mRNA localization to the mitochondrial surface allows the efficient translocation inside the organelle of a nuclear recoded ATP6 protein.
As previously established in yeast, two sequences within mRNAs are responsible for their specific localization to the mitochondrial surface-the region coding for the mitochondrial targeting sequence and the 3'UTR. This phenomenon is conserved in human cells. Therefore, we decided to use mRNA localization as a tool to address to mitochondria, a protein that is not normally imported. For this purpose, we associated a nuclear recoded ATP6 gene with the mitochondrial targeting sequence and the 3'UTR of the nuclear SOD2 gene, which mRNA exclusively localizes to the mitochondrial surface in HeLa cells. The ATP6 gene is naturally located into the organelle and encodes a highly hydrophobic protein of the respiratory chain complex V. In this study, we demonstrated that hybrid ATP6 mRNAs, as the endogenous SOD2 mRNA, localize to the mitochondrial surface in human cells. Remarkably, fusion proteins localize to mitochondria in vivo. Indeed, ATP6 precursors synthesized in the cytoplasm were imported into mitochondria in a highly efficient way, especially when both the MTS and the 3'UTR of the SOD2 gene were associated with the re-engineered ATP6 gene. Hence, these data indicate that mRNA targeting to the mitochondrial surface represents an attractive strategy for allowing the mitochondrial import of proteins originally encoded by the mitochondrial genome without any amino acid change in the protein that could interfere with its biologic activity.
Kaltimbacher
Valérie
V
Laboratoire de Physiopathologie Cellulaire et Moléculaire de la Rétine, INSERM U592 and Université Pierre et Marie Curie (UPMC-Paris6), Hôpital St. Antoine, 75571 Paris, Cedex 12 France.
Bonnet
Crystel
C
Lecoeuvre
Gaëlle
G
Forster
Valérie
V
Sahel
José-Alain
JA
Corral-Debrinski
Marisol
M
D020413:3' Untranslated Regions / Q000378:metabolism; D001483:Base Sequence; D001692:Biological Transport; D002467:Cell Nucleus / Q000378:metabolism*; D017931:DNA Primers; D006367:HeLa Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism* / Q000648:ultrastructure*; D025261:Mitochondrial Proton-Translocating ATPases / Q000378:metabolism*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013482:Superoxide Dismutase / Q000378:metabolism; D000099156:Superoxide Dismutase 2
D020413:3' Untranslated Regions; D017931:DNA Primers; D025261:Mitochondrial Proton-Translocating ATPases; D012333:RNA, Messenger; D013482:Superoxide Dismutase; D000099156:Superoxide Dismutase 2; C501005:MT-ATP6 protein, human