Fish CDK2 recruits Dtx4 to degrade TBK1 through ubiquitination in the antiviral response

Lu, L.-F.; Zhang, C.; Li, Z.-C.; Cui, B.-J.; Wang, Y.-Y.; Han, K.-J.; Xu, X.; Zhou, C.-J.; Zhou, X.-Y.; Wu, Y.; Xu, N.; Yang, X.-L.; Chen, D.-D.; Li, X.; Zhou, L.; Li, S.

doi:10.1101/2024.04.23.590743

bioRxiv · 10.1101/2024.04.23.590743

Fish CDK2 recruits Dtx4 to degrade TBK1 through ubiquitination in the antiviral response

Lu, L.-F.·Zhang, C.·Li, Z.-C.·Cui, B.-J.·Wang, Y.-Y.·Han, K.-J.·Xu, X.·Zhou, C.-J.·Zhou, X.-Y.·Wu, Y.·Xu, N.·Yang, X.-L.·Chen, D.-D.·Li, X.·Zhou, L.·Li, S.

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Abstract

Although the classical biological protein cell cycle protein kinase CDK2 has been extensively studied in higher vertebrates, its function in lower vertebrates beyond the regulation of mitosis remains unknown. In this study, we report a distinct mechanism whereby IFN expression is negatively regulated in fish by CDK2. After infection with the spring viremia of carp virus (SVCV), fish CDK2 expression significantly increased in tissues and cells. Moreover, antiviral resistance was improved in cdk2-/- homozygotes, and the antiviral cytokine interferon (IFN) expression was significantly higher. At the cellular level, CDK2 overexpression reduced IFN expression, while cdk2 knockdown increased the ability of cells to produce IFN. Subsequently, it was discovered that fish CDK2 binds and degrades TBK1, resulting in reduced IFN. CDK2 increases the K48-linked ubiquitination of TBK1, causing its degradation, while E3 ubiquitin ligase Dtx4 was found to be involved in this process following the significant enhancement of TBK1 K48-linked ubiquitination. Protein mass spectrometry and immunoblot analysis confirmed that the K567 site on TBK1 is essential for CDK2 to engage with Dtx4 and degrade TBK1; thus, after mutating the K567 site, K48-linked ubiquitination of TBK1 was not enhanced by Dtx4, and TBK1 was not degraded by CDK2. Our data demonstrate that fish CDK2 recruits the E3 ubiquitin ligase Dtx4 to target the K567 site of TBK1 and promote its degradation. These results suggest that CDK2 in lower vertebrates is implicated in a specialized role for antiviral innate immunity.

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Lu, L.-F., Zhang, C., Li, Z.-C., Cui, B.-J., Wang, Y.-Y., Han, K.-J., Xu, X., Zhou, C.-J., Zhou, X.-Y., Wu, Y., Xu, N., Yang, X.-L., Chen, D.-D., Li, X., Zhou, L., Li, S.. 2024-04-27. Fish CDK2 recruits Dtx4 to degrade TBK1 through ubiquitination in the antiviral response. https://doi.org/10.1101/2024.04.23.590743

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