Immune regulation of intestinal-stem-cell function in Drosophila.

Intestinal progenitor cells integrate signals from their niche, and the gut lumen, to divide and differentiate at a rate that maintains an epithelial barrier to microbial invasion of the host interior. Despite the importance of evolutionarily conserved innate immune defenses to maintain stable host-microbe relationships, we know little about contributions of stem-cell immunity to gut homeostasis. We used Drosophila to determine the consequences of intestinal-stem-cell immune activity for epithelial homeostasis. We showed that loss of stem-cell immunity greatly impacted growth and renewal in the adult gut. In particular, we found that inhibition of stem-cell immunity impeded progenitor-cell growth and differentiation, leading to a gradual loss of stem-cell numbers with age and an impaired differentiation of mature enteroendocrine cells. Our results highlight the importance of immune signaling in stem cells for epithelial function in the adult gut.

Differential effects of commensal bacteria on progenitor cell adhesion, division symmetry and tumorigenesis in the Drosophila intestine.

Microbial factors influence homeostatic and oncogenic growth in the intestinal epithelium. However, we know little about immediate effects of commensal bacteria on stem cell division programs. In this study, we examined the effects of commensal Lactobacillus species on homeostatic and tumorigenic stem cell proliferation in the female Drosophila intestine. We identified Lactobacillus brevis as a potent stimulator of stem cell divisions. In a wild-type midgut, L.brevis activates growth regulatory pathways that drive stem cell divisions. In a Notch-deficient background, L.brevis-mediated proliferation causes rapid expansion of mutant progenitors, leading to accumulation of large, multi-layered tumors throughout the midgut. Mechanistically, we showed that L.brevis disrupts expression and subcellular distribution of progenitor cell integrins, supporting symmetric divisions that expand intestinal stem cell populations. Collectively, our data emphasize the impact of commensal microbes on division and maintenance of the intestinal progenitor compartment.

Vibrio cholerae-Symbiont Interactions Inhibit Intestinal Repair in Drosophila.

Pathogen-mediated damage to the intestinal epithelium activates compensatory growth and differentiation repair programs in progenitor cells. Accelerated progenitor growth replenishes damaged tissue and maintains barrier integrity. Despite the importance of epithelial renewal to intestinal homeostasis, we know little about the effects of pathogen-commensal interactions on progenitor growth. We find that the enteric pathogen Vibrio cholerae blocks critical growth and differentiation pathways in Drosophila progenitors, despite extensive damage to epithelial tissue. We show that the inhibition of epithelial repair requires interactions between the Vibrio cholerae type six secretion system and a community of common symbiotic bacteria, as elimination of the gut microbiome is sufficient to restore homeostatic growth in infected intestines. This work highlights the importance of pathogen-symbiont interactions for intestinal immune responses and outlines the impact of the type six secretion system on pathogenesis.

Constitutive Immune Activity Promotes Tumorigenesis in Drosophila Intestinal Progenitor Cells.

Gut innate immune defenses control bacterial populations and protect the host interior from invasion. Although excess intestinal immune activity frequently promotes inflammatory illnesses, we know little about the consequences of chronic innate immune activity exclusively in endodermal gut cells of an otherwise normal animal. To address this question, we examined the consequences of persistent inflammatory signals in adult fly intestinal progenitor cells. We found that constitutive immune activity disrupts expression of homeostatic regulators such as Notch pathway components and induces hyperplasia throughout the gut. Consistent with these observations, we found that persistent immune signals interfere with progenitor cell differentiation and exacerbate the formation of Notch-dependent intestinal tumors. These findings uncover a link between constitutive immune activity and tumorigenesis in intestinal stem cells.

Comparative evaluation of the genomes of three common Drosophila-associated bacteria.

Drosophila melanogaster is an excellent model to explore the molecular exchanges that occur between an animal intestine and associated microbes. Previous studies in Drosophila uncovered a sophisticated web of host responses to intestinal bacteria. The outcomes of these responses define critical events in the host, such as the establishment of immune responses, access to nutrients, and the rate of larval development. Despite our steady march towards illuminating the host machinery that responds to bacterial presence in the gut, there are significant gaps in our understanding of the microbial products that influence bacterial association with a fly host. We sequenced and characterized the genomes of three common Drosophila-associated microbes: Lactobacillus plantarum, Lactobacillus brevis and Acetobacter pasteurianus For each species, we compared the genomes of Drosophila-associated strains to the genomes of strains isolated from alternative sources. We found that environmental Lactobacillus strains readily associated with adult Drosophila and were similar to fly isolates in terms of genome organization. In contrast, we identified a strain of A. pasteurianus that apparently fails to associate with adult Drosophila due to an inability to grow on fly nutrient food. Comparisons between association competent and incompetent A. pasteurianus strains identified a short list of candidate genes that may contribute to survival on fly medium. Many of the gene products unique to fly-associated strains have established roles in the stabilization of host-microbe interactions. These data add to a growing body of literature that examines the microbial perspective of host-microbe relationships.

A deregulated intestinal cell cycle program disrupts tissue homeostasis without affecting longevity in Drosophila.

Recent studies illuminate a complex relationship between the control of stem cell division and intestinal tissue organization in the model system Drosophila melanogaster. Host and microbial signals drive intestinal proliferation to maintain an effective epithelial barrier. Although it is widely assumed that proliferation induces dysplasia and shortens the life span of the host, the phenotypic consequences of deregulated intestinal proliferation for an otherwise healthy host remain unexplored. To address this question, we genetically isolated and manipulated the cell cycle programs of adult stem cells and enterocytes. Our studies revealed that cell cycle alterations led to extensive cell death and morphological disruptions. Despite the extensive tissue damage, we did not observe an impact on longevity, suggesting a remarkable degree of plasticity in intestinal function.

Defense genes missing from the flight division.

Birds have a smaller repertoire of immune genes than mammals. In our efforts to study antiviral responses to influenza in avian hosts, we have noted key genes that appear to be missing. As a result, we speculate that birds have impaired detection of viruses and intracellular pathogens. Birds are missing TLR8, a detector for single-stranded RNA. Chickens also lack RIG-I, the intracellular detector for single-stranded viral RNA. Riplet, an activator for RIG-I, is also missing in chickens. IRF3, the nuclear activator of interferon-beta in the RIG-I pathway is missing in birds. Downstream of interferon (IFN) signaling, some of the antiviral effectors are missing, including ISG15, and ISG54 and ISG56 (IFITs). Birds have only three antibody isotypes and IgD is missing. Ducks, but not chickens, make an unusual truncated IgY antibody that is missing the Fc fragment. Chickens have an expanded family of LILR leukocyte receptor genes, called CHIR genes, with hundreds of members, including several that encode IgY Fc receptors. Intriguingly, LILR homologues appear to be missing in ducks, including these IgY Fc receptors. The truncated IgY in ducks, and the duplicated IgY receptor genes in chickens may both have resulted from selective pressure by a pathogen on IgY FcR interactions. Birds have a minimal MHC, and the TAP transport and presentation of peptides on MHC class I is constrained, limiting function. Perhaps removing some constraint, ducks appear to lack tapasin, a chaperone involved in loading peptides on MHC class I. Finally, the absence of lymphotoxin-alpha and beta may account for the observed lack of lymph nodes in birds. As illustrated by these examples, the picture that emerges is some impairment of immune response to viruses in birds, either a cause or consequence of the host-pathogen arms race and long evolutionary relationship of birds and RNA viruses.

Avian influenza rapidly induces antiviral genes in duck lung and intestine.

Ducks are the natural reservoir of influenza A and survive infection by most strains. To characterize the duck immune response to influenza, we sought to identify innate immune genes expressed early in an infection. We used suppressive subtractive hybridization (SSH) to construct 3 libraries enriched in differentially expressed genes from lung RNA of a duck infected with highly pathogenic avian influenza virus A/Vietnam/1203/04 (H5N1), or lung and intestine RNA of a duck infected with low pathogenic avian influenza A/mallard/BC/500/05 (H5N2) compared to a mock-infected duck. Sequencing of 1687 clones identified a transcription profile enriched in genes involved in antiviral defense and other cellular processes. Major histocompatibility complex class I (MHC I), interferon induced protein with tricopeptide repeats 5 (IFIT5), and 2'-5' oligoadenylate synthetase-like gene (OASL) were increased more than 1000-fold in relative transcript abundance in duck lung at 1dpi with highly pathogenic VN1203. These genes were induced much less in lung or intestine following infection with low pathogenic BC500. The expression of these genes following infection suggests that ducks initiate an immediate and robust response to a potentially lethal influenza strain, and a minimal response to a low pathogenic strain.