168 research outputs found
mRNA Translation Is Dynamically Regulated to Instruct Stem Cell Fate
Stem cells preserve tissue homeostasis by replacing the cells lost through damage or natural turnover. Thus, stem cells and their daughters can adopt two identities, characterized by different programs of gene expression and metabolic activity. The composition and regulation of these programs have been extensively studied, particularly by identifying transcription factor networks that define cellular identity and the epigenetic changes that underlie the progressive restriction in gene expression potential. However, there is increasing evidence that post-transcriptional mechanisms influence gene expression in stem cells and their progeny, in particular through the control of mRNA translation. Here, we review the described roles of translational regulation in controlling all aspects of stem cell biology, from the decision to enter or exit quiescence to maintaining self-renewal and promoting differentiation. We focus on mechanisms controlling global translation rates in cells, mTOR signaling, eIF2É‘ phosphorylation, and ribosome biogenesis and how they allow stem cells to rapidly change their gene expression in response to tissue needs or environmental changes. These studies emphasize that translation acts as an additional layer of control in regulating gene expression in stem cells and that understanding this regulation is critical to gaining a full understanding of the mechanisms that underlie fate decisions in stem cells
Rhombomere boundaries and hindbrain patterning in the zebrafish.
The vertebrate hindbrain is a segmented structure, divided into repeating segments called rhombomeres, at the interface of which boundary cells are induced as a specific cell type. Previous work has found that, in the zebrafish hindbrain, there is a distinctive organisation of glia and neurons along the anterior-posterior axis within each rhombomere, with specific neurons at the centre or boundary regions, separated by a glial curtain adjacent to the boundary. Using molecular markers, I have characterised the organisation of several cell types within each rhombomeres, and found that from 24 hours of development, neurogenesis occurs predominantly adjacent to the rhombomere boundaries, which suggests a role for boundaries in establishing this pattern. The aim of this work was to test a possible patterning role of boundaries, and to establish a genetic hierarchy between boundary genes by carrying out knockdowns of boundary-specific genes. The secreted factor wntl is expressed in dorsal boundaries and is a candidate mediator of a potential signalling role of boundaries. From many lines of evidence in previous work, wntl was known to promote proliferation of neural progenitors and inhibit neuronal differentiation. I found that in the zebrafish hindbrain, however, knockdown of wntl or of the Wnt pathway effector tcj3b causes expansion of boundary-specific markers and loss of neurogenesis in non- boundary regions. Knockdown of rfng, a component of the Notch pathway required for boundary expression of wntl also causes expansion of boundary markers. Wntl is necessary for the expression of ash and ngnl proneural genes, and of delta genes, which in turn block ectopic boundary marker expression. Thus, wntl mediates a lateral inhibition of boundary fate by regulating neurogenesis in hindbrain segments. Taken together with concurrent work in the lab, the model derived from this is remarkably similar to the regulatory interactions occurring at the dorso-ventral boundary of the Drosophila wing imaginal disc, and indicates recruitment of a conserved network of genes in non homologous tissues
Germ cells commit somatic stem cells to differentiation following priming by PI3K/Tor activity in the Drosophila testis
How and when potential becomes restricted in differentiating stem cell daughters is poorly understood. While it is thought that signals from the niche are actively required to prevent differentiation, another model proposes that stem cells can reversibly transit between multiple states, some of which are primed, but not committed, to differentiate. In the Drosophila testis, somatic cyst stem cells (CySCs) generate cyst cells, which encapsulate the germline to support its development. We find that CySCs are maintained independently of niche self-renewal signals if activity of the PI3K/Tor pathway is inhibited. Conversely, PI3K/Tor is not sufficient alone to drive differentiation, suggesting that it acts to license cells for differentiation. Indeed, we find that the germline is required for differentiation of CySCs in response to PI3K/Tor elevation, indicating that final commitment to differentiation involves several steps and intercellular communication. We propose that CySC daughter cells are plastic, that their fate depends on the availability of neighbouring germ cells, and that PI3K/Tor acts to induce a primed state for CySC daughters to enable coordinated differentiation with the germline
Socs36E Controls Niche Competition by Repressing MAPK Signaling in the Drosophila Testis
The Drosophila testis is a well-established system for studying stem cell self-renewal and competition. In this tissue, the niche supports two stem cell populations, germ line stem cells (GSCs), which give rise to sperm, and somatic stem cells called cyst stem cells (CySCs), which support GSCs and their descendants. It has been established that CySCs compete with each other and with GSCs for niche access, and mutations have been identified that confer increased competitiveness to CySCs, resulting in the mutant stem cell and its descendants outcompeting wild type resident stem cells. Socs36E, which encodes a negative feedback inhibitor of the JAK/STAT pathway, was the first identified regulator of niche competition. The competitive behavior of Socs36E mutant CySCs was attributed to increased JAK/STAT signaling. Here we show that competitive behavior of Socs36E mutant CySCs is due in large part to unbridled Mitogen-Activated Protein Kinase (MAPK) signaling. In Socs36E mutant clones, MAPK activity is elevated. Furthermore, we find that clonal upregulation of MAPK in CySCs leads to their outcompetition of wild type CySCs and of GSCs, recapitulating the Socs36E mutant phenotype. Indeed, when MAPK activity is removed from Socs36E mutant clones, they lose their competitiveness but maintain self-renewal, presumably due to increased JAK/STAT signaling in these cells. Consistently, loss of JAK/STAT activity in Socs36E mutant clones severely impairs their self-renewal. Thus, our results enable the genetic separation of two essential processes that occur in stem cells. While some niche signals specify the intrinsic property of self-renewal, which is absolutely required in all stem cells for niche residence, additional signals control the ability of stem cells to compete with their neighbors. Socs36E is node through which these processes are linked, demonstrating that negative feedback inhibition integrates multiple aspects of stem cell behavior
An improved Erk biosensor reveals oscillatory Erk dynamics driven by mitotic erasure during early development
Erk signaling dynamics elicit distinct cellular responses in a variety of contexts. The early zebrafish embryo is an ideal model to explore the role of Erk signaling dynamics in vivo, as a gradient of activated diphosphorylated Erk (P-Erk) is induced by Fgf signaling at the blastula embryonic margin. Here we describe an improved Erk-specific biosensor which we term modified Erk Kinase Translocation Reporter (modErk-KTR). We demonstrate the utility of this biosensor in vitro and in developing zebrafish and Drosophila embryos. Moreover, we show that Fgf/Erk signaling is dynamic and coupled to tissue growth during both early zebrafish and Drosophila development. Signaling is rapidly extinguished just prior to mitosis, which we refer to as mitotic erasure, inducing periods of inactivity, thus providing a source of heterogeneity in an asynchronously dividing tissue. Our modified reporter and transgenic lines represent an important resource for interrogating the role of Erk signaling dynamics in vivo
A kinase translocation reporter reveals real-time dynamics of ERK activity in Drosophila
Extracellular signal-regulated kinase (ERK) lies downstream of a core signalling cascade that controls all aspects of development and adult homeostasis. Recent developments have led to new tools to image and manipulate the pathway. However, visualising ERK activity in vivo with high temporal resolution remains a challenge in Drosophila. We adapted a kinase translocation reporter (KTR) for use in Drosophila, which shuttles out of the nucleus when phosphorylated by ERK. We show that ERK-KTR faithfully reports endogenous ERK signalling activity in developing and adult tissues, and that it responds to genetic perturbations upstream of ERK. Using ERK-KTR in time-lapse imaging, we made two novel observations: firstly, sustained hyperactivation of ERK by expression of dominant-active epidermal growth factor receptor raised the overall level but did not alter the kinetics of ERK activity; secondly, the direction of migration of retinal basal glia correlated with their ERK activity levels, suggesting an explanation for the heterogeneity in ERK activity observed in fixed tissue. Our results show that KTR technology can be applied in Drosophila to monitor ERK activity in real-time and suggest that this modular tool can be further adapted to study other kinases. This article has an associated First Person interview with the first author of the paper
Neutral competition of stem cells is skewed by proliferative changes downstream of Hh and Hpo.
Neutral competition, an emerging feature of stem cell homeostasis, posits that individual stem cells can be lost and replaced by their neighbors stochastically, resulting in chance dominance of a clone at the niche. A single stem cell with an oncogenic mutation could bias this process and clonally spread the mutation throughout the stem cell pool. The Drosophila testis provides an ideal system for testing this model. The niche supports two stem cell populations that compete for niche occupancy. Here, we show that cyst stem cells (CySCs) conform to the paradigm of neutral competition and that clonal deregulation of either the Hedgehog (Hh) or Hippo (Hpo) pathway allows a single CySC to colonize the niche. We find that the driving force behind such behavior is accelerated proliferation. Our results demonstrate that a single stem cell colonizes its niche through oncogenic mutation by co-opting an underlying homeostatic process.This is the final version. It was first published by Wiley at http://onlinelibrary.wiley.com/doi/10.15252/embj.201387500/abstract
An improved Erk biosensor detects oscillatory Erk dynamics driven by mitotic erasure during early development
Extracellular signal-regulated kinase (Erk) signaling dynamics elicit distinct cellular responses in a variety of contexts. The early zebrafish embryo is an ideal model to explore the role of Erk signaling dynamics in vivo, as a gradient of activated diphosphorylated Erk (P-Erk) is induced by fibroblast growth factor (Fgf) signaling at the blastula margin. Here, we describe an improved Erk-specific biosensor, which we term modified Erk kinase translocation reporter (modErk-KTR). We demonstrate the utility of this biosensor in vitro and in developing zebrafish and Drosophila embryos. Moreover, we show that Fgf/Erk signaling is dynamic and coupled to tissue growth during both early zebrafish and Drosophila development. Erk activity is rapidly extinguished just prior to mitosis, which we refer to as mitotic erasure, inducing periods of inactivity, thus providing a source of heterogeneity in an asynchronously dividing tissue. Our modified reporter and transgenic lines represent an important resource for interrogating the role of Erk signaling dynamics in vivo
Cell-cycle exit and stem cell differentiation are coupled through regulation of mitochondrial activity in the Drosophila testis
Summary
Whereas stem and progenitor cells proliferate to maintain tissue homeostasis, fully differentiated cells exit the cell cycle. How cell identity and cell-cycle state are coordinated during differentiation is still poorly understood. The Drosophila testis niche supports germline stem cells and somatic cyst stem cells (CySCs). CySCs give rise to post-mitotic cyst cells, providing a tractable model to study the links between stem cell identity and proliferation. We show that, while cell-cycle progression is required for CySC self-renewal, the E2f1/Dp transcription factor is dispensable for self-renewal but instead must be silenced by the Drosophila retinoblastoma homolog, Rbf, to permit differentiation. Continued E2f1/Dp activity inhibits the expression of genes important for mitochondrial activity. Furthermore, promoting mitochondrial biogenesis rescues the differentiation of CySCs with ectopic E2f1/Dp activity but not their cell-cycle exit. In sum, E2f1/Dp coordinates cell-cycle progression with stem cell identity by regulating the metabolic state of CySCs
Somatic stem cell differentiation is regulated by PI3K/Tor signaling in response to local cues
Stem cells reside in niches that provide signals to maintain self-renewal, and differentiation is viewed as a passive process that depends on loss of access to these signals. Here, we demonstrate that the differentiation of somatic cyst stem cells (CySCs) in the Drosophila testis is actively promoted by PI3K/Tor signaling, as CySCs lacking PI3K/Tor activity cannot differentiate properly. We find that an insulin peptide produced by somatic cells immediately outside of the stem cell niche acts locally to promote somatic differentiation through Insulin-like receptor (InR) activation. These results indicate that there is a local ‘differentiation' niche that upregulates PI3K/Tor signaling in the early daughters of CySCs. Finally, we demonstrate that CySCs secrete the Dilp-binding protein ImpL2, the Drosophila homolog of IGFBP7, into the stem cell niche, which blocks InR activation in CySCs. Thus, we show that somatic cell differentiation is controlled by PI3K/Tor signaling downstream of InR and that the local production of positive and negative InR signals regulates the differentiation niche. These results support a model in which leaving the stem cell niche and initiating differentiation are actively induced by signaling
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