SUPPRESSION OF A -T-MUTATION BY A NEARBY SUBSTITUTION IN THE MITOCHONDRIAL COX1 GENE OF CHLAMYDOMONAS-REINHARDTII - A NEW-TYPE OF FRAMESHIFT SUPPRESSION IN AN ORGANELLE GENOME(1)

In Chlamydomonas reinhardtii, mutants defective in the cytochrome path way of respiration lack the capacity to grow under heterotrophic condi tions (in darkness on acetate). In the dark(-) strain dum18, a + 1 T a ddition in a run of four Ts, located at codon 145 of the mitochondrial cox1 gene encoding subunit I of cytochrome c oxidase, is responsible for the mutant phenotype. A leaky revertant (su11) that grows heterotr ophically at a lower rate than wild-type cells was isolated from dum18 . Its respiration sensitivity to cyanide was low and its cytochrome c oxidase activity was only 4% of that of the wild-type enzyme. Meiotic progeny obtained from crosses between revertant and wild-type cells in herited the phenotype of the mt(-) parent, showing that the suppressor mutation, like dum18 itself, is located in the mitochondrial genome. In order to map the su11 mutation relative to dum18, a recombinational analysis was performed on the diploid progeny. It demonstrated that s u11 was very closely linked to the dum18 mutation - less than 20-30 bp away. The cox1 gene of the su11 revertant was then sequenced. In addi tion to the + 1 T frameshift mutation still present at codon 145, an A --> C substitution was found at codon 146, leading to the replacement of a glutamic acid by an alanine in the polypeptide chain. No other m utations were detected in the cox1 coding sequence. As the new GCG cod on (Ala) created at position 146 is very seldom used in the mitochondr ial genome of C. reinhardtii, we suggest that the partial frameshift s uppression by the nearby substitution is due to an occasional abnormal translocation of the ribosome (+ 1 base shift) facilitated both by th e run of Ts and the low level of weak interaction of alanyl-tRNA.