Molecular characterization of tick salivary gland glutaminyl cyclase

Abstract

Glutaminyl cyclase (QC) catalyzes the cyclization of N-terminal glutamine residues into pyroglutamate. This post-translational modification extends the half-life of peptides and, in some cases, is essential in binding to their cognate receptor. Due to its potential role in the post-translational modification of tick neuropeptides, we report the molecular, biochemical and physiological characterization of salivary gland QC during the prolonged blood-feeding of the black-legged tick (Ixodes scapularis) and the gulf-coast tick (Amblyomma maculatum). QC sequences from I. scapularis and A. maculatum showed a high degree of amino acid identity to each other and other arthropods and residues critical for zinc-binding/catalysis (D159, E202, and H330) or intermediate stabilization (E201, W207, D248, D305, F325, and W329) are conserved. Analysis of QC transcriptional gene expression kinetics depicts an upregulation during the blood-meal of adult female ticks prior to fast feeding phases in both I. scapularis and A. maculatum suggesting a functional link with blood meal uptake. QC enzymatic activity was detected in saliva and extracts of tick salivary glands and midguts. Recombinant QC was shown to be catalytically active. Furthermore, knockdown of QC-transcript by RNA interference resulted in lower enzymatic activity, and small, unviable egg masses in both studied tick species as well as lower engorged tick weights for I. scapularis. These results suggest that the post-translational modification of neurotransmitters and other bioactive peptides by QC is critical to oviposition and potentially other physiological processes. Moreover, these data suggest that tick-specific QC-modified neurotransmitters/hormones or other relevant parts of this system could potentially be used as novel physiological targets for tick control.

Transcriptional activation of antioxidants may compensate for selenoprotein deficiencies in Amblyomma maculatum (Acari: Ixodidae) injected with selK‐ or selM‐dsRNA

“…Tissues and saliva were collected from ticks seven days post-infestation. Soluble protein extracts from tick salivary glands and midguts were prepared as previously described (Adamson et al 2013). Saliva collected from GFP-dsRNA injected ticks was the control.…”

“…Our laboratory is investigating the impact of antioxidants within the framework of tick feeding, oviposition, and R. parkeri acquisition, maintenance, trafficking, and transmission (Adamson et al, 2013). SelM and SelK represent two interesting candidates for further study.…”

Immunoprotective potential of a Rhipicephalus (Boophilus) microplus metalloprotease

“…MPs have been identified in various tick species, and are considered crucial for maintenance of blood meal related functions (Francischetti et al, 2003;Harnnoi et al, 2007;Decrem et al, 2008a;Barnard et al, 2012). The up-regulation of MPs transcription in salivary glands has been correlated with tick blood feeding (Francischetti et al, 2003;Harnnoi et al, 2007;Decrem et al, 2008aDecrem et al, , 2008bBarnard et al, 2012;Adamson et al, 2013). In Ixodes scapularis, MPs are secreted in saliva, playing an essential role in fibrin and fibrinogen breakdown (Francischetti et al, 2003).…”

“…Despite the characterization of some tick MPs (Francischetti et al, 2003;Harnnoi et al, 2007;Decrem et al, 2008aDecrem et al, , 2008bBarnard et al, 2012;Adamson et al, 2013), vaccination trials against R. microplus infestation using these proteins as antigen remains to be evaluated. It has been speculated that the cysteine-rich domain in MPs is a limiting factor in the recombinant production of these proteins by prokaryotes (Ramos and SelistreDe-Araujo, 2006), since a native-like conformation of recombinant MPs is supposed to be critical -as far as retaining immunogenic properties is concerned.…”

Probing the functional role of tick metalloproteases

“…The increase in gene transcription of this protein during the blood meal of adult female ticks of both species indicates that QCs have a functional role in blood feeding. Glutaminyl cyclase enzyme activity is found in the saliva and in salivary gland and midgut extracts of ticks (Adamson et al ., ). Additionally, gene‐specific RNA interference (RNAi) decreases Is‐QC activity, and leads to small and unviable egg masses in both tick species ( I scapularis , A. maculatum ), as well as lower engorged weights for I. scapularis females when feeding on the host.…”