Suppressor of IKKepsilon forms direct interactions with cytoskeletal proteins, tubulin and alpha-actinin, linking innate immunity to the cytoskeleton

ABSTRACT: Suppressor of IKKepsilon (SIKE) is associated with the innate immune system’s type I interferon response through TANK binding kinase 1 (TBK1). Characterized as an endogenous inhibitor of TBK1 when overexpressed in viral infection and pathological cardiac hypertrophic models, mechanistic study revealed SIKE acted as a high affinity substrate of TBK1, but SIKE’s function remains unknown. This study aimed to investigate the SIKE interaction network to identify by association a potential function for SIKE. Our affinity purification/mass spectrometry results showed that SIKE formed interactions with cytoskeletal proteins, nucleic acid-associated proteins, enzymes, and chaperones. In immunofluorescence assays, endogenous SIKE localized to cytosolic puncta in epithelial and myeloid cells whilst in epithelial cells additional staining occurred in stress fiber-like structures and adjacent to the plasma membrane, whereas myeloid cells revealed SIKE associated with nuclear puncta. Using cellular markers, significant co-occurrence of SIKE fluorescence with actin, a-actinin, tubulin, ezrin, FAK and b-catenin was detected. Reciprocal immunoprecipitation and in vitro precipitation assays confirmed a direct SIKE interaction with tubulin and a-actinin. These results indicate that SIKE directly associates with the cytoskeleton thus demonstrating its potential role in mediating cytoskeletal rearrangement necessary in innate immunity but also its ability to link a key catalytic hub, TBK1, to the cytoskeleton to direct TBK1 activity.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Kevin Knitter  

LAB HEAD: Kristina T Nelson

PROVIDER: PXD007262 | Pride | 2018-05-29

REPOSITORIES: Pride