Salivary Tick Cystatin OmC2 Targets Lysosomal Cathepsins S and C in Human Dendritic Cells.

Tina Zavašnik-Bergant,Andreja Sekirnik,Marko Fonović,Jiří Salát,Lenka Grunclová,Petr Kopáček,Robert Vidmar,Boris Turk

doi:10.3389/fcimb.2017.00288

Abstract

To ensure successful feeding tick saliva contains a number of inhibitory proteins that interfere with the host immune response and help to create a permissive environment for pathogen transmission. Among the potential targets of the salivary cystatins are two host cysteine proteases, cathepsin S, which is essential for antigen- and invariant chain-processing, and cathepsin C (dipeptidyl peptidase 1, DPP1), which plays a critical role in processing and activation of the granule serine proteases. Here, the effect of salivary cystatin OmC2 from Ornithodoros moubata was studied using differentiated MUTZ-3 cells as a model of immature dendritic cells of the host skin. Following internalization, cystatin OmC2 was initially found to inhibit the activity of several cysteine cathepsins, as indicated by the decreased rates of degradation of fluorogenic peptide substrates. To identify targets, affinity chromatography was used to isolate His-tagged cystatin OmC2 together with the bound proteins from MUTZ-3 cells. Cathepsins S and C were identified in these complexes by mass spectrometry and confirmed by immunoblotting. Furthermore, reduced increase in the surface expression of MHC II and CD86, which are associated with the maturation of dendritic cells, was observed. In contrast, human inhibitor cystatin C, which is normally expressed and secreted by dendritic cells, did not affect the expression of CD86. It is proposed that internalization of salivary cystatin OmC2 by the host dendritic cells targets cathepsins S and C, thereby affecting their maturation.

Highlights

As external parasites, ticks have adapted to their hosts to successfully feed while warding off the host immune system
Recombinant cystatin OmC2 from insect cells was preferentially applied for two reasons; (1) it was added to the immune cells later on, and (2) it was expressed with His-tag at the C-terminal end of cystatin molecule, i.e., at the part which does not participate in binding to the active site of targeted proteases
Two additional species were determined, the first of which corresponded to cystatin OmC2 that was N-terminally conjugated with Alexa Fluor 488 (13629 Da), and the second was a minor species that corresponded to the additional labeling of a lysine side chain (14147 Da)

Summary

Introduction

Ticks have adapted to their hosts to successfully feed while warding off the host immune system. The saliva of ticks contains multiple proteins with anti-haemostatic, antiinflammatory and immunomodulatory properties (Díaz-Martin et al, 2013). Tick saliva influences the production and secretion of numerous cytokines, often simultaneously, from various types of immune cells (Kotál et al, 2015). The tick salivary proteins inhibit the components of the host immune system (Francischetti et al, 2009) and have an impact on the transmission of vector-borne pathogens. Many species of Borrelia causing relapsing fever in infected humans, are transmitted by the soft ticks of Ornithodoros genus (Parola and Raoult, 2001).

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Conclusion