Cooperative stabilisation of 14-3-3σ protein-protein interactions via covalent protein modification.

Marta Falcicchio,Richard G Doveston,Jake A Ward,Pietro Roversi,Alisha Mohindra,Salvador Macip,Sara Y Chothia,Jaswir Basran

doi:10.1039/d1sc02120f

Abstract

14-3-3 proteins are an important family of hub proteins that play important roles in many cellular processes via a large network of interactions with partner proteins. Many of these protein–protein interactions (PPI) are implicated in human diseases such as cancer and neurodegeneration. The stabilisation of selected 14-3-3 PPIs using drug-like ‘molecular glues’ is a novel therapeutic strategy with high potential. However, the examples reported to date have a number of drawbacks in terms of selectivity and potency. Here, we report that WR-1065, the active species of the approved drug amifostine, covalently modifies 14-3-3σ at an isoform-unique cysteine residue, Cys38. This modification leads to isoform-specific stabilisation of two 14-3-3σ PPIs in a manner that is cooperative with a well characterised molecular glue, fusicoccin A. Our findings reveal a novel stabilisation mechanism for 14-3-3σ, an isoform with particular involvement in cancer pathways. This mechanism can be exploited to harness the enhanced potency conveyed by covalent drug molecules and dual ligand cooperativity. This is demonstrated in two cancer cell lines whereby the cooperative behaviour of fusicoccin A and WR-1065 leads to enhanced efficacy for inducing cell death and attenuating cell growth.

Highlights

The 14-3-3 family of hub proteins play diverse and important roles in maintaining normal cell function through interaction with over 200 partner proteins.[1,2] These protein–protein interactions (PPIs) are typically dependent on the phosphorylation of speci c recognition motifs within disordered domains of the partner protein
This modification leads to isoformspecific stabilisation of two 14-3-3s PPIs in a manner that is cooperative with a well characterised molecular glue, fusicoccin A
Our findings reveal a novel stabilisation mechanism for 14-3-3s, an isoform with particular involvement in cancer pathways. This mechanism can be exploited to harness the enhanced potency conveyed by covalent drug molecules and dual ligand cooperativity. This is demonstrated in two cancer cell lines whereby the cooperative behaviour of fusicoccin A and WR-1065 leads to enhanced efficacy for inducing cell death and attenuating cell growth

Summary

Introduction

The 14-3-3 family of hub proteins (seven isoforms: b, g, 3, h, s, s, z) play diverse and important roles in maintaining normal cell function through interaction with over 200 partner proteins.[1,2] These protein–protein interactions (PPIs) are typically dependent on the phosphorylation of speci c recognition motifs within disordered domains of the partner protein. We report that 3 stabilises two distinct 14-3-3s PPIs (with p53 and ERa) via covalent protein modi cation of 14-3-3s at Cys[38], a solvent exposed residue unique to this human 14-3-3 isoform (Fig. 1B).

Results

Conclusion