Identification and Characterization of Cold-Tolerance Associated Genes in Wheat

The low temperature remains as one of the major limiting factors of crop productivity in the temperate region, and identification of cold tolerance related genes is crucial for developing cold tolerant crop plants to increase agricultural productivity. The objective of my thesis is to identify cold tolerance related candidate genes in wheat, one of the major crops in the temperate region. In Chapter 2, I have reviewed the literature pertaining to the mechanisms of cold tolerance in plants with specific emphasis on Wheat. In Chapter 3, forty candidate genes with increased expression under cold exposure based on published microarray data were selected and further characterized. These genes belonging to four categories namely defense-related regulators; transcriptional and epigenetic regulators; post-transcriptional and post-translational regulators; and genes of unknown functions revealed many differentially expressed genes including Remorin – upregulated in response to cold; a novel gene in wheat homologous to RD29B of Arabidopsis-upregulated in response to cold and ABA; and another novel gene regulated by both ABA and MetJA. In chapter 4, the results of genome-wide identification and characterization of the wheat remorin family and its association with cold tolerance are presented. A search of the wheat database revealed the existence of twenty different remorin genes that we classified into six groups sharing a common structure and phylogenetic origin. Promoter analysis of TaREM genes revealed the presence of putative cis-elements related to diverse functions like development, hormonal regulation, biotic and abiotic stress responsiveness. Expression levels of TaREM genes were measured in plants grown under in field and laboratory conditions and in response to hormone treatment. Our analyses revealed twelve members of the remorin family that are regulated during cold acclimation of wheat in four different tissues (root, crown, stem and leaves), with the highest expression in roots. Differential gene expression was found between wheat cultivars with contrasting degree of cold tolerance suggesting the implication of TaREM genes in cold response and tolerance. Additionally, eight genes were induced in response to ABA and MetJA treatment. This genome-wide analysis of TaREM genes provides valuable resources for functional analysis aimed at understanding their role in stress adaptation. The chapter 5 is focused on gaining insights into the evolutionary history and in-silico functional characterization of a novel cold-responsive gene in wheat. This gene in wheat has distant homology to known abiotic stress-related genes in other plants including CAP160 in Spinacia oleracea, RD29B in Arabidopsis and CDeT11-24 in Craterostigma plantagineum. The results show that these genes are homologous and may have evolved from a common ancestor. The Bayesian phylogenetic analyses of the protein sequences of this gene from various plant species revealed three distinctive clades. Further analyses revealed that this gene has predominantly evolved through neutral processes with some regions experiencing signatures of negative selections and some regions showing signatures of episodic positive selections. These genes contained common K-like segments and function predictions revealed that these protein-coding genes may share at least two functions related to abiotic stress conditions. One function is similar to the cryoprotective function of LEA protein, and the second function as a signalling molecule by binding specifically to phosphatidic acid.