Drosophila Boi limits Hedgehog levels to suppress follicle stem cell proliferation

Stem cells depend on signals from cells within their microenvironment, or niche, as well as factors secreted by distant cells to regulate their maintenance and function. Here we show that Boi, a Hedgehog (Hh)-binding protein, is a novel suppressor of proliferation of follicle stem cells (FSCs) in the Drosophila ovary. Hh is expressed in apical cells, distant from the FSC niche, and diffuses to reach FSCs, where it promotes FSC proliferation. We show that Boi is expressed in apical cells and exerts its suppressive effect on FSC proliferation by binding to and sequestering Hh on the apical cell surface, thereby inhibiting Hh diffusion. Our studies demonstrate that cells distant from the local niche can regulate stem cell function through ligand sequestration, a mechanism that likely is conserved in other epithelial tissues.     

An epithelial niche in the Drosophila ovary undergoes long-range stem cell replacement

Adult epithelial stem cells are thought to reside in specific niches, where they are maintained by adhesion to stromal cells and by intercellular signals. In niches that harbor multiple adjacent stem cells, such as those maintaining Drosophila germ cells, lost stem cells are replaced by division of neighboring stem cells or reversion of transit cells. We have characterized the Drosophila follicle stem cell (FSC) niche as a model of the epithelial niche to learn whether nonneighboring cells can also generate stem cell replacements. Exactly two stroma-free FSC niches holding single FSCs are located in fixed locations on opposite edges of the Drosophila ovariole. FSC daughters regularly migrate across the width of the ovariole to the other niche before proliferating and contributing to the follicle cell monolayer. Crossmigrating FSC daughters compete with the resident FSC for niche occupancy and are the source of replacement FSCs. The ability of stem cell daughters to target a distant niche and displace its resident stem cell suggests that precancerous mutations might spread from niche to niche within stem cell-based tissues.     

Novel Tools for Genetic Manipulation of Follicle Stem Cells in the Drosophila Ovary Reveal an Integrin-Dependent Transition from Quiescence to Proliferation

In many tissues, the presence of stem cells is inferred by the capacity of the tissue to maintain homeostasis and undergo repair after injury. Isolation of self-renewing cells with the ability to generate the full array of cells within a given tissue strongly supports this idea, but the identification and genetic manipulation of individual stem cells within their niche remain a challenge. Here we present novel methods for marking and genetically altering epithelial follicle stem cells (FSCs) within the Drosophila ovary. Using these new tools, we define a sequential multistep process that comprises transitioning of FSCs from quiescence to proliferation. We further demonstrate that integrins are cell-autonomously required within FSCs to provide directional signals that are necessary at each step of this process. These methods may be used to define precise roles for specific genes in the sequential events that occur during FSC division after a period of quiescence.     

Basolateral Junction Proteins Regulate Competition for the Follicle Stem Cell Niche in the Drosophila Ovary

Epithelial stem cells are routinely lost or damaged during adult life and must therefore be replaced to maintain homeostasis. Recent studies indicate that stem cell replacement occurs through neutral competition in many types of epithelial tissues, but little is known about the factors that determine competitive outcome. The epithelial follicle stem cells (FSCs) in the Drosophila ovary are regularly lost and replaced during normal homeostasis, and we show that FSC replacement conforms to a model of neutral competition. In addition, we found that FSCs mutant for the basolateral junction genes, lethal giant larvae (lgl) or discs large (dlg), undergo a biased competition for niche occupancy characterized by increased invasion of neighboring FSCs and reduced loss. Interestingly, FSCs mutant for a third basolateral junction gene, scribble (scrib), do not exhibit biased competition, suggesting that Lgl and Dlg regulate niche competition through a Scrib-independent process. Lastly, we found that FSCs have a unique cell polarity characterized by broadly distributed adherens junctions and the lack of a mature apical domain. Collectively, these observations indicate that Lgl and Dlg promote the differentiation of FSC progeny to a state in which they are less prone to invade the neighboring niche. In addition, we demonstrate that the neutral drift model can be adapted to quantify non-neutral behavior of mutant clones.     

Rap-GEF signaling controls stem cell anchoring to their niche through regulating DE-cadherin-mediated cell adhesion in the Drosophila testis

Stem cells will undergo self-renewal to produce new stem cells if they are maintained in their niches. The regulatory mechanisms that recruit and maintain stem cells in their niches are not well understood. In Drosophila testes, a group of 12 nondividing somatic cells, called the hub, identifies the stem cell niche by producing the growth factor Unpaired (Upd). Here, we show that Rap-GEF/Rap signaling controls stem cell anchoring to the niche through regulating DE-cadherin-mediated cell adhesion. Loss of function of a Drosophila Rap-GEF (Gef26) results in loss of both germline and somatic stem cells. The Gef26 mutation specifically impairs adherens junctions at the hub-stem cell interface, which results in the stem cells "drifting away" from the niche and losing stem cell identity. Thus, the Rap signaling/E-cadherin pathway may represent one mechanism that regulates polarized niche formation and stem cell anchoring.     

Rab11 is required for epithelial cell viability, terminal differentiation, and suppression of tumor-like growth in the Drosophila egg chamber

Background

The Drosophila egg chamber provides an excellent system in which to study the specification and differentiation of epithelial cell fates because all of the steps, starting with the division of the corresponding stem cells, called follicle stem cells, have been well described and occur many times over in a single ovary.

Methodology/Principal Findings

Here we investigate the role of the small Rab11 GTPase in follicle stem cells (FSCs) and in their differentiating daughters, which include main body epithelial cells, stalk cells and polar cells. We show that rab11-null FSCs maintain their ability to self renew, even though previous studies have shown that FSC self renewal is dependent on maintenance of E-cadherin-based intercellular junctions, which in many cell types, including Drosophila germline stem cells, requires Rab11. We also show that rab11-null FSCs give rise to normal numbers of cells that enter polar, stalk, and epithelial cell differentiation pathways, but that none of the cells complete their differentiation programs and that the epithelial cells undergo premature programmed cell death. Finally we show, through the induction of rab11-null clones at later points in the differentiation program, that Rab11 suppresses tumor-like growth of epithelial cells. Thus, rab11-null epithelial cells arrest differentiation early, assume an aberrant cell morphology, delaminate from the epithelium, and invade the neighboring germline cyst. These phenotypes are associated with defects in E-cadherin localization and a general loss of cell polarity.

Conclusions/Significance

While previous studies have revealed tumor suppressor or tumor suppressor-like activity for regulators of endocytosis, our study is the first to identify such activity for regulators of endocytic recycling. Our studies also support the recently emerging view that distinct mechanisms regulate junction stability and plasticity in different tissues.     

The Drosophila ovarian and testis stem cell niches: similar somatic stem cells and signals

The stem cell niches at the apex of Drosophila ovaries and testes have been viewed as distinct in two major respects. While both contain germline stem cells, the testis niche also contains "cyst progenitor" stem cells, which divide to produce somatic cells that encase developing germ cells. Moreover, while both niches utilize BMP signaling, the testis niche requires a key JAK/STAT signal. We now show, by lineage marking, that the ovarian niche also contains a second type of stem cell. These "escort stem cells" morphologically resemble testis cyst progenitor cells and their daughters encase developing cysts before undergoing apoptosis at the time of follicle formation. In addition, we show that JAK/STAT signaling also plays a critical role in ovarian niche function, and acts within escort cells. These observations reveal striking similarities in the stem cell niches of male and female gonads, and suggest that they are largely governed by common mechanisms.     

Integrin-mediated adhesion and stem-cell-niche interactions

The regulation of stem cell behavior and maintenance typically involves the integration of both intrinsic and extrinsic cues. One such external cue, integrin-mediated cell adhesion to the extracellular matrix, plays an important part in regulating stem cell function and maintenance. In particular, integrins help define and shape the microenvironment in which stem cells are found: the stem cell niche. Integrins have a diverse array of roles in this context including homing of stem cells to their niche, maintaining stem cells in the niche, developing stem-cell-niche architecture, regulating stem cell proliferation and self renewal, and finally, controlling the orientation of dividing stem cells. Because of their various roles in directing stem cell behavior, integrin-mediated adhesion and signaling in the niche have been implicated in processes that underlie cancer progression and metastasis.     

The Drosophila ovary: an active stem cell community

Only a small number of cells in adult tissues (the stem cells) possess the ability to self-renew at every cell division,while producing differentiating daughter cells to maintain tissue homeostasis for an organism's lifetime.The Drosophilaovary harbors three different types of stem cell populations (germline stem cell (GSC),somatic stem cell (SSC) andescort stem cell (ESC)) located in a simple anatomical structure known as germarium,rendering it one of the best modelsystems for studying stem cell biology due to reliable stem cell identification and available sophisticated genetic toolsfor manipulating gene functions.Particularly,the niche for the GSC is among the first and best studied ones,and studieson the GSC and its niche have made many unique contributions to a better understanding of relationships between stemcells and their niche.So far,both the GSC and the SSC have been shown to be regulated by extrinsic factors originatingfrom their niche and intrinsic factors functioning within.Multiple signaling pathways are required for controlling GSCand SSC self-renewal and differentiation,which provide unique opportunities to investigate how multiple signals fromthe niche are interpreted in the stem cell.Since the Drosophila ovary contains three types of stem cells,it also providesoutstanding opportunities to study how multiple stem cells in a given tissue work collaboratively to contribute to tissuefunction and maintenance.This review highlights recent major advances in studying Drosophila ovarian stem cells andalso discusses future directions and challenges.     

Proteomic profiling of secretome and adherent plasma membranes from distinct mammary epithelial cell subpopulations

The stem cell niche comprises stem cells (SCs), stromal cells, soluble factors, extracellular matrix constituents and vascular networks. The ability to identify signals that regulate SC self-renewal and differentiation is confounded by the difficulty in isolating pure SC niche components in sufficient quantities to enable their biochemical characterisation. Here, we report the extracellular (secretome) and adherent plasma membrane proteomes of three distinct epithelial cell subpopulations isolated and immortalized from the mouse mammary gland--basal and mammary stem cell (basal/MaSC), luminal progenitor (LP) and mature luminal (ML) cell lines. GeLC-MS/MS-based proteomic profiling revealed a distinct switch in components modulating Wnt and ephrin signalling, and integrin-mediated interactions amongst the three cell subpopulations. For example, expression of ephrin B2, ephrin receptors A1, and A2, as well as integrins α2β1 and α6β4 were shown to be enriched in basal/MaSCs, relative to LP and ML cells. Conspicuously, Wnt10a was uniquely detected in basal/MaSCs, and may modulate the canonical Wnt signalling pathway to maintain basal/MaSC activity. By contrast, non-canonical Wnt signalling might be elevated in ML cells, as evidenced by the high expression levels of Wnt5a, Wnt5b, and the transmembrane tyrosine kinase Ror2.     

精原干细胞niche的研究进展

精原干细胞(spermatogonial stem cells,SSCs)是指睾丸内位于曲精细管基膜上既能自我更新维持自身适量恒定,又能定向分化产生精母细胞的一类原始精原细胞。随着干细胞深入的研究,人们发现了一种控制着干细胞可塑性与命运的微环境,此微环境被称为干细胞niche,干细胞niche由niche细胞、细胞外基质、细胞因子等构成。精原干细胞niche是由黏附因子、生长因子、支持细胞、间质细胞以及小管周肌肉细胞组成。大量的研究表明支持细胞在睾丸中是主要的成体细胞,通过分泌可溶性的因子来影响精原干细胞niche的结构与功能,同时支持细胞还能够间接的影响其他的成体细胞。随着年龄的增长使得精原干细胞niche的功能下降。精原干细胞数量以及精原干细胞niche为我们研究组织特异性干细胞生物学以及保持再生组织平衡提供了很宝贵的线索,精原干细胞对于保持组织的自我更新具有很重要的作用,并且受到人们大量的关注,然而精原干细胞niche也起到很重要的作用,它为治疗一些疾病提供新途径.本文将综述精原干细胞niche及其变化对精原干细胞功能调节的相关研究进展。     

Integrin-dependent anchoring of a stem-cell niche

Interactions between stem cells and their surrounding microenvironment, or niche, are critical for the establishment and maintenance of stem-cell properties. The adult Drosophila testis contains a morphologically discrete stem-cell niche, the 'hub'. The small cluster of non-dividing, somatic hub cells at the anterior tip of the fly testis is contacted by the germline stem cells (GSCs), which retain their stem-cell character through the direct association with the hub. Here we show that integrin-mediated adhesion is important for maintaining the correct position of embryonic hub cells during gonad morphogenesis. The misplaced hub in integrin-deficient embryos directs the orientation of cell divisions in the presumptive GSCs, a hallmark of the active germline stem-cell niche. A decrease in integrin-mediated adhesion in adult testes, which resulted in a loss of the hub and the stem-cell population, revealed the importance of hub-cell anchoring. Finally, we show that an extracellular matrix (ECM) is present around the gonad during late embryogenesis and that this ECM is defective in integrin-deficient gonads. On the basis of our data, we propose that integrins are required for the attachment of the hub cells to the ECM, which is essential for maintaining the stem-cell niche.     

Anchoring stem cells in the niche by cell adhesion molecules

Adult stem cells generally reside in supporting local micro environments or niches, and intimate stem cell and niche association is critical for their long-term maintenance and function. Recent studies in model organisms especially Drosophila have started to unveil the underlying mechanisms of stem anchorage in the niche at the molecular and cellular level. Two types of cell adhesion molecules are emerging as essential players: cadherin-mediated cell adhesion for keeping stem cells within stromal niches, whereas integrin-mediated cell adhesion for keeping stem cells within epidermal niches. Further understanding stem cell anchorage and release in coupling with environmental changes should provide further insights into homeostasis control in tissues that harbor stem cells.Key words: stem cell, niche, anchorage, cell adhesion, extracellular matrix, cadherin, integrinTissue-specific adult stem cells are characterized by their prolonged self-renewal ability and potentiality to differentiate into one or more types of mature cells. These unique properties make stem cells essential for maintaining tissue homeostasis throughout life. It is generally believed that all adult stem cells reside in specific microenvironments named niches, which provide physical support and produce critical signals to maintain stem cell identity and govern their behavior.^1–4 Consequently, intimate stem cell and niche association is a pre-requisite for stem cell''s long-term maintenance and function. How stem cells are kept within the niche is thus an important issue in stem cell biology. Characterization of a number of stem cell niches in model organisms has led to the classification of niches into two general types: stromal niches where stem cells have direct membrane contact with the niche cells and epidermal niches where stem cells are usually associated with the extracellular matrix (ECM), and do not directly contact any fixed stromal cells.¹ Studies in Drosophila have led to the cellular and functional verification of the stem cell niche theory^5,6 and not surprisingly, have also led to the discovery of the molecular mechanisms anchoring stem cells to the niche. Here I consider recent studies in Drosophila on types of cell adhesions used to anchor stem cells in the niches, and summarize cell adhesion molecules utilized in the most characterized niches in the mammalian tissues, and suggest that cadherin-mediated cell-to-cell adhesion and integrin-mediated cell-to-ECM adhesion are possibly two general mechanisms that function in respective stromal or epidermal niches for stem cell anchorage in diverse organisms.     

Scanning electron microscopy and the study of Fc and C3b receptors of cultured fat-storing cells from rat liver

Non-parenchymal fat-storing cells (FSCs) were isolated from rat liver by Metrizamide density centrifugation after liver perfusion with media containing pronase and/or collagenase. These cells were cultured in Williams' E medium with 20% fetal calf serum under the usual conditions. FSCs, identified by vitamin A-specific fluorescence (VAF), were viewed under conventional light microscopy 24, 48 and 72 h after seeding. At 24 h, some FSCs remained hemispheric; at 48 h all FSCs were well spread out, and at 72 h the number of FSCs had increased and the VAF intensity had become weaker. Scanning electron microscopy revealed that the features of the FSCs clearly differed from those of the other non-parenchymal cells. The in vitro FSC features closely resembled the in vivo features. FSCs cultured for 48 h had flattened, triangular or oval shapes with fairly smooth surfaces that showed scattered microvilli 0.1 micron long. Two types of processes, long slender (130 microns maximum length) and short broad ones, protruded from the cell body. These processes had smaller projections 0.2 micron wide that looked like fern leaves. Many lipid droplets could be seen under the cell surface. IgG-coated sheep erythrocytes (EAs) and IgM-C3-coated ox erythrocytes (EACs) were used to investigate whether there are Fc and C3b receptors on the FSCs. No rosette-formation characteristic of EAs or EACs was present on the FSCs; proof that FSCs have neither receptor.     

Insulin signals control the competence of the Drosophila female germline stem cell niche to respond to Notch ligands

Adult stem cells reside in specialized microenvironments, or niches, that are essential for their function in vivo. Stem cells are physically attached to the niche, which provides secreted factors that promote their self-renewal and proliferation. Despite intense research on the role of the niche in regulating stem cell function, much less is known about how the niche itself is controlled. We previously showed that insulin signals directly stimulate germline stem cell (GSC) division and indirectly promote GSC maintenance via the niche in Drosophila. Insulin-like peptides are required for maintenance of cap cells (a major component of the niche) via modulation of Notch signaling, and they also control attachment of GSCs to cap cells and E-cadherin levels at the cap cell–GSC junction. Here, we further dissect the molecular and cellular mechanisms underlying these processes. We show that insulin and Notch ligands directly stimulate cap cells to maintain their numbers and indirectly promote GSC maintenance. We also report that insulin signaling, via phosphoinositide 3-kinase and FOXO, intrinsically controls the competence of cap cells to respond to Notch ligands and thereby be maintained. Contrary to a previous report, we also find that Notch ligands originated in GSCs are not required either for Notch activation in the GSC niche, or for cap cell or GSC maintenance. Instead, the niche itself produces ligands that activate Notch signaling within cap cells, promoting stability of the GSC niche. Finally, insulin signals control cap cell–GSC attachment independently of their role in Notch signaling. These results are potentially relevant to many systems in which Notch signaling modulates stem cells and demonstrate that complex interactions between local and systemic signals are required for proper stem cell niche function.     

Nerve-derived sonic hedgehog defines a niche for hair follicle stem cells capable of becoming epidermal stem cells

In adult skin, stem cells in the hair follicle bulge cyclically regenerate the follicle, whereas a distinct stem cell population maintains the epidermis. The degree to which all bulge cells have equal regenerative potential is not known. We found that Sonic hedgehog (Shh) from neurons signals to a population of cells in the telogen bulge marked by the Hedgehog response gene Gli1. Gli1-expressing bulge cells function as multipotent stem cells in their native environment and repeatedly regenerate the anagen follicle. Shh-responding perineural bulge cells incorporate into healing skin wounds where, notably, they can change their lineage into epidermal stem cells. The perineural niche (including Shh) is dispensable for follicle contributions to acute wound healing and skin homeostasis, but is necessary to maintain bulge cells capable of becoming epidermal stem cells. Thus, nerves cultivate a microenvironment where Shh creates a molecularly and phenotypically distinct population of hair follicle stem cells.     

The receptor tyrosine phosphatase Lar regulates adhesion between Drosophila male germline stem cells and the niche

The stem cell niche provides a supportive microenvironment to maintain adult stem cells in their undifferentiated state. Adhesion between adult stem cells and niche cells or the local basement membrane ensures retention of stem cells in the niche environment. Drosophila male germline stem cells (GSCs) attach to somatic hub cells, a component of their niche, through E-cadherin-mediated adherens junctions, and orient their centrosomes toward these localized junctional complexes to carry out asymmetric divisions. Here we show that the transmembrane receptor tyrosine phosphatase Leukocyte-antigen-related-like (Lar), which is best known for its function in axonal migration and synapse morphogenesis in the nervous system, helps maintain GSCs at the hub by promoting E-cadherin-based adhesion between hub cells and GSCs. Lar is expressed in GSCs and early spermatogonial cells and localizes to the hub-GSC interface. Loss of Lar function resulted in a reduced number of GSCs at the hub. Lar function was required cell-autonomously in germ cells for proper localization of Adenomatous polyposis coli 2 and E-cadherin at the hub-GSC interface and for the proper orientation of centrosomes in GSCs. Ultrastructural analysis revealed that in Lar mutants the adherens junctions between hub cells and GSCs lack the characteristic dense staining seen in wild-type controls. Thus, the Lar receptor tyrosine phosphatase appears to polarize and retain GSCs through maintenance of localized E-cadherin-based adherens junctions.     

Artificial niche substrates for embryonic and induced pluripotent stem cell cultures

Stem cells possess the ability to self-renew and differentiate into other cell types. In vivo, stem cells reside in their own anatomic niches in a defined physiological environment, from which they are released to differentiate into a required cell type when deemed appropriate. While a resident within the niche, the stem cell receives signals that in turn maintain the cell in a pluripotent state. In addition, the niche also provides nourishment to the cell. Physically, the niche also serves to anchor the cell via various ECM components and cell-adhesion molecules. Therefore, in vitro models that replicate the in vivo niche will lead to a better understanding of stem cell fate and turnover. In turn, this will help inform attempts to culture stem cells in vitro on artificial niche-like substrates. In this review, we have highlighted recent studies describing artificial niche-like substrates used to culture embryonic and induced pluripotent stem cells in vitro.