bims-algemi Biomed News
on Allotopic expression and gene therapy for mitochondrial disease
Issue of 2020‒03‒22
two papers selected by
Atif Towheed
Columbia University Irving Medical Center


  1. Methods Cell Biol. 2020 ;pii: S0091-679X(19)30159-1. [Epub ahead of print]155 441-487
    Bacman SR, Gammage PA, Minczuk M, Moraes CT.
      Most patients with mitochondrial DNA (mtDNA) mutations have a mixture of mutant and wild-type mtDNA in their cells. This phenomenon, known as mtDNA heteroplasmy, provides an opportunity to develop therapies by selectively eliminating the mutant fraction. In the last decade, several enzyme-based gene editing platforms were developed to cleave specific DNA sequences. We have taken advantage of these enzymes to develop reagents to selectively eliminate mutant mtDNA. The replication of intact mitochondrial genomes normalizes mtDNA levels and consequently mitochondrial function. In this chapter, we describe the methodology used to design and express these nucleases in mammalian cells in culture and in vivo.
    Keywords:  Gene therapy; Heteroplasmy; Mitochondrial diseases; mitoTALEN; mtDNA; mtZFN
    DOI:  https://doi.org/10.1016/bs.mcb.2019.12.004
  2. Methods Cell Biol. 2020 ;pii: S0091-679X(19)30136-0. [Epub ahead of print]155 45-79
    Priesnitz C, Pfanner N, Becker T.
      Mitochondria are deeply integrated into crucial functions of eukaryotic cells, including ATP production via oxidative phosphorylation, biosynthesis of iron-sulfur clusters, amino acids, lipids and heme, signaling pathways, and programmed cell death. The import of about 1000 different proteins that are produced as precursors on cytosolic ribosomes is essential for mitochondrial functions and biogenesis. The translocase of the outer mitochondrial membrane (TOM) forms the entry gate for the vast majority of mitochondrial proteins. Research of the last years has uncovered a complicated network of protein translocases and pathways that sort proteins into the mitochondrial subcompartments: outer and inner membranes, intermembrane space, and matrix. The in vitro import of a large number of different precursor proteins into mitochondria has been a pivotal experimental assay to identify these protein-sorting routes. This experimental set-up enables studies on the kinetics of protein transport into isolated mitochondria, on the processing of precursor proteins, and on their assembly into functional protein machineries. In vitro protein import assays are widely used and are indispensable for research on mitochondrial protein biogenesis.
    Keywords:  Blue native electrophoresis; Mitochondria; Protein assembly; Protein import; Protein sorting; TIM23 complex; TOM complex
    DOI:  https://doi.org/10.1016/bs.mcb.2019.11.006