Project description:The closely related Coffea arabica cultivars ‘Tall Mokka’ and ‘Typica’, with excellent flavor, but differing distinctively in the size of aerial organs, branching pattern and branch numbers. Differential gene expression analysis of shoot tips of arabica coffee cultivars 'Tall Mokka' and 'Typica' were done using Potato cDNA microarray as cross-species platform. Using cross-species microarray hybridization, we identified a prolyl oligopeptidase (CaPOP) gene as differentially expressed between the shoot tips of ‘Tall Mokka’ and ‘Typica’. Isolation and sequencing of POP genes from coffee identified three paralogs, CaPOP1, CaPOP2 and CaPOP3. All three genes were present in both cultivars, which suggest that differences in the expression of CaPOP are caused by factor(s) regulating the transcription of CaPOPs. CaPOP1 differs in sequence from CaPOP2 primarily in having two large deletions in the promoter region. CaPOP genes are homologous to arabidopsis At1g20380, encoding a post-proline cleaving enzyme that acts on substrates shorter than 30 amino acids. Ectopic expression of CaPOP1 under its native promoter in transgenic arabidopsis resulted in more secondary branches than in the wild type. This is the first study to successfully isolate CaPOP genes and characterize their expression in the developing tissues of coffee. This study also identified a novel role for prolyl oligopeptidase in control of branching. Eight coffee trees of 'Typica' ('K') and six trees of 'Tall Mokka' ('M') cultivar were used in this study. The trees were equally divided into two groups 'A' and 'B' for each cultivar ('MA','MB', 'KA' and 'KB') and treated as biological replicates. Eight two channel microarray hybridizations were done in following pairs: MA x KA, MA x KB, MB x KA, MB x KB and dye swap replicate of each pair. Summary: Two-sample experiment: Tall Mokka vs. Typica . 8 Hybridizations. 2 Biological replicates per sample. 1 Dye swap per array.
Project description:The closely related Coffea arabica cultivars ‘Tall Mokka’ and ‘Typica’, with excellent flavor, but differing distinctively in the size of aerial organs, branching pattern and branch numbers. Differential gene expression analysis of shoot tips of arabica coffee cultivars 'Tall Mokka' and 'Typica' were done using Potato cDNA microarray as cross-species platform. Using cross-species microarray hybridization, we identified a prolyl oligopeptidase (CaPOP) gene as differentially expressed between the shoot tips of ‘Tall Mokka’ and ‘Typica’. Isolation and sequencing of POP genes from coffee identified three paralogs, CaPOP1, CaPOP2 and CaPOP3. All three genes were present in both cultivars, which suggest that differences in the expression of CaPOP are caused by factor(s) regulating the transcription of CaPOPs. CaPOP1 differs in sequence from CaPOP2 primarily in having two large deletions in the promoter region. CaPOP genes are homologous to arabidopsis At1g20380, encoding a post-proline cleaving enzyme that acts on substrates shorter than 30 amino acids. Ectopic expression of CaPOP1 under its native promoter in transgenic arabidopsis resulted in more secondary branches than in the wild type. This is the first study to successfully isolate CaPOP genes and characterize their expression in the developing tissues of coffee. This study also identified a novel role for prolyl oligopeptidase in control of branching.
Project description:Caffeine is a metabolite of great economic importance, especially in coffee, where it influences the sensorial and physiological impacts of the beverage. Caffeine metabolism in the Coffea species begins with the degradation of purine nucleotides through three specific N-methyltransferases: XMT, MXMT and DXMT. A comparative analysis was performed to clarify the molecular reasons behind differences in caffeine accumulation in two Coffea species, namely Coffea arabica and Coffea canephora var. robusta. Three different genes encoding N-methyltransferase were amplified in the doubled haploid Coffea canephora: CcXMT1, CcMXMT1 and CcDXMT. Six genes were amplified in the haploid Coffea arabica: CaXMT1, CaXMT2, CaMXMT1, CaMXMT2, CaDXMT1, and CaDXMT2. A complete phylogenic analysis was performed to identify specific key amino acids defining enzymatic function for each protein identified. Furthermore, a quantitative gene-expression analysis was conducted on leaves and on maturing coffee beans, simultaneously analyzing caffeine content. In the different varieties analyzed, caffeine accumulation is higher in leaves than in the coffee bean maturation period, higher in Robusta than in Arabica. In Robusta, CcXMT1 and CcDXMT gene expressions are predominant and transcriptional activity is higher in leaves than in maturing beans, and is highly correlated to caffeine accumulation. In Arabica, the CaXMT1 expression level is high in leaves and CaDXMT2 as well to a lesser extent, while global transcriptional activity is weak during bean maturation, suggesting that the transcriptional control of caffeine-related genes differs within different organs and between Arabica and Robusta. These findings indicate that caffeine accumulation in Coffea species has been modulated by a combination of differential transcriptional regulation and genome evolution.
Project description:As microRNAs (miRNAs) are important regulators of many biological processes, a series of small RNAomes from plants have been produced in the last decade. However, miRNA data from several groups of plants are still lacking, including some economically important crops. Here microRNAs from Coffea canephora leaves were profiled and 58 unique sequences belonging to 33 families were found, including two novel microRNAs that have never been described before in plants. Some of the microRNA sequences were also identified in Coffea arabica that, together with C. canephora, correspond to the two major sources of coffee production in the world. The targets of almost all miRNAs were also predicted on coffee expressed sequences. This is the first report of novel miRNAs in the genus Coffea, and also the first in the plant order Gentianales. The data obtained establishes the basis for the understanding of the complex miRNA-target network on those two important crops.