FIGURE 3.

Pet54 plays a role in COX1 mRNA untranslated regions that depends on the presence of the Cox1 protein. A, the cox1Δ::ARG8^m construct was inserted at the COX1 locus, where the ARG8^m reporter replaced the coding sequence of COX1. In addition, the COX1 coding sequence flanked by the COX2 UTRs was inserted at an ectopic site, 295 bp upstream of COX2 on the mtDNA (20). Mitochondria from cells carrying the wild-type mtDNA or the ectopic chimeric mtDNA were analyzed by SDS-PAGE and Western blotting. The membrane was probed with an antibody for Cox1 and afterward for citrate synthase (CS) as a loading control. The Δpet111 mutation was introduced as indicated. B, serial dilutions of the indicated mutants bearing the ectopic chimeric mtDNA were spotted on medium lacking (−ARG) or containing (+ARG) arginine and were grown for 2 days at 30 °C. C, cells carrying the ectopic chimeric mtDNA and either an empty plasmid or a plasmid expressing a high copy number of PET111 were grown on medium lacking (−ARG) or containing (+ARG) arginine. Serial dilutions were grown for 2 days at 30 °C. D, the COX1 coding region was replaced by the reporter gene ARG8^m (cox1Δ::ARG8^m); however, in this construct, no chimeric COX1 construct was inserted. The indicated mutants bearing this mtDNA were grown on serial dilutions as in B.