Interaction modulation through arrays of clustered methyl-arginine protein modifications.

Jonathan Woodsmith,Nouhad Benlasfer,Christian L Heine,Victoria Casado-Medrano,Dorothee Dormann,Oliver Rocks,Ulrich Stelzl,Rebecca L Eccles,Claudia Abou-Ajram,Verena Thormann,Saskia Hutten

doi:10.26508/lsa.201800178

Abstract

Systematic analysis of human arginine methylation identifies two distinct signaling modes; either isolated modifications akin to canonical post-translational modification regulation, or clustered arrays within disordered protein sequence. Hundreds of proteins contain these methyl-arginine arrays and are more prone to accumulate mutations and more tightly expression-regulated than dispersed methylation targets. Arginines within an array in the highly methylated RNA-binding protein synaptotagmin binding cytoplasmic RNA interacting protein (SYNCRIP) were experimentally shown to function in concert, providing a tunable protein interaction interface. Quantitative immunoprecipitation assays defined two distinct cumulative binding mechanisms operating across 18 proximal arginine-glycine (RG) motifs in SYNCRIP. Functional binding to the methyltransferase PRMT1 was promoted by continual arginine stretches, whereas interaction with the methyl-binding protein SMN1 was arginine content-dependent irrespective of linear position within the unstructured region. This study highlights how highly repetitive modifiable amino acid arrays in low structural complexity regions can provide regulatory platforms, with SYNCRIP as an extreme example how arginine methylation leverages these disordered sequences to mediate cellular interactions.

Highlights

Protein post-translational modifications (PTMs) are known to regulate a vast array of cellular processes governing all facets of human biology
We and others have previously shown that PTMs can cluster across linear protein sequences (Beltrao et al, 2012; Woodsmith et al, 2013), a finding that has been extended to 3D protein structures (Dewhurst et al, 2015)
Protein structures provide a more detailed viewpoint from which to study PTM distributions, they are inherently biased against unstructured regions and limited in number, and as such would impose a large constraint on the PTM dataset

Summary

Introduction

Protein post-translational modifications (PTMs) are known to regulate a vast array of cellular processes governing all facets of human biology. To achieve a comprehensive overview of the entire disordered region, we took a genetic approach to define the binding preferences of a stretch of 19 arginines in the C-terminal SYNCRIP tail using a panel of 37 fulllength mutants in quantitative immunoprecipitation experiments. To define both the unmodified and modified states of the arginine array, we leveraged the methyltransferase PRMT1 and the methylbinding protein SMN1 as functional readouts for arginine and methyl-arginine, respectively. This study reveals how extensive RG repeats within low structural complexity regions can generate cumulative binding mechanisms and, how extensive PTMs allow for a second, distinct recognition mode in a single repeat region

Results

Discussion

Experimental procedures