Enolase, a plasminogen receptor isolated from salivary gland transcriptome of the ixodid tick Haemaphysalis flava

Abstract

Enolase, a multifunctional protein, is shown to act as a plasminogen receptor that contributes to fibrinolysis, which plays an important role in preventing the formation of blood clots during tick feeding. The study of enolase genes provides opportunities to develop a potential antigen target for tick control. So far, enolase has been identified in only a few species of ticks. Knowledge of the exact mechanisms of plasminogen activation and fibrinolysis by enolase as a plasminogen receptor is limited. Here, we cloned the enolase full-length complementary DNA (cDNA) from the salivary glands of Haemaphysalis flava, expressed it, and analyzed the function of the recombinant H. flava enolase. The enolase cDNA was 1988 bp in length and encoded 433 amino acid residues. It contained two domains and some highly conserved functional motifs including an assumed membrane re-association region "AAVPSGASTGI." The enolase exhibited 83.3 % amino acid similarity to that of the putative enolase of Ixodes ricinus, and 85 % to that of Ornithodoros moubata enolase. After eukaryotic expression in insect cells, Western blot analysis showed that the mouse antiserum against the hexahistidine-tagged recombinant enolase protein recognized a band of approximately 48 kDa. The recombinant enolase bound human plasminogen in a dose-dependent manner and enhanced plasminogen activation in the presence of host tissue plasminogen activator (t-PA), most probably to promote fibrinolysis and maintain blood flow at the host-tick interface. Real-time quantitative polymerase chain reaction (qPCR) analysis showed that the expression level of enolase in salivary glands was significantly higher than in other tested tissues. Although the enolase was expressed in all developmental stages, it had the highest expression in the rapid blood feeding period of ticks. These findings indicate that the enolase might play an important role in blood feeding of H. flava.

WITHDRAWN: Molecular characterization of enolase fromHaemaphysalis longicornistick

“…Saliva consists of anti-immunomodulatory, anti-vasodilator, anti-coagulant, anti-complement, and anti-platelet aggregation factors (Neelakanta and Sultana, 2022). Recently, sialo transcriptome analysis of H. flava performed by our research group (Xu et al, 2016) showed the presence of significant amounts of an enolase-like homologous protein in salivary fluid in the adult and nymph stages. Enolase, a secretory protein the tick injects into the host through its saliva, may play a part in the success of the tick’s feeding.…”

Proteomic informed by transcriptomic for salivary glands components of the camel tick Hyalomma dromedarii

“…The latter enzymes have also been identified in the saliva of Ornithodoros moubata and shown to act as a profibrinolytic plasminogen receptor, most likely helping the tick to maintain the fluidity of host blood during feeding [59]. In H. dromedarii , enolases might be specifically secreted into the saliva where they could act as anti-hemostatic, anti-inflammatory or immunomodulatory compounds [60].…”

“…Interestingly, a human homolog proteins containing these three domains, named MC25, has several effects in tissue growth and differentiation [58, 59]. It has also been shown to inhibit vascular endothelial growth factor and keratinocyte growth [60, 61]. Therefore, proteins belonging to this family, identified in this proteome, could serve as binders of growth factors affecting angiogenesis, tissue repair, and immunity [57].…”

Acquired tick resistance: The trail is hot

“…69 Haemaphysalis ticks have a particularly large number of demonstrated and putative thrombin inhibitors, including: madanin-1 and -2, 70,71 chimadanin, 72 and the serpin HLS1 73 from Haemaphysalis longicornis; haemathrin from Haemaphysalis bispinosa 74 ; and serpins HDS1 and HDS2 from H. doenitzi 75 Iris from I. ricinus 68 has also demonstrated anti-thrombin activity, in addition to BmGTI, 76 the serpin RmS-15, 77 BmAP, 78 and microphilin 79 from R. microplus. Haemaphysalis proteins longistatin 80 and enolase 81 stimulate this pathway by activating plasminogen that negatively regulates the coagulation cascade.…”