bioRxiv ScienceSearch

Biology subjects

Alfonso Martinez Arias

Publications and source records attributed to Alfonso Martinez Arias.

7 recordsLinked to original sources

Evo-engineering and the Cellular and Molecular Origins of the Vertebrate Spinal Cord

The formation of the spinal cord during early embryonic development in vertebrate embryos is a continuous process that begins at gastrulation and continues through to the completion of somitogenesis. Despite the conserved usage of patterning mechanisms and gene regulatory networks that act to generate specify spinal cord progenitors, there now exists two seemingly disparate models to account for their action. In the first, a posterior localized signalling source transforms previously anterior-specified neural plate into the spinal cord. In the second, a population of bipotent stem cells undergo continuous self-renewal and differentiation to progressively lay down the spinal cord and axial mesoderm by posterior growth. Whether this represents fundamental differences between the experimental model organisms utilised in the generation of these models remains to be addressed. Here we review lineage studies across four key vertebrate models: mouse, chicken, Xenopus and zebrafish and relate this to the underlying gene regulatory networks that are known to be required for spinal cord formation. We propose that by applying a dynamical systems approach to understanding how distinct neural and mesodermal fates arise from a bipotent progenitor pool, it is possible to begin to understand how differences in the dynamical cell behaviours such as proliferation rates and cell movements can map onto conserved regulatory networks to generate diversity in the timing of tissue generation and patterning during development.

Developmental Biology

Interactions between Nodal and Wnt signalling Drive Robust Symmetry Breaking and Axial Organisation in Gastruloids (Embryonic Organoids)

Generation of asymmetry within the early embryo is a critical step in the establishment of the three body axes, providing a reference for the patterning of the organism. To study the establishment of asymmetry and the development of the anteroposterior axis (AP) in culture, we utilised our Gastruloid model system. Gastruloids, highly reproducible embryonic organoids formed from aggregates of mouse embryonic stem cells, display symmetry-breaking, polarised gene expression and axial development, mirroring the processes on a time-scale similar to that of the mouse embyro. Using Gastruloids formed from mouse ESCs containing reporters for Wnt, FGF and Nodal signalling, we were able to quantitatively assess the contribution of these signalling pathways to the establishment of asymmetry through single time-point and live-cell fluorescence microscopy.\n\nDuring the first 24-48h of culture, interactions between the Wnt/{beta}-Catenin and Nodal/TGF{beta} signalling pathways promote the initial symmetry-breaking event, manifested through polarised Brachyury (T/Bra) expression. Neither BMP nor FGF signalling is required for the establishment of asymmetry, however Wnt signalling is essential for the amplification and stability of the initial patterning event. Additionally, low, endogenous levels of FGF (24-48h) has a role in the amplification of the established pattern at later time-points.\n\nOur results confirm that Gastruloids behave like epiblast cells in the embryo, leading us to translate the processes and signalling involved in pattern formation of Gastruloids in culture to the development of the embryo, firmly establishing Gastruloids as a highly reproducible, robust model system for studying cell fate decisions and early pattern formation in culture.

Developmental Biology

FGF/MAPK signaling sets the switching threshold of a bistable circuit controlling cell fate decisions in ES cells

Intracellular transcriptional regulators and extracellular signaling pathways together regulate the allocation of cell fates during development, but how their molecular activities are integrated to establish the correct proportions of cells with particular fates is not known. Here we study this question in the context of the decision between the epiblast (Epi) and the primitive endoderm (PrE) fate that occurs in the mammalian preimplantation embryo. Using an embryonic stem (ES) cell model, we discover two successive functions of FGF/MAPK signaling in this decision. First, the pathway needs to be inhibited to make the PrE-like gene expression program accessible for activation by GATA transcription factors in ES cells. In a second step, MAPK signaling levels determine the threshold concentration of GATA factors required for PrE-like differentiation, and thereby control the proportion of cells differentiating along this lineage. Our findings can be explained by a simple mutual repression circuit modulated by FGF/MAPK signaling. This may be a general network architecture to integrate the activity of signal transduction pathways and transcriptional regulators, and serve to balance proportions of cell fates in several contexts.

Developmental Biology

A framework for the analysis of symmetric and asymmetric divisions in developmental processes

Animal tissue development relies on precise generation and deployment of specific cell types into tissue sub-structures. Understanding how this process is regulated remains a major challenge of biology. In many tissues, development progresses through a sequence of dividing progenitor cells, each with decreasing potency, that balance their growth and differentiation. Dividing progenitor cells thus face a decision on whether their offspring shall differentiate or self-renew. This results in three possible modes of division (symmetric self-renewing, symmetric differentiating, and asymmetric) all of which have been observed in developing animal tissues. In some instances, the frequencies of occurrence of these division modes are incompatible with the possibility that sibling cells take the decision to differentiate independently of each other. Rather, an excess of symmetric divisions, both proliferating and differentiating, is usually observed in so far no general mechanism by which this fate entanglement takes place has been put forward. Here we propose a simple model of progenitor priming that provides a rationale on how the fate of sibling cells might be linked. Analysis of the model suggests that commitment to the cycle completion of cells primed for differentiation might be the cause of the observed excess of symmetric divisions. The model presented is applicable to a broad range of developmental systems and provides a testing framework to explain the dynamics of cell division and differentiation are related.

Developmental Biology

Generation of Aggregates of Mouse ES Cells that Show Symmetry Breaking, Polarisation and Emergent Collective Behaviour in vitro.

Dissociated mouse embryonic stem (ES) cells were cultured to form aggregates in small volumes of basal medium in U-bottomed, non tissue-culture-treated 96-well plates and subsequently maintained in suspension culture. After growth for 48 hours, the aggregates are competent to respond to ubiquitous experimental signals which result in their symmetry-breaking and generation of defined polarised structures by 96 hours. It is envisaged that this system can be applied both to the study of early developmental events and more broadly to the processes of self-organisation and cellular decision-making. It may also provide a suitable niche for the generation of cell types present in the embryo but unobtainable from conventional adherent culture.

Developmental Biology

Brachyury cooperates with Wnt/β-Catenin signalling to elicit Primitive Streak like behaviour in differentiating mouse ES cells.

The formation of the Primitive Streak is the first visible sign of gastrulation, the process by which the three germ layers are formed from a single epithelium during early development. Embryonic Stem Cells (ESCs) provide a good system to understand the molecular and cellular events associated with these processes. Previous work, both in embryos and in culture, has shown how converging signals from both Nodal/TGF{beta}R and Wnt/{beta}-Catenin signalling pathways specify cells to adopt a Primitive Streak like fate and direct them to undertake an epithelial to mesenchymal transition (EMT). However, many of these approaches have relied on genetic analyses without taking into account the temporal progression of events within single cells. In addition, it is still unclear as to what extent events in the embryo are able to be reproduced in culture. Here, we combine flow-cytometry and a quantitative live single-cell imaging approach to demonstrate how the controlled differentiation of mouse ESCs (mESCs) towards a Primitive Streak fate in culture results in cells displaying many of the characteristics observed during early mouse development including transient Brachyury expression, EMT and increased motility. We also find that the EMT initiates the process, and this is both fuelled and terminated by the action of Bra, whose expression is dependent on the EMT and {beta}-Catenin activity. As a consequence of our analysis, we propose that a major output of Brachyury expression is in controlling the velocity of the cells that are transiting out of the Primitive Streak.

Developmental Biology

An interplay between extracellular signalling and the dynamics of the exit from pluripotency drives cell fate decisions in mouse ES cells

Embryonic Stem cells derived from the epiblast tissue of the mammalian blastocyst, retain the capability to differentiate into any adult cell type and are able to self-renew indefinitely under appropriate culture conditions. Despite the large amount of knowledge that we have accumulated to date about the regulation and control of self-renewal, efficient directed differentiation into specific tissues remains elusive. In this work, we have analyzed in a systematic manner the interaction between the dynamics of loss of pluripotency and Activin/Nodal, BMP4 and Wnt signalling in fate assignment during the early stages of differentiation of mouse ES cells in culture. During the initial period of differentiation cells exit from pluripotency and enter an Epi-like state. Following this transient stage, and under the influence of Activin/Nodal and BMP signalling, cells face a fate choice between differentiating into neuroectoderm and contributing to Primitive Streak fates. We find that Wnt signalling does not suppress neural development as previously thought and that it aids both fates in a context dependent manner. Our results suggest that as cells exit pluripotency they are endowed with a primary neuroectodermal fate and that the potency to become endomesodermal rises with time. We suggest that this situation translates into a \"race for fates\" at the level of single cells in which the neuroectodermal fate has an advantage.

Developmental Biology