The alternative oxidase genes and their expression in Chlamydomonas

Mitochondria of Chlamydomonas possess a terminal cyanide-resistant alternative oxidase (AOX) that reduces oxygen to water with electrons derived from the ubiquinone pool. Mutants inactivated in the cytochrome pathway of respiration are still able to consume oxygen via the alternative oxidase pathway. – By combining PCR, RT-PCR and screening of a Chlamydomonas cDNA library, two genes, AOX1 and AOX2, were identified. The cDNAs from AOX1 and AOX2 contain ORFs encoding predicted proteins of 360 and 347 amino acids, respectively. In both cases, a potential targeting presequence was identified in the protein sequence. The two AOX proteins have only 57.6% identity. Compared to AOX proteins from plants and fungi, the highest degree of identity (35-38%) was found with proteins from Aspergillus niger and Ajellomyces capsulatus. – The expression of AOX1 and AOX2 was analyzed at transcriptional level in wild-type and in dum19 mutant cells deprived of cytochrome c oxidase activity. Very low amounts of the 1.9-kb AOX2 transcript were present compared to the 2.3-kb AOX1 transcript. The expression of AOX1 was 2-6 times higher in the mutant than in wild-type, indicating that the absence of the cytochrome pathway of respiration enhances the transcriptional activity of the gene. Supported by FRFC grant 2.4527.97.