Comparative Transcriptomics Analyses across Species, Organs, and Developmental Stages Reveal Functionally Constrained lncRNAs

Fabrice Daniel Darbellay, Anamaria Necsulea
OXFORD UNIV PRESS, 2020
Article

Résumé

The functionality of long noncoding RNAs (lncRNAs) is disputed. In general, lncRNAs are under weak selective pressures, suggesting that the majority of lncRNAs may be nonfunctional. However, although some surveys showed negligible phenotypic effects upon lncRNA perturbation, key biological roles were demonstrated for individual lncRNAs. Most lncRNAs with proven functions were implicated in gene expression regulation, in pathways related to cellular pluripotency, differentiation, and organ morphogenesis, suggesting that functional lncRNAs may be more abundant in embryonic development, rather than in adult organs. To test this hypothesis, we perform a multidimensional comparative transcriptomics analysis, across five developmental time points (two embryonic stages, newborn, adult, and aged individuals), four organs (brain, kidney, liver, and testes), and three species (mouse, rat, and chicken). We find that, overwhelmingly, lncRNAs are preferentially expressed in adult and aged testes, consistent with the presence of permissive transcription during spermatogenesis. LncRNAs are often differentially expressed among developmental stages and are less abundant in embryos and newborns compared with adult individuals, in agreement with a requirement for tighter expression control and less tolerance for noisy transcription early in development. For differentially expressed lncRNAs, we find that the patterns of expression variation among developmental stages are generally conserved between mouse and rat. Moreover, lncRNAs expressed above noise levels in somatic organs and during development show higher evolutionary conservation, in particular, at their promoter regions. Thus, we show that functionally constrained lncRNA loci are enriched in developing organs, and we suggest that many of these loci may function in an RNA-independent manner.

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https://infoscience.epfl.ch/record/276104?ln=fr

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Fabrice Daniel Darbellay, Anamaria Necsulea
OXFORD UNIV PRESS, 2020
Article

Résumé

Official source

https://infoscience.epfl.ch/record/276104?ln=fr

À propos de ce résultat

Concepts associés (18)

Concepts associés

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Publications associées (26)

Publications associées

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Gene expression across mammalian organ development

Maria del Carmen Sandi Perez, Ioannis Xenarios

The evolution of gene expression in mammalian organ development remains largely uncharacterized. Here we report the transcriptomes of seven organs (cerebrum, cerebellum, heart, kidney, liver, ovary and testis) across developmental time points from early organogenesis to adulthood for human, rhesus macaque, mouse, rat, rabbit, opossum and chicken. Comparisons of gene expression patterns identified correspondences of developmental stages across species, and differences in the timing of key events during the development of the gonads. We found that the breadth of gene expression and the extent of purifying selection gradually decrease during development, whereas the amount of positive selection and expression of new genes increase. We identified differences in the temporal trajectories of expression of individual genes across species, with brain tissues showing the smallest percentage of trajectory changes, and the liver and testis showing the largest. Our work provides a resource of developmental transcriptomes of seven organs across seven species, and comparative analyses that characterize the development and evolution of mammalian organs.

2019

Chargement

Previous attempts to recapitulate embryogenesis in a developmentally relevant context started with aggregates composed of a few thousand ESCs, termed embryoid bodies (EB), that upon induction of differentiation reveal a surprising level of autonomous cell fate patterning, in some cases giving rise to spatially arranged ectoderm, mesoderm and endoderm layers. However, cell fate patterning and morphogenesis in EBs is poorly reproducible and cannot be controlled. These limitations necessitate further research on the self-organization potential of ESCs, and in particular the development of more robust in vitro models mimicking post-implantation mouse embryos. This PhD thesis explored three novel approaches towards this goal. Firstly, together with collaborators, I focused on a previously established in vitro model of gastrulation termed gastruloids. Gastruloids emerge from small ESC aggregates that, upon activation of Wnt signaling, break symmetry and undergo axial morphogenesis to create structures that are similar to the developing tail of the mouse embryo. A systematic improvement of the existing culture conditions allowed gastruloids to be cultured for an extended period of time, resulting in an embryo-like patterning along the three major body axes. Furthermore, gene expression analyses, combined with immunohistochemistry and in situ hybridization assays, demonstrated that gastruloids exhibit spatio-temporal activation of Hox genes in a way that is remarkably similar to post-implantation mouse embryos. Secondly, I report a novel gastruloid culture approach, which for the first time promotes the formation of anterior neural tissues. Mouse ESCs were aggregated and treated in high-throughput in a bioengineered microwell array system to generate homogeneous âartificial epiblastsâ that transcriptionally and morphologically resemble post-implantation epiblasts. These epiblast-like (EPI) aggregates break symmetry and undergo axial elongation to establish antero-posterior patterning. I provide evidence for attenuated Wnt signaling and the presence of an epithelium in the starting EPI aggregates as the major determinants for the formation of anterior neural tissues, that are completely absent in conventional gastruloids. Lastly, I systematically explored the role of extraembryonic tissues in the patterning of artificial epiblasts. By combining EPI aggregates with trophoblast stem cell aggregates, hybrid self-organizing structures termed EpiTS embryoids were generated that mimic epiblast and extraembryonic ectoderm, respectively. EpiTS embryoids were found to execute an elaborate self-organization program that is initiated through highly localized gastrulation-like processes at the embryonic-extraembryonic interface. Upon axial elongation, EpiTS embryoids give rise to primitive brain-like and preplacodal ectoderm-like regions. These observations highlight the role of extraembryonic tissues for proper induction of gastrulation and in particular the formation of anterior neural tissues. Altogether, this thesis introduces three innovative and highly versatile stem cell culture technologies to study post-implantation mouse embryonic development. The approaches are amenable to large-scale studies aimed at identifying novel regulators of gastrulation and anterior neural development that is currently out of reach with existing experimental tools. This work should contribute to the advancement of the nascent field of synthetic embryology.

EPFL2020

Chargement

In vitro endoderm emergence and self-organisation in the absence of extraembryonic tissues and embryonic architecture

Stefano Davide Vianello

In humans, mice, and other mammals key internal organs such as the gut, the lungs, the pancreas, and the liver all derive from the same embryonic tissue: the endoderm. The development of all of these structures thus depends on a same set of early cells, and on the developmental instructions provided to them by the embryo. One approach to study what these instructions might be is to study the behaviour of endoderm cells when they can not rely on embryonic instructions. That is, when they are generated and made to develop outside of the embryo. The rationale behind this approach, widely embraced by the field of synthetic embryology, is that the unguided development of specific embryonic cell types can provide a window to the observation of intrinsic characteristics of these cells and of fundamental developmental principles which might not be executed or manifested within the regulated environment of the embryo. Accordingly, in the work here described I use gastruloids, synthetic models of development made by aggregation of mouse embryonic stem cells in vitro to characterise the developmental behaviour of endoderm cells, in the absence of embryonic cues and in the absence of the extraembryonic tissues that they would usually rely on in vivo. In the first chapter of this thesis, I provide an introduction, overview, and review of our current understanding of the developmental biology of mouse endoderm as this develops in the embryo. I also introduce gastruloids as stem-cell-based experimental systems to study developmental mechanisms in vitro, decoupled from the constraints of experimental work with animals and animal embryos. Chapter 2 is dedicated to the earliest steps of endoderm development in gastruloids. I show when and where these cells appear, and how they sort as the gastruloid starts to mature. I find that endoderm cells follow similar spatial and temporal developmental patterns as those they would follow if they were in the embryo. I notably find that they always remain in an epithelial state, even as other cell types undergo mesenchymal transition. In Chapter 3 I describe surprising dynamics of endoderm cells over time, and how they self-organise into complex architectures at the core of the gastruloids. The epithelial domain that forms is dynamic and plastic, and partitions the gastruloids into interfacing compartments. Importantly, I reveal a significant endodermal component to this in vitro system, usually considered to be unstructured and neural/mesodermal in character. In Chapter 4 I show how the core endoderm compartment that I see forming in gastruloids is patterned along the longitudinal axis of the aggregate, with different cellular identities at different anterior-posterior position of the domain. By comparison with published embryonic datasets, I identify endodermal identities corresponding to the full spectrum of those that make up the gut tube in the embryo. Specifically, I describe how most endodermal cells adopt an early intestinal fate, and how many acquire anterior foregut identities. The work described in this thesis presents gastruloids as promising in vitro systems for the study of endoderm biology, morphogenesis, and mature fate specification, filling a gap within the field of synthetic embryology. It also notably opens translational possibilities to the use of gastruloids to produce endodermal cell types which may be otherwise difficult to generate using conventional directed differentiation methods.

EPFL2022

Chargement

EPFL2020

In vitro endoderm emergence and self-organisation in the absence of extraembryonic tissues and embryonic architecture

Stefano Davide Vianello

EPFL2022