Functional dissection of the TBK1 molecular network

Area of Science:

• Immunology
• Molecular Biology
• Cell Signaling

Background:

• TANK-binding kinase 1 (TBK1) and inducible IκB-kinase (IKK-i) are key regulators of type-I interferon induction.
• The functional relationship between TBK1/IKK-i and their associated adaptor proteins (TANK, Sintbad, NAP1) is not well understood.

Purpose of the Study:

• To comprehensively map the molecular network of TBK1/IKK-i.
• To elucidate the distinct roles of adaptor proteins in kinase function and cellular responses.

Main Methods:

• Systematic affinity purification-mass spectrometry to identify protein interactions.
• Immunofluorescence microscopy to determine subcellular localization.
• Reconstitution of kinase-deficient cells with point mutants to assess functional relevance.

Main Results:

• A comprehensive TBK1/IKK-i molecular network revealed mutually exclusive interactions between kinases and adaptors.
• Adaptor proteins TANK, Sintbad, and NAP1 bind to the C-terminal coiled-coil 2 region of TBK1.
• TBK1 activation in response to viral stimuli is critically dependent on the TBK1/TANK interaction.

Conclusions:

• The adaptor proteins form distinct alternative complexes with TBK1/IKK-i.
• The TBK1/TANK interaction is essential for TBK1 activation and downstream signaling in response to viral infection.