DEAD box protein is required for formation of a hidden break in Arabidopsis chloroplast 23S rRNA

• Published in: The Plant journal : for cell and molecular biology 2010-09, Vol.63 (5), p.766-777
• Main Authors: Nishimura, Kenji, Ashida, Hiroki, Ogawa, Taro, Yokota, Akiho
• Format: Article
• Language: English
• Subjects: 23S rRNA; Amino Acid Sequence; Arabidopsis; Arabidopsis - genetics; Arabidopsis - metabolism; Arabidopsis Proteins - genetics; Arabidopsis Proteins - metabolism; Arabidopsis thaliana; Blotting, Northern; Blotting, Western; chloroplast; Chloroplasts; Chloroplasts - metabolism; DEAD box protein; DEAD-box RNA Helicases - genetics; DEAD-box RNA Helicases - metabolism; Flowers & plants; Genes, Essential - genetics; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; hidden break; Molecular biology; Molecular modelling; Molecular Sequence Data; Mutation; Plant biology; Protein Binding; Proteins; Recombinant Proteins - metabolism; Ribonucleic acid; ribulose-1,5-bisphosphate; Ribulose-Bisphosphate Carboxylase - genetics; Ribulose-Bisphosphate Carboxylase - metabolism; RNA; RNA processing; RNA, Chloroplast - genetics; RNA, Chloroplast - metabolism; RNA, Ribosomal, 23S - genetics; RNA, Ribosomal, 23S - metabolism; rRNA; rRNA 23S; Sequence Homology, Amino Acid; Transcription; Translation
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1111/j.1365-313X.2010.04276.x

In plant chloroplasts, the ribosomal RNA (rRNA) of the large subunit of the ribosome undergoes post-maturation fragmentation processing. This processing consists of site-specific cleavage that generates gapped, discontinuous rRNA molecules. However, the molecular mechanism underlying introduction of the gap structure (the 'hidden break') is poorly understood. Here, we found that the DEAD box protein RH39 plays a key role in introduction of the hidden break into the 23S rRNA in Arabidopsis chloroplasts. Genetic screening for an Arabidopsis plant with a drastically reduced level of ribulose-1,5-bisphosphate carboxylase/oxygenase identified an RH39 mutant. The levels of other chloroplast-encoded photosynthetic proteins were also severely reduced. The reductions were not due to a failure of transcription, but rather inefficiency in translation. RNA gel blotting revealed incomplete fragmentation of 23S rRNA in chloroplasts during maturation. In vitro analysis with recombinant RH39 suggested that the protein binds to the adjacent sequence upstream of the hidden break site to exert its function. We propose a molecular mechanism for the RH39-mediated fragmentation processing of 23S rRNA in chloroplasts.