Hlcyst-1 and Hlcyst-2 Are Potential Inhibitors of HlCPL-A in the Midgut of the Ixodid Tick Haemaphysalis longicornis

Abstract

Although the actions of cysteine proteases are controlled in part by endogenous tight-binding cysteine protease inhibitors from the cystatin superfamily, regulatory mechanisms used by ticks to control protease activities are unknown. We report here the interaction of two endogenous midgut cysteine protease inhibitors, Hlcyst-1 and Hlcyst-2, with an endogenous midgut cysteine protease, HlCPL-A in Haemaphysalis longicornis. In vitro inhibition assays demonstrated that the hydrolytic activity of HlCPL-A was inhibited by Hlcyst-1 and Hlcyst-2 in dose dependent manner. Immunofluorescent studies revealed that Hlcyst-1 and Hlcyst-2 are co-localized with HlCPL-A in the epithelial cells of the midgut. The hemoglobin degradation activity of HlCPL-A was dose-dependently inhibited by Hlcyst-1 and Hlcyst-2. These results strongly indicate that, Hlcyst-1 and Hlcyst-2 are possible inhibitor of HlCPL-A and play a key role in regulatory mechanisms of hemoglobin degradation process in ticks.