Expression of the CcDREB1D promoter in Coffea arabica: functional genomics and transcriptome analysis

Abstract

Climate change is posing a major challenge to coffee worldwide production and one of the strategies to overcome these problems consists in the generation of crops with increased tolerance to drought and other abiotic stresses, through biotechnological approaches and advanced molecular breeding techniques. Throughout its evolution, plants have developed a series of mechanisms of tolerance to abiotic stresses. However, the activation of these mechanisms varies depending on the plant species, which determines your level of tolerance to stress. The elucidation of the function of the genes of tolerance to stress and also of the process of vegetal defense in these conditions is of fundamental importance for the obtaining of more resistant agricul tural varieties, which consequently can contribute to the reduction of losses in the crops in adverse climatic conditions. Recent studies resulted in the identification of many candidate genes in Coffea for drought tolerance presenting contrasted differential expression profiles between genotypes. Among those are found the genes involved in the water stress response pathway, such as the DREB transcription factors. DREBs (Dehydration Responsive Element Binding) are important plant transcription factors that regulate the expression of many stress-inducible genes and play a critical role in improving the abiotic stress tolerance of plants by interacting with a DRE/CRT cis-element present in the promoter region of various abiotic stress-responsive genes. Among others DREBs, the CcDREB1D gene displayed an increase in mRNA levels during drought stress acclimation. This indicates a distinct regulation for homologous gene expression in the two genotypes and suggests that this gene might contribute to the diversity observed between genotypes. Currently, highperformance sequencing strategies of the transcriptome (RNA-Seq) allow to generate a large volume of data, at low cost and in a short period of time. This information allows the realization of studies of transcriptional profiles and gene networks in an in-depth understanding of several gene profiles. Facing this context, the Chapter 1 presents a review of the literature encompassing from the origin and commercial characteristics of coffee, to the study of genes and their transcriptome. The Chapter 2 evaluated the activity of three CcDREB1D promoter haplotypes isolated from drought-tolerant and drought-susceptible clones of Coffea canephora by evaluating their ability to control the expression of the uidA reporter gene in response to water deficit. The Chapter 3 comprised the study of the haplotype pHP16L isolated from C. canephora and its participation in the expression of the uidA reporter gene in response to drought, high and low temperatures, high light intensity and exogenous ABA application stresses, accompanied by RT-qPCR data. The Chapter 4 comprised analyzing the expression conditions of the pHP16L haplotype of the DREB1D promoter under a representative range of the most common abiotic stresses using the RNA-Seq technique as a tool to understand the mechanism of plant stress tolerance and induced stress genes expression as well as DREBs gene, accompanied by data validation by the RT-qPCR technique.