Exploring Suppressor of IKK Epsilon's Interaction with Cytoskeletal Proteins

Abstract

The innate immune system rapidly responds to challenges by pathogens via activation of an inflammatory response. As a convergence point for multiple inflammatory and anti-viral signaling pathways, TANK binding kinase 1 (TBK1) serves as a catalytic hub to initiate host defenses. Suppressor of IKK epsilon (SIKE) is a newly identified TBK1 substrate that, due to its high affinity for the kinase, shifts TBK1's catalytic function towards SIKE phosphorylation and reduces type I interferon production via interferon regulatory factor 3/7 phosphorylation. The goal of this project was to identify the function of SIKE within the host's anti-viral response. Co-immunoprecipitation – tandem MS/MS analyses of the SIKE interaction network identified several interactions with cytoskeletal components. Immunofluorescence assays of endogenous SIKE with cytoskeletal markers and colocalization analyses indicated that SIKE colocalized with actin and a-actinin, but not focal adhesion kinase. Reciprocal immunoprecipitation (RcIPs) studies were used to confirm and further delineate the SIKE:actin interaction. RcIPs employed epitope tagged actin and full-length (residues 1-207), N-terminal (residues 1-112) or C –terminal (residues 113-207) FLAG-tagged SIKE. To determine the basal versus viral induced SIKE:actin interaction, studies were completed +/- double stranded RNA stimulation to mimic a viral infection. These studies support a role for SIKE in cytoskeletal rearrangements, such as cell migration and formation/resolution of RNA granules, that contribute to the antiviral response. This work was supported by NIH grant R21AI107447.