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Elevated Wnt/β‐catenin signaling has been commonly associated with tumorigenesis especially colorectal cancer (CRC). Here, an MST4‐pβ‐cateninThr40 signaling axis essential for intestinal stem cell (ISC) homeostasis and CRC development is uncovered. In response to Wnt3a stimulation, the kinase MST4 directly phosphorylates β‐catenin at Thr40 to block its Ser33 phosphorylation by GSK3β. Thus, MST4 mediates an active process that prevents β‐catenin from binding to and being degraded by β‐TrCP, leading to accumulation and full activation of β‐catenin. Depletion of MST4 causes loss of ISCs and inhibits CRC growth. Mice bearing either MST4T178E mutation with constitutive kinase activity or β‐cateninT40D mutation mimicking MST4‐mediated phosphorylation show overly increased ISCs/CSCs and exacerbates CRC. Furthermore, the MST4‐pβ‐cateninThr40 axis is upregulated and correlated with poor prognosis of human CRC. Collectively, this work establishes a previously undefined machinery for β‐catenin activation, and further reveals its function in stem cell and tumor biology, opening new opportunities for targeted therapy of CRC., A novel MST4‐pβ‐cateninThr40 signaling axis essential for maintaining intestinal stem cell (ISC) homeostasis is identified. In response to Wnt signal, MST4 directly phosphorylates β‐catenin at Thr40 to actively block the GSK3β/β‐TrCP‐mediated phosphorylation/degradation of β‐catenin. Genetic activation of the MST4‐pβ‐cateninThr40 axis exacerbates ISC‐driven colorectal cancer (CRC); and hyperactivation of this axis is clinically correlated with poor prognosis.

Stem cells are regulated both intrinsically and externally, including by signals from the local environment and distant organs. To identify genes and pathways that regulate stem-cell fates in the whole organism, we perform a genome-wide transgenic RNAi screen through ubiquitous gene knockdowns, focusing on regulators of adult Drosophila testis germline stem cells (GSCs). Here we identify 530 genes that regulate GSC maintenance and differentiation. Of these, we further knock down 113 selected genes using cell-type-specific Gal4s and find that more than half were external regulators, that is, from the local microenvironment or more distal sources. Some genes, for example, versatile (vers), encoding a heterochromatin protein, regulates GSC fates differentially in different cell types and through multiple pathways. We also find that mitosis/cytokinesis proteins are especially important for male GSC maintenance. Our findings provide valuable insights and resources for studying stem cell regulation at the organismal level., Both intrinsic and external signals regulate stem cell fate. Here, the authors perform a genome-wide RNAi screen to identify regulators of testis germline stem cells (GSC) in Drosophila, finding more than half of the genes were external regulators and heterochromatin/cytokinesis proteins mediated GSC fate.

The proportion of stem cells versus differentiated progeny is well balanced to maintain tissue homeostasis, which in turn depends on the balance of the different signaling pathways involved in stem cell self-renewal versus lineage-specific differentiation. In a screen for genes that regulate cell lineage determination in the posterior midgut, we identified that the Osa-containing SWI/SNF (Brahma) chromatin-remodeling complex regulates Drosophila midgut homeostasis. Mutations in subunits of the Osa-containing complex result in intestinal stem cell (ISC) expansion as well as enteroendocrine (EE) cell reduction. We further demonstrated that Osa regulates ISC self-renewal and differentiation into enterocytes by elaborating Notch signaling, and ISC commitment to differentiation into EE cells by regulating the expression of Asense, an EE cell fate determinant. Our data uncover a unique mechanism whereby the commitment of stem cells to discrete lineages is coordinately regulated by chromatin-remodeling factors.

Like the mammalian intestine, the Drosophila adult midgut is constantly replenished by multipotent intestinal stem cells (ISCs). Although it is well known that adult ISCs arise from adult midgut progenitors (AMPs), relatively little is known about the mechanisms that regulate AMP specification. Here, we demonstrate that Broad (Br)-mediated hormone signaling regulates AMP specification. Br is highly expressed in AMPs temporally during the larva-pupa transition stage, and br loss of function blocks AMP differentiation. Furthermore, Br is required for AMPs to develop into functional ISCs. Conversely, br overexpression drives AMPs toward premature differentiation. In addition, we found that Br and Notch (N) signaling function in parallel pathways to regulate AMP differentiation. Our results reveal a molecular mechanism whereby Br-mediated hormone signaling directly regulates stem cells to generate adult cells during metamorphosis.

The homeostasis of Drosophila midgut is maintained by multipotent intestinal stem cells (ISCs), each of which gives rise to a new ISC and an immature daughter cell, enteroblast (EB), after one asymmetric cell division. In Drosophila, the Gal4-UAS system is widely used to manipulate gene expression in a tissue- or cell-specific manner, but in Drosophila midgut, there are no ISC- or EB-specific Gal4 lines available. Here we report the generation and characterization of Dl-Gal4 and Su(H)GBE-Gal4 lines, which are expressed specifically in the ISCs and EBs separately. Additionally, we demonstrate that Dl-Gal4 and Su(H)GBE-Gal4 are expressed in adult midgut progenitors (AMPs) and niche peripheral cells (PCs) separately in larval midgut. These two Gal4 lines will serve as invaluable tools for navigating ISC behaviors.

All multicellular organisms have a specialized organ to concentrate and excrete wastes from the body. The kidneys in vertebrates and the malpighian tubules in Drosophila accomplish these functions. Mammals and Drosophila share some similar features during renal tubular development. Vertebrate kidneys are derived through the mutual induction of the ureteric bud and metanephric mesoderm, whereas the malpighian tubules of Drosophila develop from the hindgut primordium and visceral mesoderm. The vertebrate kidney also has the capacity to recover and regenerate following episodes of acute injury. Previous studies suggest that stem cells and progenitor cells may be involved in the repair and regeneration of injured renal tissue. However, studies differ as to the source of the regenerating renal cells. Recently, multipotent stem cells in Drosophila malpighian tubules were identified, and it was demonstrated that several differentiated cells in the malpighian tubules arise from these stem cells. In this article, the current understanding of kidney development and stem cell fate in mammal and Drosophila is compared. Furthermore, the potential application of the adult renal stem cells in kidney repair and the treatment of kidney cancers are discussed.

In adult Drosophila testis, asymmetric division of germline stem cells (GSCs) is specified by an oriented spindle and cortically localized adenomatous coli tumor suppressor homolog 2 (Apc2). However, the molecular mechanism underlying these events remains unclear. Here we identified Megator (Mtor), a nuclear matrix protein, which regulates GSC maintenance and asymmetric division through the spindle assembly checkpoint (SAC) complex. Loss of Mtor function results in Apc2 mis-localization, incorrect centrosome orientation, defective mitotic spindle formation, and abnormal chromosome segregation that lead to the eventual GSC loss. Expression of mitotic arrest-deficient-2 (Mad2) and monopolar spindle 1 (Mps1) of the SAC complex effectively rescued the GSC loss phenotype associated with loss of Mtor function. Collectively our results define a new role of the nuclear matrix-SAC axis in regulating stem cell maintenance and asymmetric division., Author Summary Like many stem cells, the adult Drosophila male GSC often divides asymmetrically to produce one new stem cell and one gonialblast. The asymmetric division of GSC is specified by perpendicular orientation of the mitotic spindle to the hub-GSC interface and localization of Apc2. Here we show that Tpr/Mtor regulates GSC self-renewal and asymmetric division through the SAC complex. We found that Mtor cell-autonomously required in both GSCs and CySCs to regulate their self-renewal. Loss of Mtor function affects expression and localization of Apc2 and E-cadherin. We further found that Mtor is required for the correct centrosome orientation, mitotic spindle formation, and chromosome segregation. These defects are rescued by SAC complex components, Mps1 and Mad2. These data together suggest that Mtor regulates GSC asymmetric division and maintenance through the mitotic spindle checkpoint complex. We suggest that disruption of the Tpr-SAC pathway might lead to chromosome instability, chromosome lagging, and aneuploidy, stem cell division defects, and thereby tumor development.

Stem cell competition has emerged as a mechanism for selecting fit stem cells/progenitors and controlling tumourigenesis. However, little is known about the underlying molecular mechanism. Here we identify Mlf1-adaptor molecule (Madm), a novel tumour suppressor that regulates the competition between germline stem cells (GSCs) and somatic cyst stem cells (CySCs) for niche occupancy. Madm knockdown results in overexpression of the EGF receptor ligand vein (vn), which further activates EGF receptor signalling and integrin expression non-cell autonomously in CySCs to promote their overproliferation and ability to outcompete GSCs for niche occupancy. Conversely, expressing a constitutively activated form of the Drosophila JAK kinase (hopTum−l) promotes Madm nuclear translocation, and suppresses vn and integrin expression in CySCs that allows GSCs to outcompete CySCs for niche occupancy and promotes GSC tumour formation. Tumour suppressor-mediated stem cell competition presented here could be a mechanism of tumour initiation in mammals., Stem cell competition mediates the balance between tissue homeostasis and tumour formation, but how this occurs is unclear. Here, Singh et al. show that the tumour suppressor Mlfl-adaptor molecule regulates the balance between germline stem cell and somatic cyst stem cell growth in the Drosophila testis niche.

In multicellular organisms, biological activities are regulated by cell signaling. The various signal transduction pathways regulate cell fate, proliferation, migration, and polarity. Miscoordination of the communicative signals will lead to disasters like cancer and other fatal diseases. The JAK/STAT signal transduction pathway is one of the pathways, which was first identified in vertebrates and is highly conserved throughout evolution. Studying the JAK/STAT signal transduction pathway in Drosophila provides an excellent opportunity to understand the molecular mechanism of the cell regulation during development and tumor formation. In this review, we discuss the general overview of JAK/STAT signaling in Drosophila with respect to its functions in the eye development and stem cell fate determination.

SummaryThe intestinal epithelium is the most rapidly self-renewing tissue in adult animals and maintained by intestinal stem cells (ISCs) in both Drosophila and mammals. To comprehensively identify genes and pathways that regulate ISC fates, we performed a genome-wide transgenic RNAi screen in adult Drosophila intestine and identified 405 genes that regulate ISC maintenance and lineage-specific differentiation. By integrating these genes into publicly available interaction databases, we further developed functional networks that regulate ISC self-renewal, ISC proliferation, ISC maintenance of diploid status, ISC survival, ISC-to-enterocyte (EC) lineage differentiation, and ISC-to-enteroendocrine (EE) lineage differentiation. By comparing regulators among ISCs, female germline stem cells, and neural stem cells, we found that factors related to basic stem cell cellular processes are commonly required in all stem cells, and stem-cell-specific, niche-related signals are required only in the unique stem cell type. Our findings provide valuable insights into stem cell maintenance and lineage-specific differentiation.

Adult stem cells maintain tissue homeostasis by continuously replenishing damaged, aged and dead cells in any organism. Five types of region and organ-specific multipotent adult stem cells have been identified in the Drosophila digestive system: intestinal stem cells (ISCs) in the posterior midgut; hindgut intestinal stem cells (HISCs) at the midgut/hindgut junction; renal and nephric stem cells (RNSCs) in the Malpighian Tubules; type I gastric stem cells (GaSCs) at foregut/midgut junction; and type II gastric stem cells (GSSCs) at the middle of the midgut. Despite the fact that each type of stem cell is unique to a particular organ, they share common molecular markers and some regulatory signaling pathways. Due to the simpler tissue structure, ease of performing genetic analysis, and availability of abundant mutants, Drosophila serves as an elegant and powerful model system to study complex stem cell biology. The recent discoveries, particularly in the Drosophila ISC system, have greatly advanced our understanding of stem cell self-renewal, differentiation, and the role of stem cells play in tissue homeostasis/regeneration and adaptive tissue growth.

Stem cell-mediated tissue repair is a promising approach in regenerative medicine. Intestinal epithelium is the most rapidly self-renewing tissue in adult mammals. Recently, using lineage tracing and molecular marker labeling, intestinal stem cells (ISCs) have been identified in Drosophila adult midgut. ISCs reside at the basement membrane and are multipotent as they produce both enterocytes and enteroendocrine cells. The adult Drosophila midgut provides an excellent in vivo model organ to study ISC behavior during aging, stress, regeneration, and infection. It has been demonstrated that Notch, Janus kinase/signal transducer and activator of transcription, epidermal growth factor receptor/mitogen-activated protein kinase, Hippo, and wingless signaling pathways regulate ISCs proliferation and differentiation. There are plenty of genetic tools and markers developed in recent years in Drosophila stem cell studies. These tools and markers are essential in the precise identification of stem cells as well as manipulation of genes in stem cell regulation. Here, we describe the details of genetic tools, markers, and immunolabeling techniques used in identification and characterization of adult midgut stem cells in Drosophila.

Stomach cancer is the second most frequent cause of cancer-related death worldwide. Thus, it is important to elucidate the properties of gastric stem cells, including their regulation and transformation. To date, such stem cells have not been identified in Drosophila. Here, using clonal analysis and molecular marker labeling, we identify a multipotent stem-cell pool at the foregut/midgut junction in the cardia (proventriculus). We found that daughter cells migrate upward either to anterior midgut or downward to esophagus and crop. The cardia functions as a gastric valve and the anterior midgut and crop together function as a stomach in Drosophila; therefore, we named the foregut/midgut stem cells as gastric stem cells (GaSC). We further found that JAK-STAT signaling regulates GaSCs' proliferation, Wingless signaling regulates GaSCs' self-renewal, and hedgehog signaling regulates GaSCs' differentiation. The differentiation pattern and genetic control of the Drosophila GaSCs suggest the possible similarity to mouse gastric stem cells. The identification of the multipotent stem cell pool in the gastric gland in Drosophila will facilitate studies of gastric stem cell regulation and transformation in mammal.

The spermatogonial stem cells (SSCs) are responsible for the transmission of genetic information from an individual to the next generation. SSCs play critical roles in understanding the basic reproductive biology of gametes and treatments of human infertility. SSCs not only maintain normal spermatogenesis, but also sustain fertility by critically balancing both SSC self-renewal and differentiation. This self-renewal and differentiation in turn is tightly regulated by a combination of intrinsic gene expression within the SSC as well as the extrinsic gene signals from the niche. Increased SSCs self-renewal at the expense of differentiation result in germ cell tumours, on the other hand, higher differentiation at the expense of self-renewal can result in male sterility. Testicular germ cell cancers are the most frequent cancers among young men in industrialized countries. However, understanding the pathogenesis of testis cancer has been difficult because it is formed during foetal development. Recent studies suggest that SSCs can be reprogrammed to become embryonic stem (ES)-like cells to acquire pluripotency. In the present review, we summarize the recent developments in SSCs biology and role of SSC in testicular cancer. We believe that studying the biology of SSCs will not only provide better understanding of stem cell regulation in the testis, but eventually will also be a novel target for male infertility and testicular cancers.

Over the past 2 years, our understanding of intestinal stem cells in the Drosophila posterior midgut has advanced greatly. In this review, I will focus on the establishment of these stem cells in their niche during development and the molecular mechanisms that regulate their asymmetric division in adults.

In many tissues, two or more types of stem cells share a niche, and how the stem cells coordinate their self-renewal and differentiation is poorly understood. In the Drosophila testis, germ line stem cells (GSCs) and somatic cyst progenitor cells (CPCs) contact each other and share a niche (the hub). The hub expresses a growth factor unpaired (Upd) that activates the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway in GSCs to regulate the stem cell self-renewal. Here, we demonstrate that the JAK/STAT signaling also regulates CPCs self-renewal. We also show that a negative regulator, the suppressor of cytokine signaling 36E (SOCS36E), suppresses JAK/STAT signaling in somatic cells, preventing them from out-competing the GSCs. Furthermore, through selectively manipulating the JAK/STAT signaling level in either CPCs or GSCs, we demonstrate that the somatic JAK/STAT signaling is essential for self-renewal and maintenance of both CPCs and GSCs. These data suggest that a single JAK/STAT signal from the niche orchestrate the competitive and dependent co-existence of GSCs and CPCs in the Drosophila testis niche.

The Drosophila midgut epithelium undergoes continuous regeneration that is sustained by multipotent intestinal stem cells (ISCs) underneath. Notch signaling has dual functions to control ISC behavior: it slows down the ISC proliferation and drives the activated ISCs into different differentiation pathways at a dose-dependent manner. Here we identified a molecular mechanism to unite these two contradictory functions. We found JAK-STAT signaling controls ISC proliferation and this ability is negatively regulated by Notch at least through a transcriptional control of the JAK-STAT signaling ligand, unpaired (upd). This study provides insight into how stem cells, under steady conditions, balance the processes of proliferation and differentiation to maintain the stable cellular composition of a healthy tissue.

The digestive systems in mammals and Drosophila are quite different in terms of their complexity and organization, but their biological functions are similar. The Drosophila midgut is a functional equivalent of the mouse small intestine. Adult intestinal stem cells (ISCs) have been identified in both the mouse small intestine and Drosophila midgut. The anatomy and cell renewal in the Drosophila midgut are similar to those in the mouse small intestine: the intestinal epithelium in both systems is a tube composed of epithelial cells with absorptive and secretory functions; the Notch signaling controls absorptive versus secretory fate decisions in the intestinal epithelium; cell renewal in both systems starts from stem cells in the basal cell layer, and the differentiated cells then move toward the lumen. However, it is clear that the stem cells in the two systems are regulated in different ways. In this review, we will compare cell renewal and stem cell regulation in the two systems.

Summary 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.