Structural and Biochemical Analysis of the Hordeum vulgare L. HvGR-RBP1 Protein, a Glycine-Rich RNA-Binding Protein Involved in the Regulation of Barley Plant Development and Stress Response

Abstract

The timing of whole-plant senescence influences important agricultural traits such as yield and grain protein content. Post-transcriptional regulation by plant RNA-binding proteins is essential for proper control of gene expression, development, and stress responses. Here, we report the three-dimensional solution NMR structure and nucleic acid-binding properties of the barley glycine-rich RNA-binding protein HvGR-RBP1, whose transcript has been identified as being >45-fold up-regulated in early—as compared to late—senescing near-isogenic barley germplasm. NMR analysis reveals that HvGR-RBP1 is a multidomain protein comprising a well-folded N-terminal RNA Recognition Motif (RRM) and a structurally disordered C-terminal glycine-rich domain. Chemical shift differences observed in 2D 1H–15N correlation (HSQC) NMR spectra of full-length HvGR-RBP1 and N-HvGR-RBP1 (RRM domain only) suggest that the two domains can interact both in-trans and intramolecularly, similar to what is observed in the tobacco NtGR-RBP1 protein. Further, we show that the RRM domain of HvGR-RBP1 binds single-stranded DNA nucleotide fragments containing the consensus nucleotide sequence 5′-TTCTGX-3′ with low micromolar affinity in vitro. We also demonstrate that the C-terminal glycine-rich (HvGR) domain of Hv-GR-RBP1 can interact nonspecifically with ssRNA in vitro. Structural similarities with other plant glycine-rich RNA-binding proteins suggest that HvGR-RBP1 may be multifunctional. Based on gene expression analysis following cold stress in barley and E. coli growth studies following cold shock treatment, we conclude that HvGR-RBP1 functions in a manner similar to cold-shock proteins and harbors RNA chaperone activity. HvGR-RBP1 is therefore not only involved in the regulation of barley development including senescence, but also functions in plant responses to environmental stress.

Quantitative Proteomic Analysis of Alligator Weed Leaves Reveals That Cationic Peroxidase 1 Plays Vital Roles in the Potassium Deficiency Stress Response

“…In our study, one annexin was up-regulated, and the other was down-regulated, suggesting that this family of proteins has various functions. Glycine-rich RNA-binding protein (GRP) functions in a manner similar to cold-shock proteins and has RNA chaperone activity in barley [36]. Further research suggested that this protein is a key molecular component of hormone-regulated development and regulates gene expression at multiple levels, such as by modifying mRNA alternative splicing as well as via export, translation, and degradation [37].…”

The Glycine-Rich RNA-Binding Protein Is a Vital Post-Transcriptional Regulator in Crops

“…In E. coli cells expressing HvGR-RBP1 heterologously, these cells exhibited faster growth compared to control cells following exposure to cold shock. It is worth noting that the RRM structural domain of HvGR-RBP1 is crucial for this function [89]. And, also, the mRNA level of HvGRP3 was significantly increased under cold treatment and fungal pathogen Erysiphe graminis and Rhynchosporium secalis infestation [90].…”

Plant RNA Binding Proteins as Critical Modulators in Drought, High Salinity, Heat, and Cold Stress Responses: An Updated Overview

“…At least eight of those GaRB-GRP and GrRB-GRP genes showed differential expression under salt stress conditions indicating they might be involved in salt stress responses of cotton species. The HvGR-RBP1 of barley was upregulated in response to cold stress [31]. A recent study investigating the functions of OsGRP3 in rice using overexpression and knockout mutants revealed that OsGRP3 expression contributes to drought tolerance by alleviating the ROS accumulation through the regulation of ROS genes.…”