A new live-cell reporter strategy to simultaneously monitor mitochondrial biogenesis and morphology

• Published in: Scientific reports 2015-11, Vol.5 (1), p.17217-17217, Article 17217
• Main Authors: Hodneland Nilsson, Linn Iren, Nitschke Pettersen, Ina Katrine, Nikolaisen, Julie, Micklem, David, Avsnes Dale, Hege, Vatne Røsland, Gro, Lorens, James, Tronstad, Karl Johan
• Format: Article
• Language: English
• Subjects: 631/1647/245/2225; 631/80/642/333; 631/80/86/2366; 631/80/86/388; Adenylate Kinase - metabolism; Biochemical markers; Biomarkers - metabolism; Biomass; Biosynthesis; Cell culture; Confocal microscopy; Cytology; Gene Expression; Genes, Reporter; Green fluorescent protein; Green Fluorescent Proteins - biosynthesis; Green Fluorescent Proteins - genetics; HeLa Cells; Homeostasis; Humanities and Social Sciences; Humans; Image processing; Kinases; Mitochondria; Mitochondria - physiology; Mitochondria - ultrastructure; Mitochondrial DNA; Mitochondrial Dynamics; Morphology; multidisciplinary; Nuclear Respiratory Factor 1 - metabolism; Organelle Biogenesis; Protein kinase; Science; Single-Cell Analysis
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/srep17217

Changes in mitochondrial amount and shape are intimately linked to maintenance of cell homeostasis via adaptation of vital functions. Here, we developed a new live-cell reporter strategy to simultaneously monitor mitochondrial biogenesis and morphology. This was achieved by making a genetic reporter construct where a master regulator of mitochondrial biogenesis, nuclear respiratory factor 1 (NRF-1), controls expression of mitochondria targeted green fluorescent protein (mitoGFP). HeLa cells with the reporter construct demonstrated inducible expression of mitoGFP upon activation of AMP-dependent protein kinase (AMPK) with AICAR. We established stable reporter cells where the mitoGFP reporter activity corresponded with mitochondrial biogenesis both in magnitude and kinetics, as confirmed by biochemical markers and confocal microscopy. Quantitative 3D image analysis confirmed accordant increase in mitochondrial biomass, in addition to filament/network promoting and protecting effects on mitochondrial morphology, after treatment with AICAR. The level of mitoGFP reversed upon removal of AICAR, in parallel with decrease in mtDNA. In summary, we here present a new GFP-based genetic reporter strategy to study mitochondrial regulation and dynamics in living cells. This combinatorial reporter concept can readily be transferred to other cell models and contexts to address specific physiological mechanisms.