Investigating The Mechanisms Of Stimulator Of Interferon Genes (Sting) Subcellular Localisation
Stimulator of Interferon Genes (STING) is a critical component of the innate immune response against microbial infection. Activation of STING is induced following the recognition of pathogen-associated molecular patterns (PAMPs) present within the host cell cytosol during infection. Following activation, STING signalling promotes the expression of type I interferons and pro-inflammatory cytokines to induce an immune response. In the steady state, STING is located on the endoplasmic reticulum (ER) membrane. Following activation, STING traffics to the Golgi, which is important for recruitment of key kinases (e.g. TBK1) for downstream signalling events. To prevent sustained inflammation, STING signalling is ultimately terminated within endolysosomal compartments. However, many questions still remain regarding the timing and mechanisms of STING relocalisation from the ER, and how this controls inflammatory STING responses. To understand the kinetics of STING signalling in macrophages I am utilising fluorescent live-cell microscopy approaches to characterise the dynamics of STING translocation across organelles and association with key known interactors. Additionally I have performed a global phosphoproteomic screen in primary murine macrophages following STING activation over time. Gene ontology (GO) analysis of proteins undergoing phosphorylation changes has shown time-dependent signatures. This screening approach also identified several novel proteins and phosphorylation events, not previously associated with the STING pathway. I am now using CRISPR/Cas9 editing to investigate the role of these proteins in regulating the STING response using both biochemical approaches and imaging tools I have generated. Aberrant cGAS-STING activation and/or disruption of STING localisation has been implicated in the pathogenesis of several autoimmune and inflammatory conditions. Therefore, understanding STING trafficking is important for effective therapeutic targeting of STING-associated inflammatory disease.