title: Expression of algal nuclear ATP synthase subunit 6 in human cells results in protein targeting to mitochondria but no assembly into ATP synthase.
abstract: Artificial transfer of mitochondrial genes to the nucleus has implications for the understanding of mitochondrial function, evolution, and human health. Therefore, we created nuclear compatible versions of human subunit a (A6) of ATP synthase, linked to a mitochondrial targeting signal. Expression and targeting of human nuclear subunit a were compared to subunit a of Chlamydomonas reinhardtii, which naturally occurs in the nucleus. Algal subunit a was targeted to mitochondria more efficiently than human nuclear subunit a variants. However, there was no evidence of improved mitochondrial function in cultured cells; on the contrary, long-term expression of algal subunit a was associated with poor survival and intolerance of growth conditions that demand heavy reliance on oxidative phosphorylation. Analysis of enriched mitochondrial membrane fractions on native gels revealed a high-molecular- weight complex containing FLAG-tagged subunit a; however, this complex did not colocalize with ATP synthase. Thus, there was no evidence of assembly of algal subunit a into holoenzyme, nor did human nuclear subunit a colocalize with ATP synthase holoenzyme. In conclusion, obstacles remain to functional expression of mitochondrial genes transferred to the nucleus.
authors.affiliation: MRC-Dunn Human Nutrition Unit, Cambridge, United Kingdom.
authors.forename: Monika
authors.initials: M
authors.lastname: Bokori-Brown
authors.forename: Ian J
authors.initials: IJ
authors.lastname: Holt
mesh_terms: D020418:Algal Proteins / Q000502:physiology*; D000818:Animals; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D016825:Chlamydomonas reinhardtii; D004272:DNA, Mitochondrial / Q000502:physiology*; D005057:Eukaryotic Cells / Q000201:enzymology*; D018014:Gene Transfer Techniques; D006801:Humans; D025261:Mitochondrial Proton-Translocating ATPases / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology
chemical_list: D020418:Algal Proteins; D004272:DNA, Mitochondrial; C412204:ATP synthase subunit 6; D025261:Mitochondrial Proton-Translocating ATPases