Initiation codon mutations in the Chlamydomonas chloroplast petD gene result in temperature-sensitive photosynthetic growth.
The chloroplast petD gene encodes subunit IV of the cytochrome b6/f complex and is required for photosynthetic electron transport. We have created Chlamydomonas strains in which the initiation codon of the petD gene has been changed to AUU or AUC. These mutants can grow photosynthetically at room temperature, but not at 35 degrees C. The accumulation of subunit IV during photosynthetic or heterotrophic growth at room temperature is reduced to 10-20% of the wild-type level; petD mRNA abundance is reduced to approximately 50% of the wild-type amount. Pulse labeling experiments indicate that at room temperature, subunit IV translation proceeds at 10-20% of the wild-type rate. Cells grown heterotrophically at 35 degrees C accumulate < 5% as much subunit IV as wild-type cells grown under the same conditions, and < 1% as much subunit IV as wild-type cells grown at room temperature. We conclude that translation initiation in these mutants is inefficient, leading to decreased translation and accumulation of subunit IV. At 35 degrees C, translational inefficiency leads directly or indirectly to insufficient accumulation of subunit IV to support photosynthetic growth.
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Boyce Thompson Institute for Plant Research, Cornell University, Ithaca, NY 14853.
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D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002696:Chlamydomonas / Q000235:genetics* / Q000254:growth & development* / Q000378:metabolism; D002736:Chloroplasts / Q000378:metabolism; D003573:Cytochrome b Group / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D045346:Cytochrome b6f Complex; D003580:Cytochromes / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D045348:Cytochromes f; D004247:DNA / Q000302:isolation & purification / Q000378:metabolism; D017931:DNA Primers; D004579:Electron Transport; D017343:Genes, Plant; D007425:Intracellular Membranes / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D015388:Organelles / Q000378:metabolism; D010788:Photosynthesis; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis / Q000378:metabolism; D015183:Restriction Mapping; D013696:Temperature
D003573:Cytochrome b Group; D003580:Cytochromes; D017931:DNA Primers; D012333:RNA, Messenger; D004247:DNA; D045346:Cytochrome b6f Complex; D045348:Cytochromes f