Investigating the role of gC1qR in Enterovirus A71-induced apoptosis

Abstract

Enterovirus A71 (EV-A71) is a non-enveloped virus which contains a single-stranded positive RNA genome encapsidated by four viral structural proteins (VPs) VP1 VP2 VP3 and VP4 Infection of EV-A71 causes an array of childhood diseases with severe neurological manifestations and has been shown to induce mitochondrial-associated apoptotic signaling and caspase-mediated signaling events in many types of cells Cellular apoptosis is a crucial cellular defense to account for EV-A71 invasion; however it remains largely unclear the spatiotemporal control of viral and cellular factors that are utilized by invading EV-A71 for the establishment of productive infection versus the initiation of apoptosis Our lab had previous identified a cellular interactome of VP4 by a combined tandem-tag affinity purification (TAP) technique and LC/MS-MS-based protein identification approach Among the 39 VP4-interacting cellular candidates receptor of the globular heads of C1q complement (gC1qR) shows the highest identification fidelity and is ubiquitously present in cells Our previous finding supports intimate gC1qR-VP4 interaction and co-localization in cytoplasm and mitochondria in infected RD cells by immunoprecipitation and confocal imagines In this study we step further the role of gC1qR in mitochondria-associated apoptosis because gC1qR is capable of entering mitochondria to modulate cellular defense machinery Firstly our results found that infection of RD cells with 5 MOI of EV-A71 resulted in increased qC1qR and viral structural protein during the productive phase of viral growth within 4-8 hours The caspase-3 was also activated after 4 hours post-infection EV-A71 infection of gC1qR-knockdown RD cells in comparison to infected sh-control cells promoted intracellular VPs production and numbers of apoptotic cells with flipped phosphatidylserine (PS) protein on the outer cell membrane Moreover VPs were found to translocate to the mitochondria which may cause mitochondrial stress as mitochondrial membrane potential were lost quickly after infection alone with increasing levels of cleaved forms of caspase-9 and caspase-3 These results suggested that gC1qR is required to negatively modulate EV-A71-induced apoptosis and productive viral production in RD cells Indeed EV-A71 infection of gC1qR-overexpressing RD cells in comparison to infected RD cells resulted in decreased VPs and membrane PS translocation Also gC1qR-overexpression protected the loss of MMP that occurred during EV-A71 infection which accompanied with decreased VPs in the cytosolic and mitochondrial fraction as well as decreased activation of caspase-9 and caspase-3 These results implied that endogenous gC1qR may interact with VP4 to retard early apoptosis and accumulate through time to negatively modulate intracellular VPs biogenesis via viral translation or cellular degradation machinery yet the in-depth mechanism remains to be clarified In conclusion this study provides a preliminary evidence that gC1qR has a novel role during EV-A71 infection to spatiotemporally modulate cellular apoptosis EV-A71-induced mitochondria-associated downstream apoptotic signaling can be retarded through overexpressing gC1qR which is likely to be critical for EV-A71 to establish early- to mid-stage (4-6 h) of intracellular viral production while accumulated gC1qR is capable of negative modulating viral protein production which is likely to occur during mid- to late-stage (6-8 h) of infection when viral protein production is no longer needed

Date of Award	2016 Jan 28
Original language	English
Supervisor	Shainn-Wei Wang (Supervisor)