The Epstein-Barr Virus Oncoprotein, LMP1, Regulates the Function of SENP2, a SUMO-protease

Thomas L Selby,Akash Patel,Tabithia Ross,Matthew Varn,Gretchen L Bentz,C Randall Moss,Sheetal Patel,Natalie Biel,Angela J Lowrey,Wyatt T Cramblet,Ashley Ray,Leslie Hilding

doi:10.1038/s41598-019-45825-5

Thomas L Selby, Akash Patel + Show 10 more

Open Access

https://doi.org/10.1038/s41598-019-45825-5

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Abstract

Epstein-Barr virus (EBV) latent membrane protein-1 (LMP1) activates numerous signal transduction pathways using its C-terminal activating regions. We reported that LMP1 increased global levels of sumoylated proteins, which aided the oncogenic nature of LMP1. Because increased protein sumoylation is detected in numerous cancers, we wanted to elucidate additional mechanisms by which LMP1 modulates the sumoylation machinery. Results indicated that SUMO-protease activity decreased in a LMP1-dependent manner, so we hypothesized that LMP1 inhibits SUMO-protease activity, resulting in reduced de-sumoylation of cellular proteins, which contributes to the detected accumulation of sumoylated proteins in EBV-positive lymphomas. Focusing on SENP2, findings revealed that LMP1 expression corresponded with increased sumoylation of SENP2 at K48 and K447 in a CTAR-dependent manner. Interestingly, independent of LMP1-induced sumoylation of SENP2, LMP1 also decreased SENP2 activity, decreased SENP2 turnover, and altered the localization of SENP2, which led us to investigate if LMP1 regulated the biology of SENP2 by a different post-translational modification, specifically ubiquitination. Data showed that expression of LMP1 inhibited the ubiquitination of SENP2, and inhibition of ubiquitination was sufficient to mimic LMP1-induced changes in SENP2 activity and trafficking. Together, these findings suggest that LMP1 modulates different post-translational modifications of SENP2 in order to modulate its biology and identify a third member of the sumoylation machinery that is manipulated by LMP1 during latent EBV infections, which can affect oncogenesis.

Highlights

Dysregulation of cellular sumoylation processes is a feature of a variety of diseases[7,8,9,10,11,12,13,14,15], and targeting Ubc[9] and the SENP expression and function has been proposed as potential new therapeutic targets[9,16,17]
Because the SUMO-proteases are required for SUMO maturation and protein de-sumoylation[33], we first investigated if latent membrane protein-1 (LMP1) affected SENP activity by quantitating the ability of the endogenous SENPs to cleave a SUMO-modified substrate in LMP1- and control-expressing Human embryonic kidney (HEK) 293 cells
In LMP1-expressing cells, the decreased SENP activity corresponded with a detectable increase of SUMO-1/2/3 levels (Fig. 1b and Supplementary Data), confirming our earlier reports that LMP1 expression coincided with increased sumoylation during Epstein-Barr virus (EBV) latency[15,27,32,40] and suggesting that inhibition of SENP2 activity may augment the accumulation of sumoylated proteins detected in LMP1-positive lymphomas

Summary

Introduction

Dysregulation of cellular sumoylation processes is a feature of a variety of diseases[7,8,9,10,11,12,13,14,15], and targeting Ubc[9] and the SENP expression and function has been proposed as potential new therapeutic targets[9,16,17]. CTAR3 hijacks enzymatically active Ubc[9] during latent EBV infections[15], which results in increased sumoylation of cellular proteins[15], aids LMP1-induced cell migration[15], modulates innate immune responses[32], and aids the maintenance of latency[27]. These findings prompted us to question if LMP1 targeted additional steps of the sumoylation processes and manipulated additional members of the sumoylation machinery. The end result is the decreased de-sumoylation of cellular proteins, which aids the documented global increase in sumoylated proteins during latent infections[40]

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