Serpins (serine protease inhibitors) are a large family of structurally related proteins found in a wide variety of organisms, including hematophagous arthropods. Protein analyses revealed that Iris, previously described as an immunomodulator secreted in the tick saliva, is related to the leukocyte elastase inhibitor and possesses serpin motifs, including the reactive center loop (RCL), which is involved in the interaction between serpins and serine proteases. Only serine proteases were inhibited by purified recombinant Iris (rIris), whereas mutants L339A and A332P were found devoid of any protease inhibitory activity. The highest K a was observed with human leukocyteelastase, suggesting that elastase-like proteases are the natural targets of Iris. In addition, mutation M340R completely changed both Iris substrate specificity and affinity. This likely identified Met-340 as amino acid P1 in the RCL. The effects of rIris and its mutants were also tested on primary hemostasis, blood clotting, and fibrinolysis. rIris increased platelet adhesion, the contact phase-activated pathway of coagulation, and fibrinolysis times in a dose-dependent manner, whereas rIris mutant L339A affected only platelet adhesion. Taken together, these results indicate that Iris disrupts coagulation and fibrinolysis via the anti-proteolytic RCL domain. One or more other domains could be responsible for primary hemostasis inhibition. To our knowledge, this is the first ectoparasite serpin that interferes with both hemostasis and the immune response.Ticks are blood-sucking arthropods that infest a large variety of vertebrate hosts (mammals, birds, reptiles, and amphibians) in many parts of the world (1). To complete their blood meal, blood-sucking arthropods express a wide range of anti-hemostatic molecules in their saliva, including vasodilators, inhibitors of the platelet aggregation, and anti-coagulants (2). Tick saliva and salivary gland extracts are also known to modulate the host's defense mechanisms (3-6). Both anti-hemostatic and immunosuppressive compounds were identified, isolated, and characterized from soft and hard ticks. These compounds include histamine-binding proteins, tissue factor pathway inhibitor-like proteins, anti-thrombin-like proteins, and anticomplement factors (7-16).These last years, several laboratories reported the construction and screening of cDNA libraries from tick salivary glands. Thus, Das et al. (17) found 14 Ixodes scapularis immunodominant antigens, whereas Leboulle et al. (18) identified 27 mRNA, the expression of which is specifically induced or up-regulated during the Ixodes ricinus blood meal. Finally, Ribeiro and coworkers explored the sialome of the tick I. scapularis (19,20) and uncovered a large variety of putative bioactive agents. These studies all identified some serine protease inhibitors, containing serpin, kunitz, kazal, or ␣-macroglobulin motifs (21).To date, ϳ500 serpins have been identified in a large variety of species, including animals, viruses, and plants. On average, serpins are 35...

Objective Growing evidence suggests that a phenotypic switch converting pancreatic acinar cells to duct-like cells can lead to pancreatic intraepithelial neoplasia (PanIN) and eventually to invasive pancreatic ductal adenocarcinoma. Histologically, the onset of this switch is characterised by the co-expression of acinar and ductal markers in acini, a lesion called acinar-to-ductal metaplasia (ADM). Transcriptional regulators required to initiate ADM still remain unknown, yet need to be identified to characterise the regulatory networks that drive ADM. Here we investigate the role of the ductal transcription factors Hepatocyte Nuclear Factor 6 (HNF6, also known as Onecut1)and SRY-related HMG box factor 9 (Sox9) in ADM. Design Expression of HNF6 and Sox9 is measured by immunostaining in normal and diseased human pancreas. The function of the factors is tested in cultured cells and in mouse models of ADM by a combination of gain- and loss-of-function experiments. Results Expression of HNF6 and Sox9 is ectopically induced in acinar cells in human ADM, as well as in mouse models of ADM. We show that these factors are required for repression of acinar genes, for modulation of ADM-associated changes in cell polarity, and for activation of ductal genes in metaplastic acinar cells. Conclusions HNF6 and Sox9 are new biomarkers of ADM and constitute candidate targets for preventive therapy in cases when ADM may lead to cancer. Our work also highlights that ectopic activation of transcription factors may underlie metaplastic processes occurring in other organs.

Endothelial cells are required to initiate pancreas development from the endoderm. They also control the function of endocrine islets after birth. Here we investigate in developing pancreas how the endothelial cells become organized during branching morphogenesis and how their development affects pancreatic cell differentiation. We show that endothelial cells closely surround the epithelial bud at the onset of pancreas morphogenesis. During branching morphogenesis, the endothelial cells become preferentially located near the central (trunk) epithelial cells and remain at a distance from the branch tips where acinar cells differentiate. This correlates with predominant expression of the angiogenic factor vascular endothelial growth factor-A (VEGF-A) in trunk cells. In vivo ablation of VEGF-A expression by pancreas-specific inactivation of floxed Vegfa alleles results in reduced endothelial development and in excessive acinar differentiation. On the contrary, acinar differentiation is repressed when endothelial cells are recruited around tip cells that overexpress VEGF-A. Treatment of embryonic day 12.5 explants with VEGF-A or with VEGF receptor antagonists confirms that acinar development is tightly controlled by endothelial cells. We also provide evidence that endothelial cells repress the expression of Ptf1a, a transcription factor essential for acinar differentiation, and stimulate the expression of Hey-1 and Hey-2, two repressors of Ptf1a activity. In explants, we provide evidence that VEGF-A signaling is required, but not sufficient, to induce endocrine differentiation. In conclusion, our data suggest that, in developing pancreas, epithelial production of VEGF-A determines the spatial organization of endothelial cells which, in turn, limit acinar differentiation of the epithelium.

The migration of cortical projection neurons is a multistep process characterized by dynamic cell shape remodeling. The molecular basis of these changes remains elusive, and the present work describes how microRNAs (miRNAs) control neuronal polarization during radial migration. We show that miR-22 and miR-124 are expressed in the cortical wall where they target components of the CoREST/REST transcriptional repressor complex, thereby regulating doublecortin transcription in migrating neurons. This molecular pathway underlies radial migration by promoting dynamic multipolar-bipolar cell conversion at early phases of migration, and later stabilization of cell polarity to support locomotion on radial glia fibers. Thus, our work emphasizes key roles of some miRNAs that control radial migration during cerebral corticogenesis.

Iris is a specific elastase inhibitor expressed in the salivary glands of the hard tick Ixodes ricinus. It belongs to the superfamily of serpins and interferes with both haemostasis and the immune response of the host. In this study, we first show that Iris is expressed in nymphs but not in the female midgut nor in males. We also show that Iris is present in the saliva. To examine its potency as anti-tick vaccine candidate, we set up three models of I. ricinus infestation on immunized animals: nymphs on mice, and adults and nymphs on rabbits. We report the rise of neutralizing antibodies following immunization of rabbits and mice. This comes with a significant protective immunity against ticks in rabbits only, resulting in a 30% mortality rate and a diminution of weight gain in both nymphs and adults and a prolongation of blood feeding time in adults. This is the first report on an anti-tick vaccine trial on I. ricinus using a protein able to interact with both host immunity and haemostasis, as a vaccinating antigen.