Formation Of Lineages Research Articles

Polyploidy Expands the Range of Centaurium (Gentianaceae).

The Mediterranean region is one of the most important worldwide hotspots in terms of number of species and endemism, and multiple hypotheses have been proposed to explain how diversification occurred in this area. The contribution of different traits to the diversification process has been evaluated in different groups of plants. In the case of Centaurium (Gentianaceae), a genus with a center of diversity placed in the Mediterranean region, polyploidy seems to have been an important driver of diversification as more than half of species are polyploids. Moreover, ploidy levels are strongly geographically structured across the range of the genus, with tetraploids distributed towards more temperate areas in the north and hexaploids in more arid areas towards the south. We hypothesize that the diversification processes and biodiversity patterns in Centaurium are explained by the coupled formation of polyploid lineages and the colonization of different areas. A MCC tree from BEAST2 based on three nuclear DNA regions of a total of 26 taxa (full sampling, of 18 species and 8 subspecies) was used to perform ancestral area reconstruction analysis in “BioGeoBEARS.” Chromosome evolution was analyzed in chromEvol and diversification in BAMM to estimate diversification rates. Our results suggest that two major clades diverged early from the common ancestor, one most likely in the western Mediterranean and the other in a widespread area including west and central Asia (but with high uncertainty in the exact composition of this widespread area). Most ancestral lineages in the western clade remained in or around the western Mediterranean, and dispersal to other areas (mainly northward and eastward), occurred at the tips. Contrarily, most ancestral lineages in the widespread clade had larger ancestral areas. Polyploidization events in the western clade occurred at the tips of the phylogeny (with one exception of a polyploidization event in a very shallow node) and were mainly tetraploid, while polyploidization events occurred in the widespread clade were at the tips and in an ancestral node of the phylogeny, and were mainly hexaploid. We show how ancestral diploid lineages remained in the area of origin, whereas recent and ancestral polyploidization could have facilitated colonization and establishment in other areas.

Quaternary climatic fluctuations influence the demographic history of two species of sky-island endemic amphibians in the Neotropics

We evaluated the role of Quaternary climatic fluctuations on the demographic history and population structure of amphibian species endemic to the ‘campo rupestre’ in the Neotropics, evaluating their distributional shifts, demographic changes, and lineage formation from the end of Pleistocene to present. We chose two anurans endemic to the high-elevation ‘campo rupestre’ in the Espinhaço Range (ER) in northeastern and southeastern Brazil (Bokermannohyla alvarengai and Bokermannohyla oxente), as models to test the role of Quaternary climatic fluctuations over their distribution range in this region. We collected tissue samples throughout their distribution range and used statistical phylogeography to examine processes of divergence and population demography. We generated spatial–temporal reconstructions using Bayesian inference in a coalescent framework in combination with hind-cast projections of species distribution models (SDMs). We also used the results and literature information to test alternative diversification scenarios via approximate Bayesian computation (ABC). Our results show that Quaternary climatic fluctuations influenced the geographic ranges of both species showing population expansion during the last glacial maximum (LGM) and range contraction during interglacial periods, as inferred from selected ABC models and from past projections of SDMs. We recovered Pleistocene diversification for both species occuring in distinctly unique periods for each taxon. An older and range-restricted lineage was recovered in a geographically isolated geological massif, deserving conservation and further taxonomic study. The diversification and distribution of these amphibian species endemic to the Neotropical ‘campo rupestre’ were influenced by Quaternary climatic fluctuations. The expansion of cold adapted species restricted to higher elevations during glacial periods and their concomitant retraction during interglacial periods may have been crucial for producing patterns of species richness and endemism along elevation gradients in tropical and subtropical domains. Such processes may influence the evolution of the biota distributed in heterogeneous landscapes with varied topography.

The versican-hyaluronan complex provides an essential extracellular matrix niche for Flk1+ hematoendothelial progenitors

Little is known about extracellular matrix (ECM) contributions to formation of the earliest cell lineages in the embryo. Here, we show that the proteoglycan versican and glycosaminoglycan hyaluronan are associated with emerging Flk1+ hematoendothelial progenitors at gastrulation. The mouse versican mutant Vcanhdf lacks yolk sac vasculature, with attenuated yolk sac hematopoiesis. CRISPR/Cas9-mediated Vcan inactivation in mouse embryonic stem cells reduced vascular endothelial and hematopoietic differentiation within embryoid bodies, which generated fewer blood colonies, and had an impaired angiogenic response to VEGF165. Hyaluronan was severely depleted in Vcanhdf embryos, with corresponding upregulation of the hyaluronan-depolymerase TMEM2. Conversely, hyaluronan-deficient mouse embryos also had vasculogenic suppression but with increased versican proteolysis. VEGF165 and Indian hedgehog, crucial vasculogenic factors, utilized the versican-hyaluronan matrix, specifically versican chondroitin sulfate chains, for binding. Versican-hyaluronan ECM is thus an obligate requirement for vasculogenesis and primitive hematopoiesis, providing a vasculogenic factor-enriching microniche for Flk1+ progenitors from their origin at gastrulation.

Chromatin regulatory dynamics of early human small intestinal development using a directed differentiation model.

The establishment of the small intestinal (SI) lineage during human embryogenesis ensures functional integrity of the intestine after birth. The chromatin dynamics that drive SI lineage formation and regional patterning in humans are essentially unknown. To fill this knowledge void, we apply a cutting-edge genomic technology to a state-of-the-art human model of early SI development. Specifically, we leverage chromatin run-on sequencing (ChRO-seq) to define the landscape of active promoters, enhancers and gene bodies across distinct stages of directed differentiation of human pluripotent stem cells into SI spheroids with regional specification. Through comprehensive ChRO-seq analysis we identify candidate stage-specific chromatin activity states, novel markers and enhancer hotspots during the directed differentiation. Moreover, we propose a detailed transcriptional network associated with SI lineage formation or regional patterning. Our ChRO-seq analyses uncover a previously undescribed pattern of enhancer activity and transcription at HOX gene loci underlying SI regional patterning. We also validated this unique HOX dynamics by the analysis of single cell RNA-seq data from human fetal SI. Overall, the results lead to a new proposed working model for the regulatory underpinnings of human SI development, thereby adding a novel dimension to the literature that has relied almost exclusively on non-human models.

Resolving spatial complexities of hybridization in the context of the gray zone of speciation in North American ratsnakes (Pantherophis obsoletus complex).

Inferring the history of divergence between species in a framework that permits the presence of gene flow has been crucial for characterizing the "gray zone" of speciation, which is the period of time where lineages have diverged but have not yet achieved strict reproductive isolation. However, estimates of both divergence times and rates of gene flow often ignore spatial information, for example when considering the location and width of hybrid zones with respect to changes in the environment between lineages. Using population genomic data from the North American ratsnake complex (Pantherophis obsoletus), we connected phylogeographic estimates of lineage structure, migration, historical demography, and timing of divergence with hybrid zone dynamics. We examined the spatial context of diversification by linking migration and timing of divergence to the location and widths of hybrid zones. Artificial neural network approaches were applied to understand how landscape features and past climate have influenced population genetic structure among these lineages. We found that rates of migration between lineages were associated with the overall width of hybrid zones. Timing of divergence was not related to migration rate or hybrid zone width across species pairs but may be related to the number of alleles weakly introgressing through hybrid zones. This research underscores how incomplete reproductive isolation can be better understood by considering differential allelic introgression and the effects of historical and contemporary landscape features on the formation of lineages as well as overall genomic estimates of migration rates through time.

Sol\u2013gel based synthesis and biological properties of zinc integrated nano bioglass ceramics for bone tissue regeneration

Bone is a flexible and electro active tissue that is vulnerable to various traumatic injuries. The self-healing of damaged bone tissue towards reconstruction is limited due to the lack of proper niche compliances. Nevertheless, the classical grafting techniques like autograft/allograft for bone repair pose challenges like bacterial infections and donor-site morbidity with unsatisfactory outcomes. The use of appropriate biomaterial with osteogenic potential can meet these challenges. In this regard, bioactive glass ceramics is widely used as a bone filler or graft material because of its bonding affinity to bone leading towards bone reconstruction applications without the challenge of post implant infections. Hence, the current study is aimed at addressing this potentiality of zinc (Zn) for doped the bioglass at nano-scale advantages for bone tissue repair. Since, Zn has been demonstrated to have not only antibacterial property but also the stimulatory effect on osteoblasts differentiation, mineralization by enhancing the osteogenic genes expression. In view of these, the present study is focused on sol–gel synthesis and pysico-chemical characterization of Zinc-doped bioglass nanoparticles (Zn-nBGC) and also analyzing its biological implications. The surface morphological and physiochemical characterizations using SEM, EDX, FT-IR and XRD analysis has shown the increased surface area of Zn-nBGC particles providing a great platform for biomolecular interaction, cytocompatibility, cell proliferation and osteogenic differentiation. The obtaining hydroxy apatite groups have initiated in vitro mineralization towards osteogenic lineage formation. Zn has not only involved in enhancing cellular actions but also strengthen the ceramic nanoparticles towards antibacterial application. Hence the finding suggests a biomaterial synthesis of better biomaterial for bone tissue engineering application by preventing post-operative bacterial infection.

The Evolution of Sex is Tempered by Costly Hybridization in Boechera (Rock Cress).

Despite decades of research, the evolution of sex remains an enigma in evolutionary biology. Typically, research addresses the costs of sex and asexuality to characterize the circumstances favoring one reproductive mode. Surprisingly few studies address the influence of common traits that are, in many organisms, obligately correlated with asexuality, including hybridization and polyploidy. These characteristics have substantial impacts on traits under selection. In particular, the fitness consequences of hybridization (i.e., reduced fitness due to interspecific reproductive isolation) will influence the evolution of sex. This may comprise a cost of either sex or asexuality due to the link between hybridity and asexuality. We examined reproductive isolation in the formation of de novo hybrid lineages between 2 widespread species in the ecological model system Boechera. Seventeen percent of 664 crosses produced F1 fruits, and only 10% of these were viable, suggesting that postmating prezygotic and postzygotic barriers inhibit hybrid success in this system. The postmating prezygotic barrier was asymmetrical, with 110 of 115 total F1 fruits produced when Boechera stricta acted as maternal parent. This asymmetry was confirmed in wild-collected lineages, using a chloroplast phylogeny of wild-collected B. stricta, Boechera retrofracta, and hybrids. We next compared fitness of F2 hybrids and selfed parental B. stricta lines, finding that F2 fitness was reduced by substantial hybrid sterility. Multiple reproductively isolating barriers influence the formation and fitness of hybrid lineages in the wild, and the costs of hybridization likely have profound impacts on the evolution of sex in the natural environment.

The BET Inhibitor, CPI-0610, Promotes Myeloid Differentiation in Myelofibrosis Patient Bone Marrow and Peripheral CD34+ Hematopoietic Stem Cells

The BET Inhibitor, CPI-0610, Promotes Myeloid Differentiation in Myelofibrosis Patient Bone Marrow and Peripheral CD34+ Hematopoietic Stem Cells

Phylogenetic analysis of SARS-CoV-2 in the first few months since its emergence.

During the first few months of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) evolution in a new host, contrasting hypotheses have been proposed about the way the virus has evolved and diversified worldwide. The aim of this study was to perform a comprehensive evolutionary analysis to describe the human outbreak and the evolutionary rate of different genomic regions of SARS‐CoV‐2. The molecular evolution in nine genomic regions of SARS‐CoV‐2 was analyzed using three different approaches: phylogenetic signal assessment, emergence of amino acid substitutions, and Bayesian evolutionary rate estimation in eight successive fortnights since the virus emergence. All observed phylogenetic signals were very low and tree topologies were in agreement with those signals. However, after 4 months of evolution, it was possible to identify regions revealing an incipient viral lineage formation, despite the low phylogenetic signal since fortnight 3. Finally, the SARS‐CoV‐2 evolutionary rate for regions nsp3 and S, the ones presenting greater variability, was estimated as 1.37 × 10−3 and 2.19 × 10−3 substitution/site/year, respectively. In conclusion, results from this study about the variable diversity of crucial viral regions and determination of the evolutionary rate are consequently decisive to understand essential features of viral emergence. In turn, findings may allow the first‐time characterization of the evolutionary rate of S protein, crucial for vaccine development.

Wwc2 Is a Novel Cell Division Regulator During Preimplantation Mouse Embryo Lineage Formation and Oogenesis.

Formation of the hatching mouse blastocyst marks the end of preimplantation development, whereby previous cell cleavages culminate in the formation of three distinct cell lineages (trophectoderm, primitive endoderm and epiblast). We report that dysregulated expression of Wwc2, a genetic paralog of Kibra/Wwc1 (a known activator of Hippo-signaling, a key pathway during preimplantation development), is specifically associated with cell autonomous deficits in embryo cell number and cell division abnormalities. Division phenotypes are also observed during mouse oocyte meiotic maturation, as Wwc2 dysregulation blocks progression to the stage of meiosis II metaphase (MII) arrest and is associated with spindle defects and failed Aurora-A kinase (AURKA) activation. Oocyte and embryo cell division defects, each occurring in the absence of centrosomes, are fully reversible by expression of recombinant HA-epitope tagged WWC2, restoring activated oocyte AURKA levels. Additionally, clonal embryonic dysregulation implicates Wwc2 in maintaining the pluripotent epiblast lineage. Thus, Wwc2 is a novel regulator of meiotic and early mitotic cell divisions, and mouse blastocyst cell fate.

GnRH neurogenesis depends on embryonic pheromone receptor expression

Gonadotropin-releasing hormone (GnRH) neurons control mammalian reproduction and migrate from their birthplace in the nasal placode to the hypothalamus during development. Despite much work on the origin and migration of GnRH neurons, the processes that control GnRH lineage formation are not fully understood. Here, we demonstrate that Nhlh genes control vomeronasal receptor expression in the developing murine olfactory placode associated with the generation of the first GnRH neurons at embryonic days (E)10–12. Inactivation of ß2-microglobulin (ß2-m), which selectively affects surface expression of V2Rs, dramatically decreased the number of GnRH neurons in the Nhlh2 mutant background, preventing rescue of fertility in female Nhlh2 mutant mice by male pheromones. In addition, we show that GnRH neurons generated after E12 fail to establish synaptic connections to the vomeronasal amygdala, suggesting the existence of functionally specialized subpopulations of GnRH neurons, which process pheromonal information.

Metabolite Patterns in Human Myeloid Hematopoiesis Result from Lineage-Dependent Active Metabolic Pathways.

Assessment of hematotoxicity from environmental or xenobiotic compounds is of notable interest and is frequently assessed via the colony forming unit (CFU) assay. Identification of the mode of action of single compounds is of further interest, as this often enables transfer of results across different tissues and compounds. Metabolomics displays one promising approach for such identification, nevertheless, suitability with current protocols is restricted. Here, we combined a hematopoietic stem and progenitor cell (HSPC) expansion approach with distinct lineage differentiations, resulting in formation of erythrocytes, dendritic cells and neutrophils. We examined the unique combination of pathway activity in glycolysis, glutaminolysis, polyamine synthesis, fatty acid oxidation and synthesis, as well as glycerophospholipid and sphingolipid metabolism. We further assessed their interconnections and essentialness for each lineage formation. By this, we provide further insights into active metabolic pathways during the differentiation of HSPC into different lineages, enabling profound understanding of possible metabolic changes in each lineage caused by exogenous compounds.

3144 – ECTOPIC EXPRESSION OF THE HOMEOBOX GENE MNX1 IS THE TRANSFORMING EVENT IN A MOUSE MODEL OF PEDIATRIC T(7;12)(Q36;P13) PEDIATRIC ACUTE MYELOID LEUKEMIA

Pediatric acute myeloid leukemia (AML) with the translocation t(7;12)(q36;p13) is a form of childhood leukemia associated with inferior outcome. In the current investigation we have studied the gene alterations that are induced by this chromosomal translocation, a gene fusion MNX1-ETV6 but also ectopic expression of the homeobox gene MNX1. In a murine bone marrow transplantation model, our results showed that ectopic expression of MNX1 was able to induce AML, but not the MNX1-ETV6 gene fusion. However, this transformation to AML was only seen using hematopoietic progenitor cells from fetal liver and not adult bone marrow cells. This was accompanied with an alteration of cell differentiation towards myeloid lineage, increased proliferation and colony formation capacity induced by MNX1. Our data also showed that ectopic expression of MNX1, and to a lower extend MNX1-ETV6 fusion, induced DNA damage and cell cycle arrest, where we saw an increased expression and binding of the MNX1 and the MNX1-ETV6 fusion to the gene CEP164 that has a critical role in cell cycle regulation and DNA damage response, and the transcriptional pioneer factor gene PBX1 that is important in development and differentiation. Furthermore, global histone modifications, specifically H3K4me3 and H3K27me, were induced by MNX1 but also the MNX1-ETV6 fusion. We could also see an enrichment of methylation/methionine proteins from mass spectrometry analysis of proteins associated with MNX1. In conclusion, these results suggest a possible mechanism by which MNX1 mediates its transforming effect through abnormal methylation of proteins that result in changed histone modifications and subsequent dysregulation of gene expression. Our finding that only fetal liver cells were able to transform into AML is also in concordance with the clinical finding that this AML subtype is only found in very young children.

Common principles of early mammalian embryo self-organisation.

Pre-implantation mammalian development unites extreme plasticity with a robust outcome: the formation of a blastocyst, an organised multi-layered structure ready for implantation. The process of blastocyst formation is one of the best-known examples of self-organisation. The first three cell lineages in mammalian development specify and arrange themselves during the morphogenic process based on cell-cell interactions. Despite decades of research, the unifying principles driving early mammalian development are still not fully defined. Here, we discuss the role of physical forces, and molecular and cellular mechanisms, in driving self-organisation and lineage formation that are shared between eutherian mammals.

A tale of two cell-fates: role of the Hippo signaling pathway and transcription factors in early lineage formation in mouse preimplantation embryos.

In mammals, the first cell-fate decision occurs during preimplantation embryo development when the inner cell mass (ICM) and trophectoderm (TE) lineages are established. The ICM develops into the embryo proper, while the TE lineage forms the placenta. The underlying molecular mechanisms that govern lineage formation involve cell-to-cell interactions, cell polarization, cell signaling and transcriptional regulation. In this review, we will discuss the current understanding regarding the cellular and molecular events that regulate lineage formation in mouse preimplantation embryos with an emphasis on cell polarity and the Hippo signaling pathway. Moreover, we will provide an overview on some of the molecular tools that are used to manipulate the Hippo pathway and study cell-fate decisions in early embryos. Lastly, we will provide exciting future perspectives on transcriptional regulatory mechanisms that modulate the activity of the Hippo pathway in preimplantation embryos to ensure robust lineage segregation.

Evolution by means of natural selection without reproduction: revamping Lewontin’s account

This paper analyzes recent attempts to reject reproduction with lineage formation as a necessary condition for evolution by means of natural selection (Bouchard in Philos Sci 75(5):560–570, 2008; Stud Hist Philos Sci Part C Stud Hist Philos Biol Biomed Sci 42(1):106–114, 2011; Bourrat in Biol Philos 29(4):517–538, 2014; Br J Philos Sci 66(4):883–903, 2015; Charbonneau in Philos Sci 81(5):727–740, 2014; Doolittle and Inkpen in Proc Natl Acad Sci 115(16):4006–4014, 2018). Building on the strengths of these attempts and avoiding their pitfalls, it is argued that a robust formulation of evolution by natural selection without reproduction can be established. The main contribution of this paper is a reformulation of Lewontin’s three principles (Lewontin in Annu Rev Ecol Syst 1:1–18, 1970) stating that minimal evolution by natural selection occurs when two conditions are met in a population: fitness-related variation and memory (population-level inheritance). Paradigmatic evolution by natural selection, which can generate adaptations, takes place when an additional condition is present, namely regeneration.

PRMT1-p53 Pathway Controls Epicardial EMT and Invasion

Epicardial cells are cardiac progenitors that give rise to the majority of cardiac fibroblasts, coronary smooth muscle cells, and pericytes during development. An integral phase of epicardial fate transition is epithelial-to-mesenchymal transition (EMT) that confers motility. We uncover an essential role for the protein arginine methyltransferase 1 (PRMT1) in epicardial invasion and differentiation. Using scRNA-seq, we show that epicardial-specific deletion of Prmt1 reduced matrix and ribosomal gene expression in epicardial-derived cell lineages. PRMT1 regulates splicing of Mdm4, which is a key controller of p53 stability. Loss of PRMT1 leads to accumulation of p53 that enhances Slug degradation and blocks EMT. During heart development, the PRMT1-p53 pathway is required for epicardial invasion and formation of epicardial-derived lineages: cardiac fibroblasts, coronary smooth muscle cells, and pericytes. Consequently, this pathway modulates ventricular morphogenesis and coronary vessel formation. Altogether, our study reveals molecular mechanisms involving the PRMT1-p53 pathway and establish its roles in heart development.

Evidence for Overlapping and Distinct Biological Activities and Transcriptional Targets Triggered by Fibroblast Growth Factor Receptor 2b Signaling between Mid- and Early Pseudoglandular Stages of Mouse Lung Development.

Branching morphogenesis is the basic developmental mode common to organs such as the lungs that undergo a process of ramification from a rudimentary tree. However, the precise molecular and cellular bases underlying the formation of branching organs are still unclear. As inactivation of fibroblast growth factor receptor 2b (Fgfr2b) signaling during early development leads to lung agenesis, thereby preventing the analysis of this pathway at later developmental stages, we used transgenic mice to induce expression of a soluble form of Fgfr2b to inactivate Fgfr2b ligands at embryonic day (E) 14.5, corresponding to the mid-pseudoglandular stage of lung development. We identified an Fgfr2b signaling signature comprised of 46 genes enriched in the epithelium, some of which were common to, but most of them distinct from, the previously identified Fgfr2b signaling signature at E12.5. Our results indicate that Fgfr2b signaling at E14.5 controls mostly proliferation and alveolar type 2 cell (AT2) differentiation. In addition, inhibition of Fgfr2b signaling at E14.5 leads to morphological and cellular impairment at E18.5, with defective alveolar lineage formation. Further studies will have to be conducted to elucidate the role of Fgfr2b signaling at successive stages (canalicular/saccular/alveolar) of lung development as well as during homeostasis and regeneration and repair after injury.

Using asexual vertebrates to study genome evolution and animal physiology: Banded (Fundulus diaphanus)xCommon Killifish (F.heteroclitus) hybrid lineages as a model system.

Wild, asexual, vertebrate hybrids have many characteristics that make them good model systems for studying how genomes evolve and epigenetic modifications influence animal physiology. In particular, the formation of asexual hybrid lineages is a form of reproductive incompatibility, but we know little about the genetic and genomic mechanisms by which this mode of reproductive isolation proceeds in animals. Asexual lineages also provide researchers with the ability to produce genetically identical individuals, enabling the study of autonomous epigenetic modifications without the confounds of genetic variation. Here, we briefly review the cellular and molecular mechanisms leading to asexual reproduction in vertebrates and the known genetic and epigenetic consequences of the loss of sex. We then specifically discuss what is known about asexual lineages of Fundulus diaphanus x F. heteroclitus to highlight gaps in our knowledge of the biology of these clones. Our preliminary studies of F. diaphanus and F. heteroclitus karyotypes from Porter's Lake (Nova Scotia, Canada) agree with data from other populations, suggesting a conserved interspecific chromosomal arrangement. In addition, genetic analyses suggest that: (a) the same major clonal lineage (Clone A) of F. diaphanus x F. heteroclitus has remained dominant over the past decade, (b) some minor clones have also persisted, (c) new clones may have recently formed, and iv) wild clones still mainly descend from F. diaphanus ♀ x F. heteroclitus ♂ crosses (96% in 2017–2018). These data suggest that clone formation may be a relatively rare, but continuous process, and there are persistent environmental or genetic factors causing a bias in cross direction. We end by describing our current research on the genomic causes and consequences of a transition to asexuality and the potential physiological consequences of epigenetic variation.

Monarquía y ciudades de realengo en Castilla. Siglos XII-XV

Étude sur les rapports entre le pouvoir royal et ceux des villes qui relevaient directement de la couronne (realengo) à travers quatre siècles (XIIe-XVe) depuis l'époque de la reconquista, en soulignant les grandes réformes de la période 1250-1350: le renforcement du pouvoir des cheva­liers urbains, la naissance des lignages, des partis (bandos) et des assem­blées locales restreintes (regimientos), la préférence des lois royales sur les règlements communales et la présence en ville d'agents de la couronne (alcaldes del rey, corregidores). La deuxième partie de l'étude traite des rapports politiques entre les villes, la royauté et les grands nobles pour remarquer leur influence sur les structures et sur le fonctionnement du gou­vernement des villes pendant l'époque Trastamara (1369-1474) et au cours du règne des Reyes Católicos (1475-1515) qui ont réussi à renforcer dura­blement l'autorité de la couronne sur le modele de pouvoir municipal qui s'était développé au cours du has Mayen Age.