Thiamine biosynthesis in algae is regulated by riboswitches

• Published in: Proceedings of the National Academy of Sciences - PNAS 2007-12, Vol.104 (52), p.20770-20775
• Main Authors: Croft, Martin T, Moulin, Michael, Webb, Michael E, Smith, Alison G
• Format: Article
• Language: English
• Subjects: Alternative Splicing; Animals; Bacteria; Bacteriology; Base Sequence; Biochemistry - methods; Biological Sciences; Biosynthesis; Chlamydomonas reinhardtii; Chlamydomonas reinhardtii - metabolism; Codon; Enzymes; Eukaryota - metabolism; Eukaryota - physiology; Exons; Gene Expression Regulation; Genes; Genes, Reporter; Introns; Luciferases - metabolism; Metabolism; Models, Chemical; Molecular Sequence Data; Mutation; Nucleic Acid Conformation; Photosynthesis; Plasmids - metabolism; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleic acid; RNA; Splicing; Thiamine - chemistry; Thiamine - metabolism; Untranslated regions; Volvox carteri
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0705786105

In bacteria, many genes involved in the biosynthesis of cofactors such as thiamine pyrophosphate (TPP) are regulated by ribo switches, regions in the 5' end of mRNAs to which the cofactor binds, thereby affecting translation and/or transcription. TPP riboswitches have now been identified in fungi, in which they alter mRNA splicing. Here, we show that addition of thiamine to cultures of the model green alga Chlamydomonas reinhardtii alters splicing of transcripts for the THI4 and THIC genes, encoding the first enzymes of the thiazole and pyrimidine branches of thiamine biosynthesis, respectively, concomitant with an increase in intracellular thiamine and TPP levels. Comparison with Volvox carteri, a related alga, revealed highly conserved regions within introns of these genes. Inspection of the sequences identified TPP riboswitch motifs, and RNA transcribed from the regions binds TPP in vitro. The THI4 riboswitch, but not the promoter region, was found to be necessary and sufficient for thiamine to repress expression of a luciferase-encoding reporter construct in vivo. The pyr1 mutant of C. reinhardtii, which is resistant to the thiamine analogue pyrithiamine, has a mutation in the THI4 riboswitch that prevents the THI4 gene from being repressed by TPP. By the use of these ribo switches, thiamine biosynthesis in C. reinhardtii can be effectively regulated at physiological concentrations of the vitamin.