Expression of Chloroplast Small Heat Shock Proteins of C. album under Different Abiotic Stresses

Abstract

Plant response to the altered environmental conditions usually includes initiations of several defense mechanisms. One of these mechanisms is the heat shock proteins (HSPs) production, a universal response. Chloroplast small heat shock proteins (Cp-sHSPs) are known to protect Photosystem II under stress conditions in plants. Cp-sHSPs production is positively correlated with the plant thermotolerance. While the protective role of HSPs has been greatly recognized against several physiological stresses such as heat, metals, cold and different developmental stages individually or in different combinations. Recent progress in this field has revealed unique aspects of regulations of these genes under stress. Cis-regulatory elements present in promoter regions with the combinations of several factors dictate the gene expression levels and specific patterns. Not much is known about the regulation and mechanism of action of Cp-sHSP genes in plant protection. We analyzed four novel promoters of Chenopodium album (two US ecotypes) Cp-sHSPs for the presence of putative cis-regulatory motifs (Shakeel et al., 2011). Interestingly we found more than one cis-regulatory elements in the promoter regions of these Ca-sHSPs and based on this unique promoter architecture, we proposed a differential regulation of a single Ca-sHSP gene under variety of different abiotic stresses including heat, metal, cold, drought and salt stresses individually. To analyze the Ca-sHSP gene expression patterns under different abiotic stress conditions, we treated C. album (Pakistani ecotype) plants at different abiotic conditions and initially studied the effect of above mentioned stresses on physiology of the plants. On the basis of physiological data, we decided the lower and upper limits of stress conditions for further analyses of Ca-sHSPs. Stress induced transcripts of Ca-sHSP genes were sequenced individually and analyzed; our data showed that one novel Cp-sHSP family member, CaHSP26.13p is regulated differently under different abiotic stresses. This observation was based on 100% amino acid sequence similarities among these transcripts. Expression of Chloroplast Small Heat Shock Proteins of C. album under Different Abiotic Stresses Relative abundance of transcript and protein levels was determined by real-time PCR analysis and immunoblotting of C. album leaf samples treated with and without stress conditions. Antibodies specific to methionine rich region were used for Cp-sHSPs detection by immunoblotting. Interestingly we detected 1-2 bands of precursor and processed Cp-sHSP proteins (~26 and 21KDa) depending upon the stress conditions, while only precursor protein (~26KDa) with differential expression was detected in all types of treatments suggesting the role of single Ca-sHSP in plant protection under heat, metal, cold, drought and salt stress. Due to the absence of correlation between transcript and protein levels in most of these cases, we speculate some role of post-transcriptional regulation in plant protection by this mechanism. This study demonstrates multiple roles of single C. album Cp-sHSP in variety of environmental conditions by differential regulation and can be used to develop stress tolerant plant species to face the challenge of next decade. The data presented here has been published in the following articles. 1. Samina Shakeel, Noor Ul Haq, Scott A. Heckathorn, E. William Hamilton, Dawn S. Luthe (2011). Ecotypic variation in chloroplast small heat-shock proteins and related thermotolerance in Chenopodium album. Plant Physiology and Biochemistry 49: 898-908. 2. Noor Ul Haq, Sana Raza, Dawn S. Luthe, Scott A. Heckathorn, Samina N Shakeel (2012). Dual role for Chenopodium album chloroplast small heat shock protein: Photosystem II protection from heat and metal stresses. Plant Molecular Biology Reporter, DOI 10.1007/s11105-012-0516-5. 3. Noor Ul Haq, Muhammad Ammar, Dawn S. Luthe, Scott A. Heckathorn, Samina N Shakeel. Molecular characterization of Chenopodium album chloroplast small heat shock protein and its expressions in response to different abiotic stresses. (Submitted to Plant Cell Reports).