Mutations in the Polycomb Group Gene polyhomeotic Lead to Epithelial Instability in both the Ovary and Wing Imaginal Disc in Drosophila

Most human cancers originate from epithelial tissues and cell polarity and adhesion defects can lead to metastasis. The Polycomb-Group of chromatin factors were first characterized in Drosophila as repressors of homeotic genes during development, while studies in mammals indicate a conserved role in body plan organization, as well as an implication in other processes such as stem cell maintenance, cell proliferation, and tumorigenesis. We have analyzed the function of the Drosophila Polycomb-Group gene polyhomeotic in epithelial cells of two different organs, the ovary and the wing imaginal disc.Clonal analysis of loss and gain of function of polyhomeotic resulted in segregation between mutant and wild-type cells in both the follicular and wing imaginal disc epithelia, without excessive cell proliferation. Both basal and apical expulsion of mutant cells was observed, the former characterized by specific reorganization of cell adhesion and polarity proteins, the latter by complete cytoplasmic diffusion of these proteins. Among several candidate target genes tested, only the homeotic gene Abdominal-B was a target of PH in both ovarian and wing disc cells. Although overexpression of Abdominal-B was sufficient to cause cell segregation in the wing disc, epistatic analysis indicated that the presence of Abdominal-B is not necessary for expulsion of polyhomeotic mutant epithelial cells suggesting that additional polyhomeotic targets are implicated in this phenomenon.Our results indicate that polyhomeotic mutations have a direct effect on epithelial integrity that can be uncoupled from overproliferation. We show that cells in an epithelium expressing different levels of polyhomeotic sort out indicating differential adhesive properties between the cell populations. Interestingly, we found distinct modalities between apical and basal expulsion of ph mutant cells and further studies of this phenomenon should allow parallels to be made with the modified adhesive and polarity properties of different types of epithelial tumors

Lack of an Antibacterial Response Defect in Drosophila Toll-9 Mutant

Toll and Toll-like receptors represent families of receptors involved in mediating innate immunity response in insects and mammals. Although Drosophila proteome contains multiple Toll paralogs, Toll-1 is, so far, the only receptor to which an immune role has been attributed. In contrast, every single mammalian TLR is a key membrane receptor upstream of the vertebrate immune signaling cascades. The prevailing view is that TLR-mediated immunity is ancient. Structural analysis reveals that Drosophila Toll-9 is the most closely related to vertebrate TLRs and utilizes similar signaling components as Toll-1. This suggests that Toll-9 could be an ancestor of TLR-like receptors and could have immune function. Consistently, it has been reported that over-expression of Toll-9 in immune tissues is sufficient to induce the expression of some antimicrobial peptides in flies. These results have led to the idea that Toll-9 could be a constitutively active receptor that maintain significant levels of antimicrobial molecules and therefore provide constant basal protection against micro-organisms. To test theses hypotheses, we generated and analyzed phenotypes associated with a complete loss-of-function allele of Toll-9. Our results suggest that Toll-9 is neither required to maintain a basal anti-microbial response nor to mount an efficient immune response to bacterial infection

Developmental regulation of regenerative potential in Drosophila by ecdysone through a bistable loop of ZBTB transcription factors

International audienceIn many organisms, the regenerative capacity of tissues progressively decreases as development progresses. However, the developmental mechanisms that restrict regenerative potential remain unclear. In Drosophila, wing imaginal discs become unable to regenerate upon damage during the third larval stage (L3). Here, we show that production of ecdysone after larvae reach their critical weight (CW) terminates the window of regenerative potential by acting on a bistable loop composed of two antagonistic Broad-complex/Tramtrack/Bric-à-brac Zinc-finger (ZBTB) genes: chinmo and broad (br). Around mid L3, ecdysone signal-ing silences chinmo and activates br to switch wing epithelial progenitors from a default self-renewing to a differentiation-prone state. Before mid L3, Chinmo promotes a strong regener-ative response upon tissue damage. After mid L3, Br installs a nonpermissive state that represses regeneration. Transient down-regulation of ecdysone signaling or Br in late L3 lar-vae enhances chinmo expression in damaged cells that regain the capacity to regenerate. This work unveils a mechanism that ties the self-renewing and regenerative potential of epi-thelial progenitors to developmental progression

Two distinct mechanisms silence chinmo in Drosophila neuroblasts and neuroepithelial cells to limit their self-renewal

International audienceWhether common principles regulate the self-renewing potential of neural stem cells (NSCs) throughout the developing central nervous system is still unclear. In the Drosophila ventral nerve cord and central brain, asymmetrically dividing NSCs, called neuroblasts (NBs), progress through a series of sequentially expressed transcription factors that limits self-renewal by silencing a genetic module involving the transcription factor Chinmo. Here, we find that Chinmo also promotes neuroepithelium growth in the optic lobe during early larval stages by boosting symmetric self-renewing divisions while preventing differentiation. Neuroepithelium differentiation in late larvae requires the transcriptional silencing of chinmo by ecdysone, the main steroid hormone, therefore allowing coordination of neural stem cell self-renewal with organismal growth. In contrast, chinmo silencing in NBs is post-transcriptional and does not require ecdysone. Thus, during Drosophila development, humoral cues or tissue-intrinsic temporal specification programs respectively limit self-renewal in different types of neural progenitors through the transcriptional and post-transcriptional regulation of the same transcription factor

fused regulates germline cyst mitosis and differentiation during Drosophila oogenesis.

The fused gene encodes a serine–threonine kinase that functions as a positive regulator of Hedgehog signal transduction in Drosophila embryogenesis, wing morphogenesis, and somatic cell development during oogenesis. Here, we have characterized the germline ovarian tumors present in adult ovaries of fused mutant females, a phenotype not observed upon deregulation of any other component of Hedgehog signaling. In the strongest fused mutant contexts, we found that tumorous ovarian follicles accumulate early spectrosome-containing germ cells corresponding to germline stem cells and/or early cystoblasts as evidenced by activated Dpp signal transduction and transcriptional repression of bag-of-marbles, encoding the cystoblast determination factor. These early germ cells are maintained far from their usual position in a specialized niche of somatic cells in the apical part of the germarium, which appears normal in size in fused mutant ovarioles. Therefore, these results indicate a novel function for fused in downregulation of Dpp signaling which is necessary for de-repression of bag-of-marbles and consequent cystoblast determination. The abnormal accumulation of these early germ cells seems to be due primarily to defects in differentiation since we show that germline stem cell proliferation in the germarium is not affected. A later block in germline development, at the 16-cell cyst stage before significant nurse cell and oocyte differentiation, was also observed in tumorous follicles when fused function was only partially lowered. Finally, fused mutant ovaries exhibit some germline cysts having undergone a supernumerary fifth mitotic division. Through clonal analysis, we provide evidence that fused regulates these cystocyte divisions cell autonomously, while the tumorous phenotype probably reflects both a somatic and germline requirement for fused for cyst and follicle development

Peptidoglycan sensing by the receptor PGRP-LE in the Drosophila gut induces immune responses to infectious bacteria and tolerance to microbiota.

International audienceGut epithelial cells contact both commensal and pathogenic bacteria, and proper responses to these bacteria require a balance of positive and negative regulatory signals. In the Drosophila intestine, peptidoglycan-recognition proteins (PGRPs), including PGRP-LE, play central roles in bacterial recognition and activation of immune responses, including induction of the IMD-NF-κB pathway. We show that bacteria recognition is regionalized in the Drosophila gut with various functional regions requiring different PGRPs. Specifically, peptidoglycan recognition by PGRP-LE in the gut induces NF-κB-dependent responses to infectious bacteria but also immune tolerance to microbiota through upregulation of pirk and PGRP-LB, which negatively regulate IMD pathway activation. Loss of PGRP-LE-mediated detection of bacteria in the gut results in systemic immune activation, which can be rescued by overexpressing PGRP-LB in the gut. Together these data indicate that PGRP-LE functions as a master gut bacterial sensor that induces balanced responses to infectious bacteria and tolerance to microbiota

In vivo AGO-APP identifies a module of microRNAs cooperatively controlling exit from neural stem cell state

Abstract MicroRNAs (miRNAs) are essential regulators of all developmental processes. Their function is particularly important during neurogenesis, when the production of large numbers of neurons from a limited number of neural stem cells depends on the precise control of determination, proliferation and differentiation. However, miRNA regulation of target mRNAs is highly promiscuous, one miRNA can target many mRNAs and vice versa, raising the question of how specificity is achieved to elicit a precise regulatory response. Here we introduce AGO-APP, a novel approach to purify Argonaute-bound miRNAs directly from cells and tissues in vivo, to isolate actively inhibiting miRNAs from different neural cell populations in the larval Drosophila central nervous system. We identify a defined group of miRNAs that redundantly target all iconic genes known to control the transition from neuroblasts to neurons. In vivo functional studies demonstrate that knockdown of individual miRNAs does not induce detectable cellular phenotypes. However, simultaneous knockdown of multiple miRNAs leads to precocious stem cell differentiation, demonstrating functional interdependence. Thus, miRNAs cooperate within a regulatory module to specify the targeted gene network

Coopted temporal patterning governs cellular hierarchy, heterogeneity and metabolism in Drosophila neuroblast tumors

International audienceIt is still unclear what drives progression of childhood tumors. During Drosophila larval development, asymmetrically-dividing neural stem cells, called neuroblasts, progress through an intrinsic temporal patterning program that ensures cessation of divisions before adulthood. We previously showed that temporal patterning also delineates an early developmental window during which neuroblasts are susceptible to tumor initiation (Narbonne-Reveau et al., 2016). Using single-cell transcriptomics, clonal analysis and numerical modeling, we now identify a network of twenty larval temporal patterning genes that are redeployed within neuroblast tumors to trigger a robust hierarchical division scheme that perpetuates growth while inducing predictable cell heterogeneity. Along the hierarchy, temporal patterning genes define a differentiation trajectory that regulates glucose metabolism genes to determine the proliferative properties of tumor cells. Thus, partial redeployment of the temporal patterning program encoded in the cell of origin may govern the hierarchy, heterogeneity and growth properties of neural tumors with a developmental origin