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bioRxiv · 10.1101/2022.03.29.486189

Regulation of EBNA1 Protein Stability by PLOD1 Lysine Hydroxylase

Dheekollu, J.·Wiedmer, A.·Soldan, S. S.·Castro Munoz, L. J.·Tang, H.-Y.·Speicher, D. W.·Lieberman, P. M.
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Abstract

Epstein-Barr virus (EBV) is a ubiquitous human {gamma}-herpesvirus that is causally associated with various malignancies and autoimmune disease. Epstein-Barr Nuclear Antigen 1 (EBNA1) is the viral-encoded DNA binding protein required for viral episome maintenance and DNA replication during latent infection in proliferating cells. EBNA1 is known to be a highly stable protein, but its mechanism of protein stability is not completely understood. Proteomic analysis of EBNA1 revealed interaction with Procollagen Lysine-2 Oxoglutarate 5 Dioxygenase (PLOD) family of proteins. Depletion of PLOD1 by shRNA or inhibition with small molecule inhibitors 2,-2 dipyridyl resulted in the loss of EBNA1 protein levels, along with a selective growth inhibition of EBV-positive lymphoid cells. PLOD1 depletion also caused a loss of EBV episomes from latently infected cells and inhibited oriP-dependent DNA replication. We used mass spectrometry to identify EBNA1 peptides with lysine hydroxylation at K460 or K461. Mutation of K460 to alanine or arginine abrogates EBNA1-driven DNA replication of oriP, while K461 mutations enhanced replication. These findings suggest that PLOD1 is a novel post-translational regulator of EBNA1 protein stability and function in viral plasmid replication, episome maintenance and host cell survival. ImportanceEBNA1 is essential for EBV latent infection and implicated in viral pathogenesis. We found that EBNA1 interacts with PLOD family of lysine hydroxylases and that this interaction is required for EBNA1 protein stability and function in viral persistence during viral latent infection. Identification of PLOD1 regulation of EBNA1 protein stability provide new opportunity to target EBNA1 for degradation in EBV associated disease.

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Dheekollu, J., Wiedmer, A., Soldan, S. S., Castro Munoz, L. J., Tang, H.-Y., Speicher, D. W., Lieberman, P. M.. 2022-03-29. Regulation of EBNA1 Protein Stability by PLOD1 Lysine Hydroxylase. https://doi.org/10.1101/2022.03.29.486189

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