Rab27 in tick extracellular vesicle biogenesis and infection

Abstract

AbstractBackgroundThe blacklegged tick,Ixodes scapularis, transmits most vector-borne diseases in the United States. It vectors seven pathogens of public health relevance, including the emerging human pathogenAnaplasma phagocytophilum. Nevertheless, it remains critically understudied when compared to other arthropod vectors.I. scapularisreleases a variety of molecules that assist in the modulation of host responses. Recently, it was found that extracellular vesicles (EVs) carry several of these molecules and may impact microbial transmission to the mammalian host. EV biogenesis has been studied in mammalian systems and is relatively well understood, but the molecular players important for the formation and secretion of EVs in arthropods of public health relevance remain elusive. RabGTPases are among the major molecular players in mammalian EV biogenesis. They influence membrane identity and vesicle budding, uncoating, and motility.MethodsUsing BLAST, anin-silicopathway for EV biogenesis in ticks was re-constructed. We identified Rab27 for further study. EVs were collected from ISE6 tick cells after knocking downrab27to examine its role in tick EV biogenesis.I. scapularisnymphs were injected with small interfering RNAs to knock downrab27then fed on naïve andA. phagocytophiluminfected mice to explore the importance ofrab27in tick feeding and bacterial acquisition.ResultsOur BLAST analysis identified several of the proteins involved in EV biogenesis in ticks, including Rab27. We show that silencingrab27inI. scapularisimpacts tick fitness. Additionally, ticks acquire lessA. phagocytophilumafterrab27silencing. Experiments in the tick ISE6 cell line show that silencing ofrab27causes a distinct range profile of tick EVs, indicating that Rab27 is needed to regulate EV biogenesis.ConclusionsRab27 is needed for successful tick feeding and may be important for acquiringA. phagocytophilumduring a blood meal. Additionally, silencingrab27in tick cells results in a shift of extracellular vesicle size. Overall, we have observed that Rab27 plays a key role in tick EV biogenesis and the tripartite interactions among the vector, the mammalian host, and a microbe it encounters.