Altered expression of the caffeine synthase gene in a naturally caffeine-free mutant of Coffea arabica

Mirian Perez Maluf,Michelle De Paula Abreu De Oliveira,Carla Cristina Da Silva,Maria Bernadete Silvarolla,Oliveiro Guerreiro Filho,Aline Gomes Tavares

doi:10.1590/s1415-47572009005000090

Abstract

In this work, we studied the biosynthesis of caffeine by examining the expression of genes involved in this biosynthetic pathway in coffee fruits containing normal or low levels of this substance. The amplification of gene-specific transcripts during fruit development revealed that low-caffeine fruits had a lower expression of the theobromine synthase and caffeine synthase genes and also contained an extra transcript of the caffeine synthase gene. This extra transcript contained only part of exon 1 and all of exon 3. The sequence of the mutant caffeine synthase gene revealed the substitution of isoleucine for valine in the enzyme active site that probably interfered with enzymatic activity. These findings indicate that the absence of caffeine in these mutants probably resulted from a combination of transcriptional regulation and the presence of mutations in the caffeine synthase amino acid sequence.

Highlights

An autogamous, allotetraploid (2n = 4x = 44 chromosomes) species that probably resulted from the natural hybridization of two diploid species, C. canephora and C. eugenioides (Lashermes et al, 1999), is the most important species cultivated for coffee production, primarily because of the excellent beverage quality provided by its beans
The sequences for all methyltransferase genes involved in the biosynthesis of caffeine have previously been identified in coffee and other plant species
All of the primers used here were selected from a conserved consensus region of each gene, Caffeine is one of the best studied compounds in coffee beans, and the characterization of its metabolic pathway is of great importance for future breeding strategies

Summary

Introduction

Several parameters are associated with the high cup quality of C. arabica, including genetic background, environmental conditions, cultivation methods and post-harvest processing. The influence of genetic background on final cup quality is related mainly to the biochemical composition of the fruits and beans. Several studies have shown that coffee species and cultivars vary in their sugar, phenolic compound, lipid and total soluble solid content (Mazzafera et al, 1998; Bradbury, 2001; Aguiar et al, 2005). The caffeine content of coffee beans (expressed as the percentage dry weight) varies among Coffea species: C. arabica beans contain ~1.2% caffeine, C. canephora 2.4% and C. eugenioides 0.3-0.8% (Mazzafera et al, 1997), whereas the beans of C. pseudozangebariae and C. richardii do not contain caffeine (Campa et al, 2005)

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Conclusion