We have identified and characterized a Macrophage Migration Inhibitory Factor (MIF) family member in the Lophotrochozoan invertebrate, Biomphalaria glabrata, the snail intermediate host of the human blood fluke Schistosoma mansoni. In mammals, MIF is a widely expressed pleiotropic cytokine with potent pro-inflammatory properties that controls cell functions such as gene expression, proliferation or apoptosis. Here we show that the MIF protein from B. glabrata (BgMIF) is expressed in circulating immune defense cells (hemocytes) of the snail as well as in the B. glabrata embryonic (Bge) cell line that has hemocyte-like features. Recombinant BgMIF (rBgMIF) induced cell proliferation and inhibited NO-dependent p53-mediated apoptosis in Bge cells. Moreover, knock-down of BgMIF expression in Bge cells interfered with the in vitro encapsulation of S. mansoni sporocysts. Furthermore, the in vivo knock-down of BgMIF prevented the changes in circulating hemocyte populations that occur in response to an infection by S. mansoni miracidia and led to a significant increase in the parasite burden of the snails. These results provide the first functional evidence that a MIF ortholog is involved in an invertebrate immune response towards a parasitic infection and highlight the importance of cytokines in invertebrate-parasite interactions. Schistosoma mansoni, a parasitic blood fluke that causes intestinal schistosomiasis in humans, requires an intermediate host, a freshwater snail of the genus Biomphalaria for its transmission. Infection of the snail triggers marked cellular and humoral immune responses, but the molecular mechanisms of these responses are not well known. We have identified and characterized the involvement of a snail homologue of the cytokine MIF (Macrophage Migration Inhibitory Factor) in the snail immune responses to infection by the parasite. By using biochemical and molecular approaches in combination with in vitro culture and in vivo gene knock down, we have demonstrated the role of snail MIF in the regulation of the snail innate immune system. In particular, MIF regulates the proliferation and activation of the hemocytes, the macrophage-like snail defense cells, and the encapsulation response. This shows for the first time that MIF has a conserved cytokine function in an invertebrate and underlines the interest of the schistosome-snail model in the study of innate immunity.

Inactivation of host Rho GTPases is a widespread strategy employed by bacterial pathogens to manipulate mammalian cellular functions and avoid immune defenses. Some bacterial toxins mimic eukaryotic Rho GTPase-activating proteins (GAPs) to inactivate mammalian GTPases, probably as a result of evolutionary convergence. An intriguing question remains whether eukaryotic pathogens or parasites may use endogenous GAPs as immune-suppressive toxins to target the same key genes as bacterial pathogens. Interestingly, a RhoGAP domain–containing protein, LbGAP, was recently characterized from the parasitoid wasp Leptopilina boulardi, and shown to protect parasitoid eggs from the immune response of Drosophila host larvae. We demonstrate here that LbGAP has structural characteristics of eukaryotic RhoGAPs but that it acts similarly to bacterial RhoGAP toxins in mammals. First, we show by immunocytochemistry that LbGAP enters Drosophila immune cells, plasmatocytes and lamellocytes, and that morphological changes in lamellocytes are correlated with the quantity of LbGAP they contain. Demonstration that LbGAP displays a GAP activity and specifically interacts with the active, GTP-bound form of the two Drosophila Rho GTPases Rac1 and Rac2, both required for successful encapsulation of Leptopilina eggs, was then achieved using biochemical tests, yeast two-hybrid analysis, and GST pull-down assays. In addition, we show that the overall structure of LbGAP is similar to that of eukaryotic RhoGAP domains, and we identify distinct residues involved in its interaction with Rac GTPases. Altogether, these results show that eukaryotic parasites can use endogenous RhoGAPs as virulence factors and that despite their differences in sequence and structure, eukaryotic and bacterial RhoGAP toxins are similarly used to target the same immune pathways in insects and mammals. Recent data have highlighted strong similarities between insect and mammalian immune pathways, raising the question whether they might be similarly targeted by parasites/pathogens. Endoparasitoid wasps are insect mortality agents that perform their development inside their host. They are widely used in biological control, but little is known of the way in which they disable the host immune response. Here, we report the first characterization to our knowledge of a protein–protein interaction between a parasitoid immune-suppressive factor, LbGAP, and its host targets, thus explaining its physiological effects on host immune cells. Strikingly, bacterial toxins containing a domain functionally related to that of LbGAP are known to alter human macrophages by targeting the same molecules. Demonstration that insect and mammalian pathogens/parasites, whether eukaryotic or prokaryotic, can use evolutionary convergent molecules “built” to target the same key factors in immune pathways, represents a new step in understanding the evolution of host–pathogen interactions. In addition to the potential of discovering new bioinsectides, studying virulence strategies of insect parasites might thus help to better characterize vertebrate–pathogen interactions and to assess the role of evolutionary convergence in the evolution of virulence.