Arthropod saliva in vector-borne disease transmission: Functional analysis of novel domains in salivary secretions

National Institute Of Allergy And Infectious Diseases

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Abstract

The main accomplishments of the Molecular Entomology Unit are: 1. We described the function of Guianensin, a Simulium guianense salivary protein, with broad anti-hemostatic and anti-inflammatory properties. Guianensin was identified in the sialotranscriptome of adult S. guianense flies and belongs to the Kunitz domain of protease inhibitors. It targets various serine proteases involved in hemostasis and inflammation. Accordingly, Guianensin significantly increased both PT (Prothrombin time) and aPTT (Activated partial thromboplastin time) in human plasma and consequently increased blood clotting time ex vivo. Guianensin also inhibited prothrombinase activity on endothelial cells and acts as a potent anti-inflammatory molecule on FXa-induced paw edema formation in vivo. We hypothesize that the secretion of Guianensin at the bite site is an evolutionary strategy to block host proteases that could facilitate blood meal acquisition by inhibiting coagulation and inflammation. This work is another example of the complexity of the biology of blood-feeding arthropods and highlights the potential use of vector salivary proteins as therapeutic applications and potential targets for transmission-blocking vaccines. 2. Blood-feeding arthropods secrete potent salivary molecules, which include platelet aggregation inhibitors, vasodilators, and anticoagulants. Among these molecules, Alboserpin, the major salivary anticoagulant from the mosquito vector Aedes albopictus, is a specific inhibitor of the human coagulation factor Xa (FXa). Here, showed that Alboserpin also displays strong anti-inflammatory activity in vivo and in vitro. Alboserpin reduced proinflammatory cytokine and chemokine release; decreased ERK1/2 expression and protease-activated receptor (PAR) signaling; and suppressed VCAM, ICAM, and NF-b gene expression. Furthermore, Alboserpin significantly reduced paw edema in vivo and attenuated inflammatory cytokine and chemokine release. Alboserpin also inhibited the ability of FXa to disrupt endothelial permeability in vitro and in vivo. This salivary molecule plays a key role blood feeding and may be implicated in arbovirus transmission. 3. Utilizing cryogenic electron microscopy (cryo-EM), we solved the first atomic structure of a native salivary protein from the Yellow Fever mosquito Aedes aegypti. We previously showed that A. aegypti salivary gland surface protein 1 (SGS1) facilitates P. gallinaceum transmission, but its mechanism remains elusive. We determined the native structure of SGS1 by cryoID and showed that the 3364 amino-acid protein has a Tc toxin-like Rhs/YD shell, four receptor domains, and a set of C-terminal daisy-chained helices. These helices are partially shielded inside the Rhs/YD shell and poised to transform into predicted transmembrane helices. This transformation, and the numerous receptor domains on the surface of SGS1, are likely key in facilitating sporozoite/arbovirus invasion into the salivary glands and manipulating host immunity. This work offers valuable insight into the possible interactions of SG1 with mosquito-borne pathogens. 4. An update insight into the sialotranscriptome of Culex quinquefasciatus, a vector of filaria parasites and West Nile virus. We revisited C. quinquefasciatus salivary gland contents using an Illumina-based sequencing approach of both male and female tissues. We identified 7,539 coding DNA sequences (CDS) functionally annotated into 25 classes. Comparison of male and female libraries allowed for the identification of female-enriched transcripts that are potentially related to blood acquisition and virus transmission. Together, these findings represent an extended reference for the identification and characterization of the proteins containing relevant pharmacological activity in the salivary glands of C. quinquefasciatus mosquitoes. The updated sialotranscriptome study reported here provided novel data on our knowledge of the salivary repertoire of this mosquito.