Zebrafish mito-mKeima and mitoQC: Novel reporter lines for real-time mitophagy analysis during embryonic development.
Paul J. Wrighton1, Jin-Mi Heo2, J. Wade Harper2, Wolfram Goessling1,3,4,5
Brigham and Women's Hospital, Medicine, Boston, MA, USA
Inappropriate mitophagy is linked to aging, neurodegenerative diseases, drug-induced liver injury, and cancer. Newly developed, pH-dependent fluorescent reporters have been integrated into mouse models to enable light microscopy analysis of mitophagy in vivo. To extend the utility of such reporters, we generated several zebrafish mitophagy reporter lines expressing mito-mKeima or mitoQC (mito-EGFP-mCherry) under control of ubiquitous and liver-specific promoters. With these tools, we can monitor mitophagy live, in real-time during normal embryonic development, and in response to stress signals, as well as probe links between molecular mitophagy receptors and specific induction modes. We observed mitophagy hot spots during normal development including the eye, vasculature, heart, and kidney. Starvation induces quantifiable mitophagy in the skeletal muscle, and the mito-mkeima signal transiently colocates with LC3-positive autolysosomes. These novel zebrafish reporter lines and the advantages of the zebrafish model system facilitate time-lapse imaging to monitor mitophagy during vertebrate development and in repose to chemical and environmental insults. This system will permit future chemical and genetic screens for novel mitophagy inducers or inhibitors. Advances in gene editing technology also enable in vivo analysis of mitophagy molecular pathways with high time resolution and tissue specificity.
1 Department of Genetics, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115
2 Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA
3 Harvard Stem Cell Institute, Cambridge, Massachusetts 02138, USA
4 Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
5 Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.