Differential Regulation of Genes Coding for Organelle and Cytosolic ClpATPases under Biotic and Abiotic Stresses in Wheat.

Senthilkumar K Muthusamy,Kailash C Bansal,Monika Dalal,Viswanathan Chinnusamy

doi:10.3389/fpls.2016.00929

Abstract

A sub-group of class I Caseinolytic proteases (Clps) function as molecular chaperone and confer thermotolerance to plants. We identified class I Clp family consisting of five ClpB/HSP100, two ClpC, and two ClpD genes from bread wheat. Phylogenetic analysis showed that these genes were highly conserved across grass genomes. Subcellular localization prediction revealed that TaClpC and TaClpD subgroup proteins and TaClpB1 proteins are potentially targeted to chloroplast, while TaClpB5 to mitochondria, and TaClpB2, TaClpB3, and TaClpB4 to cytoplasm. Spatio-temporal expression pattern analysis revealed that four TaClpB and TaClpD2 genes are expressed in majority of all tissues and developmental stages of wheat. Real-time RT-PCR analysis of expression levels of Clp genes in seven wheat genotypes under different abiotic stresses revealed that genes coding for the cytosolic Clps namely TaClpB2 and TaClpB3 were upregulated under heat, salt and oxidative stress but were downregulated by cold stress in most genotypes. In contrast, genes coding for the chloroplastic Clps TaClpC1, TaClpC2, and TaClpD1 genes were significantly upregulated by mainly by cold stress in most genotypes, while TaClpD2 gene was upregulated >2 fold by salt stress in DBW16. The TaClpB5 gene coding for mitochondrial Clp was upregulated in all genotypes under heat, salt and oxidative stresses. In addition, we found that biotic stresses also upregulated TaClpB4 and TaClpD1. Among biotic stresses, Tilletia caries induced TaClpB2, TaClpB3, TaClpC1, and TaClpD1. Differential expression pattern under different abiotic and biotic stresses and predicted differential cellular localization of Clps suggest their non-redundant organelle and stress-specific roles. Our results also suggest the potential role of Clps in cold, salt and biotic stress responses in addition to the previously established role in thermotolerance of wheat.

Highlights

Caseinolytic protease (Clp) was first identified in Escherichia coli as a heat shock inducible ATP-dependent protease complex that is capable of hydrolyzing casein (Hwang et al, 1987)
TaClpB2, TaClpB3, and TaClpB4 localized to cytoplasm, TaClpB5 localized to mitochondria, while TaClpB1, TaClpC1, TaClpC2, TaClpD1, and TaClpD2 were localized to chloroplast (Table 1)
Presence of a plastid transit peptide in the N-terminal of TaClpB1, TaClpC1, TaClpC2, TaClpD1, and TaClpD2 proteins, and a mitochondrial targeting sequence in the N-terminal of TaClpB5 conformed to the subcellular localization prediction

Summary

Introduction

Caseinolytic protease (Clp) was first identified in Escherichia coli as a heat shock inducible ATP-dependent protease complex that is capable of hydrolyzing casein (Hwang et al, 1987). Subsequent studies showed that ClpATPases fall within the AAA+ superfamily and it functions in assembly and disassembly of protein complexes (Neuwald et al, 1999; Frickey and Lupas, 2004) Members of this family form large hexameric ring structures in ATP dependent manner. These proteins have two AAA+ like domains, which consist of a RecA-like nucleotide-binding domain (NBD) and an α-helical domain (Lee et al, 2004). Under stress conditions, these proteases function as molecular chaperone by minimizing the protein aggregation and target the damaged proteins for degradation (Snider et al, 2008). Class I Clp proteins play an important role in stress responses, growth and development of plants

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