HTLV-1 Tax oncoprotein subverts the cellular DNA damage response via binding to DNA-dependent protein kinase

Area of Science:

• Virology
• Molecular Biology
• Cancer Research

Background:

• Human T-cell leukemia virus type-1 (HTLV-1) causes adult T-cell leukemia.
• The viral oncoprotein Tax impairs cell cycle control and DNA damage response.
• Tax sequesters huChk2 in chromatin, hindering response to ionizing radiation.

Purpose of the Study:

• To investigate the role of DNA-dependent protein kinase (DNA-PK) in the Tax-mediated disruption of DNA damage response.
• To elucidate the interaction between Tax, huChk2, and DNA-PK.

Main Methods:

• Nuclear complex analysis to identify DNA-PK components (DNA-PKcs, Ku70) in the Tax-Chk2 complex.
• Assays measuring DNA-PK activity in Tax-containing extracts.
• Inhibition of DNA-PK to assess its role in Tax-induced Chk2 activation.
• Immunofluorescence to detect foci formation (H2AX phosphorylation) and Tax/phospho-DNA-PK co-localization.
• Analysis of a nuclear-excluded Tax mutant.

Main Results:

• DNA-dependent protein kinase (DNA-PK), including DNA-PKcs and Ku70, is part of the Tax-Chk2 nuclear complex.
• Tax increases DNA-PK activity, and its inhibition prevents Tax-induced Chk2 activation.
• Tax expression induces H2AX phosphorylation but impairs the response to ionizing radiation by inhibiting DNA-PK phosphorylation and gammaH2AX stabilization.
• Tax co-localizes with phospho-DNA-PK, and a nuclear-excluded Tax mutant sequesters phospho-DNA-PK in the cytoplasm, indicating Tax-DNA-PK interaction is crucial.
• A novel Tax-dependent interaction between DNA-PK and Chk2 was identified.

Conclusions:

• Tax binds to and stabilizes a protein complex involving DNA-PK and Chk2.
• This Tax-DNA-PK-Chk2 complex saturates the DNA damage repair response pathway.
• Impaired DNA damage repair due to Tax interference contributes to HTLV-1-associated leukemogenesis.