Surface charge of Merkel cell polyomavirus small T antigen determines cell transformation through allosteric FBW7 WD40 domain targeting

Nnenna Nwogu,Hyun Jin Kwun,Luz E Ortiz

doi:10.1038/s41389-020-0235-y

Abstract

Merkel cell polyomavirus (MCV) small T (sT) is the main oncoprotein in Merkel cell carcinoma (MCC) development. A unique domain of sT, LT stabilization domain (LSD), has been reported to bind and inactivate multiple SCF (Skp1-Cullin-F-box) E3 ligases. These interactions impede the turnover of MCV large T (LT) antigen and cellular oncoproteins such as c-Myc and cyclin E, thereby promoting viral replication and cell transformation. However, it is currently unclear how this LSD region contributes to multiple transforming activities of sT. Structural docking simulation of sT and F-box and WD repeat domain-containing 7 (FBW7) revealed a novel allosteric interaction between sT and FBW7 WD40 domain. This model is supported by experimental evidence confirming that charge engineering in the LSD alters sT-WD40 binding. Specifically, loss of net positive charge in the LSD prevents sT-FBW7 binding by abrogating the electrostatic interaction, thus impeding inhibition of FBW7 by sT. Furthermore, positively charged mutations in the LSD significantly restored the sT function and its ability to transform rodent fibroblast cells. We infer that the surface charge of sT is a major determinant for targeting E3 ligases, which leads to sT-induced cell transformation, an observation that could be used to develop targeted therapeutics for MCC.

Highlights

E3 ubiquitin ligases are a large family of proteins engaged in the proteolytic degradation of many target proteins, regulating their turnover and activity
The mechanism by which F-box and WD-repeat domain-containing 7 (FBW7) WD40 domain interacts with target proteins is well characterized[5]
We focused our study on the potential interaction between Merkel cell polyomavirus (MCV) small T (sT) large T (LT) stabilization domain (LSD) and FBW7 WD40 domain and the resultant inhibitory effect on FBW7 function

Summary

Introduction

E3 ubiquitin ligases are a large family of proteins engaged in the proteolytic degradation of many target proteins, regulating their turnover and activity. The Skp[1], Cullin, F-box containing (SCF) complex is a key player in cell cycle regulation, and has been shown to be a major moderator of essential proteins involved in cell cycle progression[1]. The F-box protein family is classified based on the specific substrate recognition domains identified within the F-box motif. These classes include: FBWX, which contains tryptophan (W)-aspartic acid (D) dipeptide (WD40) repeats; FBXL, which includes leucine-. The F-box and WD-repeat domain-containing 7 (FBW7) protein plays essential roles in various physiological and pathological cell processes, including facilitating the degradation of various oncoproteins by means of the ubiquitin-proteasome system, thereby regulating cell growth[3].

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