Isolation and molecular characterization of rbcS in the unicellular green alga Nannochloris bacillaris (Chlorophyta, Trebouxiophyceae)

Abstract

SUMMARY The small subunit of the chloroplast enzyme ribulose-1,5-bisphosphate carboxyase/oxygenase (Rubisco) is encoded by rbcS. We isolated and characterized three rbcS genes (NbrbcS1-1, NbrbcS1-2, and NbrbcS2) from the unicellular green alga Nannochloris bacillaris (Chlorophyta, Trebouxiophyceae). In the haploid N. bacillaris genome, each is a single-copy gene located on different chromosomes. Each mature peptide contains 140 amino acid residues. NbrbcS1-1 and NbrbcS1-2 are identical and share 80% identity with NbrbcS2, while the respective transit sequences share only approximately 48% identity with that of NbrbcS2. NbrbcS1-1 transcription was suppressed in the dark and recovered drastically after transfer to light. In contrast, NbrbcS1-2 and NbrbcS2 expression were not reduced after transfer from light to dark. In chlorophyllic tobacco cells containing green fluorescent protein fusion proteins of the transit sequences of each peptide, green fluorescent protein signals were localized on particles matching chloroplasts. The first introns of NbrbcS1-1 and NbrbcS1-2 are identical to the corresponding introns of 37 rbcS genes in eight embryophyte species. While the second intron is conserved in the green algae, the NbrbcS2 intron appears to involve sliding by one base pair. The NbrbcS1-1 and NbrbcS1-2 intron conserved in green algae and embryophytes might be an ancestral intron from nuclear-encoded rbcS.