constriction sizes, the probability of encountering large pores, and air-seeding. These estimations were complemented by measurements of pit membrane thickness, embolism resistance, and number of intervessel pits per vessel (n = 31, 31, and 20 species, respectively). C_LIO_LIConstriction sizes in pores decreased with increasing pit membrane thickness, which agreed with the measured relationship between pit membrane thickness and embolism resistance. The number of pits per vessel affected constriction size and embolism resistance much less than pit membrane thickness. A strong relationship between estimated air-seeding pressures and measured embolism resistance was observed. C_LIO_LIPore constrictions provide a mechanistic explanation why pit membrane thickness determines embolism resistance, and suggest that hydraulic safety can be uncoupled from hydraulic efficiency. Although embolism spreading remains puzzling and encompasses more than pore constriction sizes, angiosperms are unlikely to have leaky pit membranes, which enables tensile transport of water. C_LI

plant biology↗

bioRxiv2020

GUN1-independent retrograde signaling targets the ethylene pathway to repress photomorphogenesis

When germinating in the light, Arabidopsis seedlings undergo photomorphogenic development, characterized by short hypocotyls, greening and expanded cotyledons. Stressed chloroplasts emit retrograde signals to the nucleus that induce developmental responses and repress photomorphogenesis. The nuclear targets of these retrograde signals are not yet fully known. Here, we show that lincomycin-treated seedlings (which lack developed chloroplasts) show strong phenotypic similarities to seedlings treated with ethylene (ET) precursor 1-aminocyclopropane-1-carboxylic acid (ACC), as both signals inhibit cotyledon separation in the light. We show that the lincomycin-induced phenotype partly requires a functioning ET signaling pathway, but could not detect increased ET emissions in response to lincomycin treatment. The two treatments show overlap in up-regulated gene transcripts, downstream of transcription factors ETHYLENE INSENSITIVE3 (EIN3) and EIN3-LIKE1 (EIL1). The induction of the ethylene signaling pathway is triggered by an unknown retrograde signal acting independently of GENOMES UNCOUPLED1 (GUN1). Our data show how two apparently different stress responses converge to optimize photomorphogenesis. One Sentence SummaryChloroplast retrograde signaling targets the ethylene-regulated gene network to repress photomorphogenesis in Arabidopsis

plant biology↗

bioRxiv2020

A semi-automated method for measuring xylem vessel length distribution

Knowledge about the length of xylem vessels is essential to understand water transport in plants because these multicellular units show a 100-fold variation, from less than a centimeter to many meters. However, the available methods to estimate vessel length distribution (VLD) are excessively time consuming and do not allow large and in-depth surveys. Here, we describe a semi-automated method to measure VLD using an automated Pneumatron device. Gas conductivity of a xylem tissue with a certain length is estimated in a straightforward and precise way with the Pneumatron in a way theoretically similar to the air-injection method. The method presented enables fast and easy measurements using multiple devices simultaneously (>50 samples day-1), which is a significant advantage. Here, the apparatus is described in detail as well as how measurements are taken. We also present the software and an R-script for data analysis. The method described represents an important contribution to studies on plant hydraulic architecture and can improve our understanding about the role of VLD in plant performance under varying water availability.

plant biology↗

bioRxiv2020

Root xylem in three woody angiosperm species is not more vulnerable to embolism than stem xylem

AimsSince plants are compartmentalised organisms, failure of their hydraulic transport system could differ between organs. We test here whether xylem tissue of stems and roots differ in their drought-induced embolism resistance, and whether intact roots are equally resistant to embolism than root segments. MethodsEmbolism resistance of stem and root xylem was measured based on the pneumatic technique for Acer campestre, A. pseudoplatanus and Corylus avellana, comparing also intact roots and root segments of C. avellana. Moreover, we compared anatomical features such as interconduit pit membrane between roots and stems. ResultsWe found a higher embolism resistance for roots than stems, although a significant difference was only found for A. pseudoplatanus. Interconduit pit membrane thickness was similar for both organs of the two Acer species, but pit membranes were thicker in roots than stems of C. avellana. Also, embolism resistance of an intact root network was similar to thick root segments for C. avellana. ConclusionOur observations show that root xylem is not more vulnerable to embolism than stem xylem, although more species need to be studied to test if this finding can be generalised. We also demonstrated that the pneumatic method can be applied to non-terminal plant samples.

ecology↗

bioRxiv2019

Is HSPG2 a modifier gene for Marfan syndrome?

Marfan syndrome (MFS) is a connective tissue disease caused by mutations in the FBN1 gene. Nevertheless, other genes influence the manifestations of the disease, characterized by high clinical variability even within families. We mapped modifier loci for cardiovascular and skeletal manifestations in the mg{Delta}loxPneo mouse model for MFS and the synthenic loci in the human genome. Corroborating our findings, one of those loci was identified also as a modifier locus in MFS patients. Here we investigate the HSPG2 gene, located in this region, as a candidate modifier gene for MFS. We show a correlation between Fbn1 and Hspg2 expression in spinal column and aorta in non-isogenic mg{Delta}loxPneo mice. Moreover, we show that mice with severe phenotypes present lower expression of Hspg2 than those mildly affected. Thus, we propose that HSPG2 is a strong candidate modifier gene for MFS and its role in modulating disease severity should be investigated in patients.

genetics↗

bioRxiv2019

Genome-wide analysis of Corsican population reveals a close affinity with Northern and Central Italy

Despite being the fourth largest island in the Mediterranean basin, the genetic variation of Corsica has not been explored as exhaustively as Sardinia, which is situated only 11 km South. However, it is likely that the populations of the two islands shared, at least in part, similar demographic histories. Moreover, the relative small size of the Corsica may have caused genetic isolation, which, in turn, might be relevant under medical and translational perspectives. Here we analysed genome wide data of 16 Corsicans, and integrated with newly (33 individuals) and previously generated samples from West Eurasia and North Africa. Allele frequency, haplotype-based, and ancient genome analyses suggest that although Sardinia and Corsica may have witnessed similar isolation and migration events, the latter is genetically closer to populations from continental Europe, such as Northern and Central Italians.

genomics↗

bioRxiv2018

Timing mechanism of sexually dimorphic nervous system differentiation

In all animals, sexual differentiation of somatic tissue is precisely timed, yet the molecular mechanisms that control the timing of sexual differentiation, particularly in the brain, are poorly understood. We have used sexually dimorphic molecular, anatomical and behavioral features of the C. elegans nervous system to decipher a regulatory pathway that controls the precise timing of sexual differentiation. We find that the sexually dimorphic differentiation of postmitotic neurons in the male nervous system is abrogated in animals that carry a mutation in the miRNA let-7 and prematurely executed in animals either lacking the let-7 inhibitor lin-28, or the direct let-7 target lin-41, an RNA-binding, posttranscriptional regulator. We show that an isoform of a phylogenetically conserved transcription factor, lin-29a, is a critical target of LIN-41 in controlling sexual maturation of sex-shared neurons. lin-29a is expressed in a male-specific manner in a subset of sex-shared neurons at the onset of sexual maturation. lin-29a acts cell-autonomously in these neurons to control the expression of sexually dimorphic neurotransmitter switches, sensory receptor expression, neurite anatomy and connectivity, and locomotor behavior. lin-29a is not only required but also sufficient to impose male-specific features at earlier stages of development and in the opposite sex. The temporal, sexual and spatial specificity of lin-29a expression is controlled intersectionally through the lin-28/let-7/lin-41 heterochronic pathway, sex chromosome configuration and neuron type-specific terminal selector transcription factors. Two Doublesex-like transcription factors represent additional neuron-type specific targets of LIN-41 and are regulated in a similar intersectional manner, indicating the existence of modular outputs downstream of the heterochronic pathway. In conclusion, we have provided insights into the molecular logic of the timing of sexual differentiation in the C. elegans nervous system. Remarkably, the lin28/let7 axis also controls the timing of sexual differentiation in mice and humans thereby hinting toward a striking universality of the control mechanisms of sexual differentiation.

neuroscience↗

bioRxiv2017

A methodology for unsupervised clustering using iterative pruning to capture fine-scale structure

SNP-based information is used in several existing clustering methods to detect shared genetic ancestry or to identify population substructure. Here, we present a methodology for unsupervised clustering using iterative pruning to capture fine-scale structure called IPCAPS. Our method supports ordinal data which can be applied directly to SNP data to identify fine-scale population structure. We compare our method to existing tools for detecting fine-scale structure via simulations. The simulated data do not take into account haplotype information, therefore all markers are independent. Although haplotypes may be more informative than SNPs, especially in fine-scale detection analyses, the haplotype inference process often remains too computationally intensive. Therefore, our strategy has been to restrict attention to SNPs and to investigate the scale of the structure we are able to detect with them. We show that the experimental results in simulated data can be highly accurate and an improvement to existing tools. We are convinced that our method has a potential to detect fine-scale structure.

bioinformatics↗

bioRxiv2017

IPCAPS: an R package for iterative pruning to capture population structure

BackgroundResolving population genetic structure is challenging, especially when dealing with closely related or geographically confined populations. Although Principal Component Analysis (PCA)-based methods and genomic variation with single nucleotide polymorphisms (SNPs) are widely used to describe shared genetic ancestry, improvements can be made especially when fine-scale population structure is the target.\n\nResultsThis work presents an R package called IPCAPS, which uses SNP information for resolving possibly fine-scale population structure. The IPCAPS routines are built on the iterative pruning Principal Component Analysis (ipPCA) framework that systematically assigns individuals to genetically similar subgroups. In each iteration, our tool is able to detect and eliminate outliers, hereby avoiding severe misclassification errors.\n\nConclusionsIPCAPS supports different measurement scales for variables used to identify substructure. Hence, panels of gene expression and methylation data can be accommodated as well. The tool can also be applied in patient sub-phenotyping contexts. IPCAPS is developed in R and is freely available from bio3.giga.ulg.ac.be/ipcaps

bioinformatics↗

bioRxiv2017

Is it possible to keep the exoskeleton of the crab Callinectes ornatus soft for several days?

Soft-shell crab is considered a gastronomic delicacy, reaching high values in the international market. The process of hardening of the crabs exoskeleton after moulting takes approximately two days to complete; however, the duration for which the shell remains at the consistency of high commercial value is only 3 hours on average. After this period, the shell assumes a consistency classified as \"paper\", later becoming \"hard\" again. The goal of this work was to evaluate the use of the crabs themselves to alter the chemical characteristics of the water and thereby increase the amount of time during which they can be marketed as \"soft-shell crab\". In this work, 241 individuals of Callinectes ornatus were used in two experiments. In the first experiment, the animals were maintained in a collective system with filtration and partial daily water renewal. In the second experiment, the crabs were maintained in a collective system with filtration but no water renewal. In Experiment 1, the chemical characteristics of the water remained unchanged over time (p > 0.05), and the median time to hardening of the exoskeleton to the paper consistency after moulting was 3 hours. Over the course of Experiment 2, there was a significant reduction (p < 0.05) in pH and significant increases in the ammonia and nitrite concentrations. When moulting occurred in water with a pH below 7.3 and total ammonia concentrations above 6.0 mg/L, the crabs shells did not harden, and it was possible to keep them soft for up to 5 days.

physiology↗