Importation of mitochodrial protein by an enhanced allotopic approach

Abstract

An expression vector containing appropriate mitochondrion-targeting sequences (MTS) and appropriate 3′UTR sequences provides efficient and stable delivery of a mRNA encoding a protein (CDS) to the mitochondrion of a mammalian cell. The MTS and 3′UTR sequences guide the CDS mRNA from the nuclear compartment of the cell to mitochondrion-bound polysomes, where the CDS is translated. This provides an efficient translocation of a mature functional protein into the mitochondria. A method of targeting mRNA expressed in the nuclear compartment of a mammalian cell to the mitochondrion is also provided. The vector and methods can be used to treat defects in mitochondrial function.

Description

FIELD OF THE INVENTION[0001]The present invention relates to the field of cell biology, molecular genetics, and medicine. It more particularly relates to the importation of proteins into the mitochondrion of animal and human cells.BACKGROUND OF THE INVENTION[0002]Mitochondria occupy a central position in the overall metabolism of eukaryotic cells; hence the oxidative phosphorylation (OXPHOS), the Krebs's cycle, the urea cycle, the heme biosynthesis and the fatty acid oxidation take place within the organelle. Recently, another major role for mitochondria in determining the cellular life span was established, as they are recognized to be a major early mediator in the apoptotic cascade. Mitochondria are also a major producer of reactive oxygen species (ROS) causing oxidative stress and therefore inducers of cell death.[0003]Primary defects in mitochondrial function are implicated in over 120 diseases and the list continues to grow, they encompass an extraordinary assemblage of clinica...

AI Technical Summary

Benefits of technology

[0013]The inventors describe an enhanced allotopic approach for importation of proteins into the mitochondrion. The present invention provides means, including compositions and methods, which enable mitochondrial importation at enhanced efficiency and stability compared to prior art techniques. The means of the invention enable a targeted localization of the mRNA to the mitochondrial surface.

[0014]Compared to prior art techniques, the means of the invention enable the efficient and stable importation of protein into the mitochondrion of an animal of human in need thereof, such as an animal or human having a cellular dysfunction caused by one or several mutations in a gene encoding a mitochondrial protein.

[0015]The inventors demonstrate that mRNA sorting to the mitochondrial surface is an efficient way to proceed to such an allotopic expression, and that this mRNA sorting can be controlled by selecting appropriate mitochondrion-targeting sequence (MTS) and appropriate 3′UTR sequences. The CDS sequence which codes for the protein to be delivered into the mitochondrion is guided by these appropriate MTS and 3′UTR sequences from the nuclear compartment to the mitochondrion-bound polysomes (where the CDS is translated), and aids in an efficient translocation of a mature functional protein into the mitochondria.

Problems solved by technology

Therefore, mitochondrial pathologies are considered among the most common genetically determined diseases, and are a major health issue since they remain inaccessible to both curative and palliative therapies.

Hence, up today important limitations are found to the allotopic expression as a therapeutic approach and require optimization to overcome the significant hurdles before it can be applied in genetic therapy.

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