A tick saliva serpin, IxsS17 inhibits host innate immune system proteases and enhances host colonization by Lyme disease agent

Abstract

Lyme disease (LD) caused by Borrelia burgdorferi is among the most important human vector borne diseases for which there is no effective prevention method. Identification of tick saliva transmission factors of the LD agent is needed before the highly advocated tick antigen-based vaccine could be developed. We previously reported the highly conserved Ixodes scapularis (Ixs) tick saliva serpin (S) 17 (IxsS17) was highly secreted by B. burgdorferi infected nymphs. Here, we show that IxsS17 promote tick feeding and enhances B. burgdorferi colonization of the host. We show that IxsS17 is not part of a redundant system, and its functional domain reactive center loop (RCL) is 100% conserved in all tick species. Yeast expressed recombinant (r) IxsS17 inhibits effector proteases of inflammation, blood clotting, and complement innate immune systems. Interestingly, differential precipitation analysis revealed novel functional insights that IxsS17 interacts with both effector proteases and regulatory protease inhibitors. For instance, rIxsS17 interacted with blood clotting proteases, fXII, fX, fXII, plasmin, and plasma kallikrein alongside blood clotting regulatory serpins (antithrombin III and heparin cofactor II). Similarly, rIxsS17 interacted with both complement system serine proteases, C1s, C2, and factor I and the regulatory serpin, plasma protease C1 inhibitor. Consistently, we validated that rIxsS17 dose dependently blocked deposition of the complement membrane attack complex via the lectin complement pathway and protected complement sensitive B. burgdorferi from complement-mediated killing. Likewise, co-inoculating C3H/HeN mice with rIxsS17 and B. burgdorferi significantly enhanced colonization of mouse heart and skin organs in a reverse dose dependent manner. Taken together, our data suggests an important role for IxsS17 in tick feeding and B. burgdorferi colonization of the host.

<em>Ixodes scapularis</em> Calreticulin Binds Complement Proteins but Does Not Protect <em>Borrelia burgdorferi </em>from Complement Killing

“…To identify plasma proteins that might interact with tick saliva CRT, protein-to-protein interaction using differential precipitation was conducted as published [33,34]. Briefly, the reaction volume of 150μl was prepared by preincubating rIxsCRT (10 μg) with 10% Normal Human Serum (NHS) for 90min at 37˚C.…”

“…The effects of rIxsCRT of complement activation was assayed using the Wieslab ® Complement System Screen (Svar Life Science AB, Malmo, Sweden) according to the manufacturer's recommendation and as previously published [34,36]. This kit evaluates effects on membrane attack complex (MAC) deposition via each of the three complement system activation pathways: Classical, Lectin, and Alternative.…”