Functional characterization of sensor DDX41 and mediator STING involved in the innate immune response to cytosolic DNA and cyclic dinucleotides

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DNA recognition in the cytosol activates interferon production. DDX41 is a DEAD-box protein that has been identified as one of the DNA sensors. STING is a membrane bound protein that acts as a mediator between the DNA sensors and the activation of interferon response. These two proteins have also been involved in activating cell innate immune reaction in response to cyclic dinucleotides, either of host or bacterial origin. So far it is unclear how DDX41 and STING activation and interaction function on a molecular level. Here, we plan to characterize DDX41 in vitro with respect to DNA and cyclic dinucleotide recognition and identify the conformational changes the protein undergoes upon ligand binding. In addition, we will use a combination of biochemical, biophysical and single molecule fluorescence techniques to understand the molecular events involved in STING activation. In particular we will I) investigate ligand-induced dimerization/oligomerization of STING, II) follow conformational changes of STING cytosolic domain and C-terminal tail upon cyclic dinucleotide binding, and III) study STING-DDX41 interaction to understand how STING is activated by sensor binding.