Homeostasis of the intestinal epithelium during a bacterial infection by Serratia marcescens in Drosophila melanogaster – DROSOGUT

These involve microscopy, transcriptomics at various stages of an early phase (fast degradation followed by epithelial recovery within a few hours). The early phase phenotype of mutants isolated in the genetic screen and from genes identified by transcriptomics is being documented.

A pore-forming toxin, hemolysin, is responsible for the early degradation of the midgut epithelium. Enterocytes are not killed but become thinner and display multiple signs of intense stress. We have found that several processes at the cellular level are involved in the recovery phase. Indeed, about half of the tested genes display an impaired regeneration after Serratia hemolysin attack.

Endurance mechanisms are likely to have been conserved throughout evolution. Hence, our discoveries may be relevant to a fuller understanding of mammalian host defense against ingested pathogens.

Talks at conferences in Europe, America, and Africa

The host response to an infection encompasses two major dimensions: the immune response to the invading pathogens and the ability of the host to withstand and repair damage inflicted either directly by the pathogen itself or indirectly by the host's immune response. Thus, host defense comprises two complementary facets : resistance and "tolerance" to infection. While the former has been intensively studied, the latter remains relatively ill-defined.

We are using the fruitfly Drosophila melanogaster as a genetic model organism to understand host defense against infections. The humoral systemic immune response has been well-studied, using mostly nonpathogenic microorganisms. In contrast, we have developed an intestinal infection model with the potent entomopathogen Serratia marcescens in which flies apparently succumb ultimately to unrecoverable gut damage. In a previous ANR grant, we have shown that host defense against such infections takes place at least at two levels : i) a mucosal immune response in the gut epithelium that is regulated by one of the pathways that controls the expression of antimicrobial peptides during the systemic immune response, ii) phagocytosis of bacteria that have crossed the gut barrier by hemocytes. We have performed the first ever genome-wide screen to identify genes required for either enhanced or decreased survival to ingested S. marcescens. We have thus identified about 160 genes that are acting in the midgut epithelium, most of which have unknown functions. We have found that intensive damages to enterocytes that result in apoptosis are compensated by the proliferation of intestinal stem cells (ISCs) under the control of the JAK-STAT pathway. Thus, host defense is not limited to classical immune response pathways but encompasses the homeostasis of epithelia, a "tolerance" mechanism.

Here, we propose four aims to investigate further our model. In Aim1, we plan an extended transcriptomics analysis of midguts during the different phases of the infection. This will be complemented by a systematic phenotypic analysis of the hits found in our screen so as to focus in greater depth on the most important genes. In Aim2, we investigate the mechanisms involved in the fast division cycles of ISCs that apparently take place during infection. We study especially a candidate gene that is specifically required for these fast cell cycles. We also tackle the problem of the differentiation of the ISCs in enterocytes by testing an hypothesis suggested by our preliminary data. In a third Aim, we dissect the exact mode of action of the JAK-STAT pathway that is required both for driving ISC proliferation but also participates in the differentiation of enteroblasts. Finally, we ask whether the thin extensions emitted apically by ISCs are functionally relevant to epithelial homeostasis.

We expect to gain a detailed understanding of the processes at play during intestinal epithelium regeneration and to better understand in vivo how the apparently simple JAK-STAT pathway is actually regulated. As there is much similarity at the molecular level between the Drosophila midgut epithelium and that of vertebrates, we hope that this project will ultimately allow a better understanding of gut physiology during inflammation and cancer initiation. The study of the gene required for fast cycles may be relevant to similar mechanisms at work, for instance in small cell lung cancer.