PLURIPOTENCY OF NOVEL TICK CYSTEINE PROTEASE INHIBITORS DURING HEMATOPHAGY

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Project Summary Ixodid ticks blood feed for several days to weeks on their hosts. Hematophagy is assisted by tick’s saliva, which is repeatedly injected into the host skin, alternating with blood intake. Tick saliva contains a mixture of bioactive molecules that target a broad spectrum of host defense mechanisms to allow the tick to blood feed on the vertebrate host for several days, many of which are host immunomodulators. Understanding the molecular interactions between tick vectors, vertebrate hosts, and pathogens is critical in developing strategies to combat ticks and tick-borne diseases. This proposal explores a fundamental mechanism of tick hematophagy success, mainly regulating vertebrate host proteolytic cascades at sites of tick feeding (skin) by multiple cysteine protease inhibitors of the cystatin family. We hypothesize that three novel Amblyomma maculatum cystatins modulate the vertebrate host immune response during blood-feeding. We will test the hypothesis by pursuing the following specific aims: 1) To complete the detailed functional characterization of three novel recombinant Amblyomma maculatum salivary cystatins as candidate host immunomodulators, 2) To investigate the role of each of the three A. maculatum salivary cystatin genes in tick feeding and pathogen transmission success by conditional knock- downs in ticks (RNAi) and, 3) To test the potential of artificially boosting the host humoral response against these three A. maculatum cystatins by vaccine studies in a preclinical animal model, with an ultimate goal that the achieved boost in the host antibody titers against tick cystatins will block their action at the tick-host interface and may impair tick feeding and/or pathogen transmission success. For Aim 2, we will also evaluate the use of bio-inspired hydrogels as non-invasive, transcuticular dsRNA delivery devices that could provide a commercially scalable approach for the delivery of these therapies to ticks in their natural environment. These aims will provide critical reagents, tools, and data to address the crucial gap in the fundamental knowledge of the role of redundancy of host immunomodulators in tick saliva.