Rubisco small subunit of Coffea arabica: cDNA sequence, gene cloning and promoter analysis in transgenic tobacco plants

Abstract

As part of our search for tissue-specific promoters in coffee ( Coffea arabica), we used 2-D gel electrophoresis to detect proteins highly and specifically expressed in coffee leaves. Among the proteins identified by N-terminal sequencing, the small subunit (SSU) of the coffee ribulose 1,5-bisphosphate carboxylase/oxygenase (EC.4.1.1.39) was one of the most abundant. Using the protein sequence information available in databases, degenerate primers were used to screen a coffee leaf cDNA library, leading to the cloning of a full-length cDNA and a genomic clone for the coffee RBCS1 gene. The promoter region of this gene was also isolated. Sequence analysis of this promoter revealed the presence of several putative DNA boxes with similarities to well-characterized light responsive elements (LRE) that have previously been shown to be critical components for transcriptional control of gene expression by light. Using a RBCS1 promoter– uidA translational fusion, we demonstrated that this coffee promoter could function as a leaf-specific and light-regulated promoter in transgenic tobacco plants. These data suggest that the 1-kb coffee RBCS1 promoter sequence contained all the cis-elements required for developmental and light-mediated control of gene expression and that trans-acting factors implicated in this mechanism are probably highly conserved between coffee and tobacco.