Isolation of cyanobacterial mutants exhibiting growth defects under microoxic conditions by transposon tagging mutagenesis of Synechocystis sp. PCC 6803.
Cyanobacteria are photosynthetic prokaryotes that perform oxygenic photosynthesis by extracting electrons from water, with the generation of oxygen as a byproduct. Cyanobacteria use oxygen not only for respiration to produce energy in the dark but also for biosynthesis of various metabolites, such as heme and chlorophyll. Oxygen levels dynamically fluctuate in the field environments, from hyperoxic at daytime to almost anaerobic at night. Thus, adaptation to anaerobiosis should be important for cyanobacteria to survive in low-oxygen and anaerobic environments. However, little is known about the molecular mechanisms of cyanobacterial anaerobiosis because cyanobacteria have been regarded as aerobic organisms. As a first step to elucidate cyanobacterial adaptation mechanisms to low-oxygen environments, we isolated five mutants, T-1-T-5, exhibiting growth defects under microoxic conditions. The mutants were obtained from a transposon-tagged mutant library of the cyanobacterium Synechocystis sp. PCC 6803, which was produced by in vitro transposon tagging of cyanobacterial genomic DNA. Southern blot analysis indicated that a kanamycin resistance gene was inserted in the genome as a single copy. We identified the chromosomal transposon-tagged locus in T-5. Two open reading frames (sll0577 and sll0578) were partially deleted by the insertion of the kanamycin resistance gene in T-5. A reverse transcription polymerase chain reaction suggested that these co-transcribed genes are constitutively expressed under both aerobic and microoxic conditions. Then, we isolated two mutants in which one of the two genes was individually disrupted. Only the mutants partially lacking an intact sll0578 gene showed growth defects under microoxic conditions, whereas it grew normally under aerobic conditions. sll0578 is annotated as purK encoding N5-carboxy-aminoimidazole ribonucleotide synthetase involved in purine metabolism. This result implies the unexpected physiological importance of PurK under low-oxygen environments.
College of Life Sciences, Ritsumeikan University.
Kazuki
K
Terauchi
School of Agricultural Sciences, Nagoya University.
Riho
R
Sobue
College of Life Sciences, Ritsumeikan University.
Yuho
Y
Furutani
Graduate School of Bioagricultural Sciences, Nagoya University.
Rina
R
Aoki
School of Agricultural Sciences, Nagoya University.|Graduate School of Bioagricultural Sciences, Nagoya University.
Yuichi
Y
Fujita
D002734:Chlorophyll / Q000378:metabolism; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D016296:Mutagenesis; D009154:Mutation; D016366:Open Reading Frames; D010100:Oxygen / Q000378:metabolism*; D046939:Synechocystis / Q000235:genetics* / Q000254:growth & development*
D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D002734:Chlorophyll; D010100:Oxygen
Synechocystis sp. PCC 6803; anaerobiosis; cyanobacteria; purK; transposon tagging