Biology subjects

Carvalho, S.

Publications and source records attributed to Carvalho, S..

29 records · Page 2Linked to original sources

bioRxiv2022

Rock inhibitors target SRSF2 leukemia by disrupting cell mitosis and nuclear morphology

Spliceosome machinery mutations are common early mutations in myeloid malignancies, however effective targeted therapies against them are still lacking. In the current study, we used an in vitro high-throughput drug screen among four different isogenic cell lines and identified ROCK inhibitors (ROCKi) as selective inhibitors of SRSF2 mutants. ROCKi targeted SRSF2 Mut primary human samples in a xenografts model and were not toxic to mice nor human cells. ROCKi induced mitotic catastrophe through their apparent effects on microtubules and nuclear organization. Transmission electron microscopy revealed that SRSF2 mutations induce deep nuclear indentation and segmentation, driven by microtubule-rich cytoplasmic intrusions, which were exacerbated by ROCKi. The severe nuclear deformation driven by the combination of SRSF2 Mut and ROCKi prevent cells from completing mitosis. These findings shed light on new ways to target SRSF2 and on the role of the microtubule system in SRSF2 Mut cells.

cancer biology↗

bioRxiv2022

High Hedgehog signaling is transduced by a multikinase-dependent switch controling the apico-basal distribution of the GPCR Smoothened.

The oncogenic GPCR Smoothened is a key transducer of the Hedgehog morphogen, which plays essential roles in the patterning of epithelial structures. Here, we examine how Hedgehog controls Smoothened subcellular localization and activity in a polarized epithelium using the Drosophila wing imaginal disc as a model. We provide evidence that Hedgehog promotes the stabilization of Smoothened by switching its fate after endocytosis toward recycling. This effect involves the sequential and additive action of Protein Kinase A, Casein Kinase I and the Fused kinase. Moreover, in the presence of very high levels of Hedgehog, a second effect of Fused leads to the local enrichment of Smoothened in the most basal domain of the cell membrane. Together, these results link the morphogenetic effects of Hedgehog to the apico-basal distribution of Smoothened and provide a novel mechanism for regulation of a GPCR by plasma membrane subcompartimentalisation.

developmental biology↗

bioRxiv2022

YAP restricts renal inflammation and mitigates kidney damage in nephronothisis related kidney disease.

Nephronophthisis (NPH) is an orphan recessive kidney disease mostly caused by mutations in NPHP1 and 20 other genes encoding proteins that localize to primary cilia. To date the pathways linking altered primary cilia function to progressive kidney scarring in NPH remain poorly defined and therapeutic options allowing NPH patients to escape end-stage kidney disease are lacking. Distinct proteins mutated in NPH interact with components of the Hippo pathway, an important regulator of cell fate. YAP (Yes-associated protein) overactivation has been shown to induce renal scarring while YAP inhibition showed protective effect in kidney diseases unrelated to NPH. Yet, the therapeutic potential of YAP inhibition in NPH has not been formerly assessed. Here we studied the impact of both genetic and pharmacologic YAP inhibition on the NPH-like phenotype caused by a bi-allelic mutation of Lkb1, a ciliary kinase interacting with NPHP1. Contrary to non NPH renal disease, our results reveal an unexpected protective role of YAP in Lkb1 mutant kidneys. Indeed, YAP genetic disruption drastically increase kidney disease burden in Lkb1 deficient mice, while pharmacologic inhibition of YAP failed to improve their phenotype. Collectively these results suggest that YAP inhibition is not a valid therapeutic strategy in NPH and suggest that LKB1 and YAP are parallel negative regulators of a yet uncharacterized pathway detrimental for kidney health.

pathology↗

bioRxiv2021

The first Miocene fossils from coastal woodlands in the southern East African Rift

The Miocene is a key time in the evolution of African mammals and their ecosystems witnessing the origin of the African apes and the isolation of eastern coastal forests through an expanding biogeographic arid corridor. Until recently, however, Miocene sites from the southeastern regions of the continent were unknown. Here we report discovery of the first Miocene fossil teeth from the shoulders of the Urema Rift in Gorongosa National Park, Mozambique, at the southern East African Rift System. We provide the first 1) radiometric age determinations of the fossiliferous Mazamba Formation, 2) reconstructions of past vegetation in the region based on pedogenic carbonates and fossil wood, and 3) description of fossil teeth from the southern rift. Gorongosa is unique in the East African Rift System in combining marine invertebrates, marine vertebrates, terrestrial mammals, and fossil woods in coastal paleoenvironments. The Gorongosa fossil sites offer the first evidence of persistent woodlands and forests on the coastal margins of southeastern Africa during the Miocene, and an exceptional assemblage of fossil vertebrates including new species. Further work will allow the testing of hypotheses positing the formation of a northeast-southwest arid corridor isolating species on the eastern coastal forests from those elsewhere in Africa. BriefThe Miocene is a key time in the evolution of African mammals and their ecosystems encompassing hominine origins and the establishment of an arid corridor that isolated eastern Africas coastal forests. Until now, however, Miocene sites from southeastern Africa have been unknown. We report the discovery of the first Miocene fossil sites from Gorongosa National Park, Mozambique, and show that these sites formed in coastal settings. We provide radiometric ages for the fossiliferous sediments, reconstructions of past vegetation based on stable isotopes and fossil wood, and a description of the first fossil teeth from the region. Gorongosa is the only paleontological site in the East African Rift that combines fossil woods, marine invertebrates, marine vertebrates, and terrestrial mammals. Gorongosa offers the first evidence of persistent woodlands and forests on the coastal margins of southeastern Africa during the Miocene.

paleontology↗

bioRxiv2021

Landscaping the behavioural ecology of primate stone tool use

Ecology is fundamental to the development, transmission, and perpetuity of primate technology. Previous studies on tool site selection have addressed the relevance of targeted resources and raw materials for tools, but few have considered the broader foraging landscape. In this first landscape-scale study of the ecological contexts of wild chimpanzee (Pan troglodytes verus) tool-use, we investigate the conditions required for nut-cracking to occur and persist over time at discrete locations in Bossou (Guinea). We examine this at three levels: selection, frequency of use, and inactivity. We find that, further to the presence of a nut tree and availability of raw materials, abundance of food-providing trees as well as proximity to nest sites were significant predictors of nut-cracking occurrence. This suggests that the spatial distribution of nut-cracking sites is mediated by the broader behavioural landscape and is influenced by non-extractive foraging of predictable resources, as well as non-foraging activities. Additionally, the number of functional tools was greater at sites with higher frequency of nut-cracking and was negatively correlated with site inactivity. Our findings indicate that the technological landscape of the Bossou chimpanzees shares affinities with the favoured places model of hominin site formation and provides new insights for reconstructing ancient patterns of landscape use. ResumeLecologie est fondamentale pour le developpement, la transmission et la perennite de la technologie des primates. Des etudes anterieures ont identifie la disponibilite des ressources cibles ainsi que les matieres premieres pour les outils comme des facteurs influents dans la selection des emplacements pour les activites technologiques. Cependant, il y a peu detudes qui abordent cette recherche a lechelle du paysage et du comportement fourrager. Dans cette premiere etude paysagere sur lutilisation doutils par le chimpanze sauvage (Pan troglodytes verus), nous recherchons les conditions ecologiques qui influencent la selection, lutilisation et linactivite des emplacements utilises pour le cassage des noix en Bossou, Guinee. Nos resultats montrent quen plus de la presence dun noyer et de la disponibilite des matieres premieres, labondance darbres nourriciers ainsi que la proximite des sites de nidification etaient des predicteurs significatifs de loccurrence du cassage des noix. Cela suggere que la distribution spatiale des sites de cassage de noix est influencee par le paysage comportemental et est influencee par le fourrage non-extractive de ressources previsibles, ainsi que par des activites non-fourragers. Nos resultats indiquent que le paysage technologique des chimpanzes de Bossou partage des affinites avec le modele des << lieux favoris >> de la formation des sites hominides et fournit de nouvelles perspectives pour reconstruire les modes dutilisation du paysage anciens.

ecology↗

bioRxiv2021

Adsorption of Lysozyme Into a Charged Confining Pore

Several applications arise from the confinement of proteins on surfaces since their stability and biological activity are enhanced. It is also known that the way a protein adsorbs on the surface is important for its biological function since its active sites should not be obstructed. In this study, the adsorption properties of hen egg-white Lysozyme, HEWL, into a negatively charged silica pore is examined employing a coarse-grained model and constant-pH Monte Carlo simulations. The role of electrostatic interactions is taken into account when including the Debye-HAijckel potentials into the C structure-based model. We evaluate the effects of pH, salt concentration, and pore radius on the protein preferential orientation and spatial distribution of its residues regarding the pore surface. By mapping the residues that stay closer to the pore surface, we find the increase of pH leads to orientational changes of the adsorbed protein when the solution pH gets closer to the HEWL isoelectric point. At these conditions, the pKa shift of these important residues caused by the adsorption into the charged confining surface results in a HEWL charge distribution that stabilizes the adsorption in the observed protein orientation. We compare our observations to the results of pKa shift for HEWL available in the literature and to some experimental data.

biophysics↗

bioRxiv2021

RAMPs regulate signalling bias and internalisation of the GIPR

Gastric inhibitory polypeptide (GIP) receptor is a class B1 GPCR, that responds to GIP and physiologically potentiates glucose-stimulated insulin secretion. Like most class B1 GPCRs, GIPR has been shown to interact with RAMPs, yet the effects of RAMPs on its signalling and trafficking remain poorly understood. We demonstrate that RAMPs modulate G protein activation and GIPR internalisation profiles. RAMP3 reduced GIPR Gs activation and cAMP production but retained GIPR at the cell surface, and this was associated with prolonged ERK1/2 phosphorylation and {beta}-arrestin association. By contrast, RAMP1/2 reduced Gq/11/15 activation of the GIPR. Through knockout mice studies, we show that RAMP1 is important to the normal physiological functioning of GIPR to regulate blood glucose levels. Thus, RAMPs act on G protein/{beta}-arrestin complexes, having both acute and chronic effects on GIPR function, while this study also raises the possibility of a more general role of RAMP3 to enhance GPCR plasma membrane localisation.

pharmacology and toxicology↗

bioRxiv2020

DNA damage independent inhibition of NF-κB transcription by anthracyclines

Anthracyclines are among the most used and effective anticancer drugs. Their activity has been attributed to DNA double-strand breaks resulting from topoisomerase II poisoning and to eviction of histones from select sites in the genome. Here we show that the extensively used anthracyclines Doxorubicin, Daunorubicin and Epirubicin, decrease the transcription of nuclear factor kappa B (NF-{kappa}B)-dependent gene targets, but not interferon responsive genes. Using an NMR-based structural approach, we demonstrate that anthracyclines disturb the complexes formed between the NF-{kappa}B subunit RelA and its DNA binding sites. The variant anthracyclines Aclarubicin, Doxorubicinone and the newly developed Dimethyl-doxorubicin, which share anticancer properties with the other anthracyclines but do not induce DNA damage, also suppressed inflammation, thus uncoupling DNA damage from the effects on inflammation. These findings have implications for anticancer therapy and for the development of novel anti-inflammatory drugs with limited side effects for life-threatening conditions such as sepsis.

immunology↗

bioRxiv2017

Partial selfing eliminates inbreeding depression while maintaining genetic diversity

Classical theory on the origin and evolution of selfing and outcrossing relies on the role of inbreeding depression created by unlinked partially-deleterious recessive alleles to predict that individuals from natural populations predominantly self or outcross. Comparative data indicates, however, that maintenance of partial selfing and outcrossing at intermediate frequencies is common in nature. In part to explain the presence of mixed reproductive modes within populations, several hypotheses regarding the evolution of inbreeding depression have been put forward based on the complex interaction of linkage and identity disequilibrium among fitness loci, together with Hill-Robertson effects. We here ask what is the genetic basis of inbreeding depression so that populations with intermediate selfing rates can eliminate it while maintain potentially adaptive genetic diversity. For this, we use experimental evolution in the nematode C. elegans under partial selfing and compare it to the experimental evolution of populations evolved under exclusive selfing and predominant outcrossing. We find that the ancestral risk of extinction upon enforced inbreeding by selfing is maintained when populations evolve under predominant outcrossing, but reduced when populations evolve under partial or exclusive selfing. Analysis of genome-wide single-nucleotide polymorphism (SNP) during experimental evolution and after enforced inbreeding suggests that, under partial selfing, populations were purged of unlinked deleterious recessive alleles that segregate in the ancestral population, which in turn allowed the expression of unlinked overdominant fitness loci. Taken together, these observations indicate that populations evolving under partial selfing gain the short-term benefits of selfing, in purging deleterious recessive alleles, but also the long-term benefits of outcrossing, in maintaining genetic diversity that may important for future adaptation.

evolutionary biology↗

bioRxiv2017

Overcoming insecticide resistance through computational inhibitor design

Insecticides allow control of agricultural pests and disease vectors and are vital for global food security and health. The evolution of resistance to insecticides, such as organophosphates (OPs), is a serious and growing concern. OP resistance often involves sequestration or hydrolysis of OPs by carboxylesterases. Inhibiting carboxylesterases could therefore restore the effectiveness of OPs for which resistance has evolved. Here, we use covalent computational design to produce nano/pico-molar boronic acid inhibitors of the carboxylesterase E7 from the agricultural pest Lucilia cuprina, as well as a common Gly137Asp E7 mutant that confers OP resistance. These inhibitors, with high selectivity against human acetylcholinesterase, and low to no toxicity in human cells and mice, act synergistically with the OPs diazinon and malathion to reduce the amount of OP required to kill L. cuprina by up to 16-fold, and abolish resistance. The compounds exhibit broad utility in significantly potentiating another OP, chlorpyrifos against the common pest, the peach-potato aphid (Myzus persicae). These compounds represent a solution to OP resistance as well as to environmental concerns regarding overuse of OPs, allowing significant reduction of use without compromising efficacy.

biochemistry↗

bioRxiv2017

Polygenicity And Epistasis Underlie Fitness-Proximal Traits In The Caenorhabditis elegans Multiparental Experimental Evolution (CeMEE) Panel

Understanding the genetic basis of complex traits remains a major challenge in biology. Polygenicity, phenotypic plasticity and epistasis contribute to phenotypic variance in ways that are rarely clear. This uncertainty is problematic for estimating heritability, for predicting individual phenotypes from genomic data, and for parameterizing models of phenotypic evolution. Here we report a recombinant inbred line (RIL) quantitative trait locus (QTL) mapping panel for the hermaphroditic nematode Caenorhabditis elegans, the C. elegans multiparental experimental evolution (CeMEE) panel. The CeMEE panel, comprising 507 RILs, was created by hybridization of 16 wild isolates, experimental evolution at moderate population sizes and predominant outcrossing for 140-190 generations, and inbreeding by selfing for 13-16 generations. The panel contains 22% of single nucleotide polymorphisms known to segregate in natural populations, and complements existing mapping resources for C. elegans by providing high nucleotide diversity across >95% of the genome. We apply it to study the genetic basis of two fitness components, fertility and hermaphrodite body size at time of reproduction, with high broad sense heritability in the CeMEE. While simulations show we should detect common alleles with additive effects as small as 5%, at gene-level resolution, the genetic architectures of these traits does not feature such alleles. We instead find that a significant fraction of trait variance, particularly for fertility, can be explained by sign epistasis with weak main effects. In congruence, phenotype prediction, while generally poor (r2 < 10%), requires modeling epistasis for optimal accuracy, with most variance attributed to the highly recombinant, rapidly evolving chromosome arms.

evolutionary biology↗