Molecular Characterization and Expression Profiling of Brachypodium distachyon L. Cystatin Genes Reveal High Evolutionary Conservation and Functional Divergence in Response to Abiotic Stress.

Saminathan Subburaj,Dong Zhu,Yueming Yan,Xiaohui Li,Yingkao Hu

doi:10.3389/fpls.2017.00743

Abstract

Cystatin is a class of proteins mainly involved in cysteine protease inhibition and plant growth and development, as well as tolerance under various abiotic stresses. In this study, we performed the first comprehensive analysis of the molecular characterization and expression profiling in response to various abiotic stresses of the cystatin gene family in Brachypodium distachyon, a novel model plant for Triticum species with huge genomes. Comprehensive searches of the Brachypodium genome database identified 25 B. distachyon cystatin (BdC) genes that are distributed unevenly on chromosomes; of these, nine and two were involved in tandem and segmental duplication events, respectively. All BdC genes had similar exon/intron structural organization, with three conserved motifs similar to those from other plant species, indicating their high evolutionary conservation. Expression profiling of 10 typical BdC genes revealed ubiquitous expression in different organs at varying expression levels. BdC gene expression in seedling leaves was particularly highly induced by various abiotic stresses, including the plant hormone abscisic acid and various environmental cues (cold, H2O2, CdCl2, salt, and drought). Interestingly, most BdC genes were significantly upregulated under multiple abiotic stresses, including BdC15 under all stresses, BdC7-2 and BdC10 under five stresses, and BdC7-1, BdC2-1, BdC14, and BdC12 under four stresses. The putative metabolic pathways of cytastin genes in response to various abiotic stresses mainly involve the aberrant protein degradation pathway and reactive oxygen species (ROS)-triggered programmed cell death signaling pathways. These observations provide a better understanding of the structural and functional characteristics of the plant cystatin gene family.

Highlights

Cystatins, which constitute a multigene family, form a class of proteins that inhibits cysteins proteases (Turk and Bode, 1991)
To obtain B. distachyon cystatin (BdC) genes, previously characterized cystatin sequences from wheat, rice, barley, and maize were used as queries to search the public Brachypodium genome database in Phytozome v9.0
A total of 25 non-redundant BdC genes and their protein encoding sequences were identified (Table S3) and serially named BdC1–BdC19 based on their location and chromosomal order (Table 1)

Summary

Introduction

Cystatins, which constitute a multigene family, form a class of proteins that inhibits cysteins proteases (Turk and Bode, 1991). Most cystatins inhibit the activities of cathepsin L-like proteases, a cysteine protease in the peptidase C1A family (Martinez et al, 2009). Plant cystatins, referred to as phytocystatins (phy-cys), are small in size, about 12–16 kDa, and have the LARFAV consensus sequence motif in the region corresponding to a predicted N-terminal αhelix (Misaka et al, 1996). It has been suggested that phy-cys with short N-terminal and longer C-terminal extensions inhibit the activities of cysteine proteases in the peptidase C13 family (Martinez et al, 2007). There are three important signature motifs necessary for the protease inhibition reactions present in all cystatins: a QxVxG reactive site, one or two glycine (G) residues in the N-terminal part of the protein, and a tryptophan residue (W) located downstream of the reactive site (Margis et al, 1998)

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Discussion

Conclusion