Discovering the RNA-Binding Proteome of Plant Leaves with an Improved RNA Interactome Capture Method.

Marcel Bach-Pages,Felix Homma,Gail M Preston,Renier A L Van Der Hoorn,Shabaz Mohammed,Alfredo Castello,Jiorgos Kourelis,Markus Kaiser,Farnusch Kaschani

doi:10.3390/biom10040661

Marcel Bach-Pages, Felix Homma + Show 7 more

Open Access

https://doi.org/10.3390/biom10040661

Copy DOI

Journal: Biomolecules	Publication Date: Apr 24, 2020
Citations: 70	License type: CC BY 4.0

Affiliation: University of Oxford, University of Duisburg-Essen

Abstract

RNA-binding proteins (RBPs) play a crucial role in regulating RNA function and fate. However, the full complement of RBPs has only recently begun to be uncovered through proteome-wide approaches such as RNA interactome capture (RIC). RIC has been applied to various cell lines and organisms, including plants, greatly expanding the repertoire of RBPs. However, several technical challenges have limited the efficacy of RIC when applied to plant tissues. Here, we report an improved version of RIC that overcomes the difficulties imposed by leaf tissue. Using this improved RIC method in Arabidopsis leaves, we identified 717 RBPs, generating a deep RNA-binding proteome for leaf tissues. While 75% of these RBPs can be linked to RNA biology, the remaining 25% were previously not known to interact with RNA. Interestingly, we observed that a large number of proteins related to photosynthesis associate with RNA in vivo, including proteins from the four major photosynthetic supercomplexes. As has previously been reported for mammals, a large proportion of leaf RBPs lack known RNA-binding domains, suggesting unconventional modes of RNA binding. We anticipate that this improved RIC method will provide critical insights into RNA metabolism in plants, including how cellular RBPs respond to environmental, physiological and pathological cues.

Highlights

RNA-binding proteins (RBPs) interact with RNAs to form dynamic ribonucleoprotein (RNP) complexes that regulate the fate and function of RNA at virtually every step of its life cycle [1]
We investigated whether the high-confidence leaf RNA-binding proteome (RBPome) had known links to RNA biology using gene ontology (GO) annotations
We discovered that RBPs exclusively identified by plant RNA interactome capture’ (ptRIC) displayed similar domain composition to those identified by multiple plant RBPomes (Figure 5) [34,35,37]

Summary

Introduction

RNA-binding proteins (RBPs) interact with RNAs to form dynamic ribonucleoprotein (RNP) complexes that regulate the fate and function of RNA at virtually every step of its life cycle [1]. In vitro studies have employed protein arrays to determine proteins that capture fluorescently-labelled RNAs [6,7], or immobilised RNAs as baits to isolate RBPs [8] These studies identified hundreds of proteins with the capacity to interact with RNA in vitro; whether these proteins interact with RNA in a physiological, native environment remained unknown [9]. Many in silico algorithms have been developed to identify novel RBPs by searching for domains with homology to well-established RNA-binding domains (RBDs) This has helped to classify hundreds of proteins as putative RBPs in different organisms [10]. These computational approaches are based on homology and cannot discover RBPs with unconventional architectures or sequences, or intrinsically disordered regions (IDRs) with in vivo RNA-binding capacity [11,12]

Methods

Results

Conclusion