Live-Cell Imaging of Mitochondria and the Actin Cytoskeleton in Budding Yeast.
Maintenance and regulation of proper mitochondrial dynamics and functions are necessary for cellular homeostasis. Numerous diseases, including neurodegeneration and muscle myopathies, and overall cellular aging are marked by declining mitochondrial function and subsequent loss of multiple other cellular functions. For these reasons, optimized protocols are needed for visualization and quantification of mitochondria and their function and fitness. In budding yeast, mitochondria are intimately associated with the actin cytoskeleton and utilize actin for their movement and inheritance. This chapter describes optimal approaches for labeling mitochondria and the actin cytoskeleton in living budding yeast cells, for imaging the labeled cells, and for analyzing the resulting images.
Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, 630 W. 168th Street, New York, NY, 10032, USA.
Ryo
R
Higuchi-Sanabria
Confocal and Specialized Microscopy Shared Resource, Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY, USA.
Theresa C
TC
Swayne
Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, 630 W. 168th Street, New York, NY, 10032, USA.
Istvan R
IR
Boldogh
Department of Pathology and Cell Biology, College of Physicians and Surgeons, Columbia University, 630 W. 168th Street, New York, NY, 10032, USA. lap5@cumc.columbia.edu.|Confocal and Specialized Microscopy Shared Resource, Herbert Irving Comprehensive Cancer Center, Columbia University, New York, NY, USA. lap5@cumc.columbia.edu.
Liza A
LA
Pon
D000085:Acetates / Q000494:pharmacology; D008841:Actin Cytoskeleton / Q000378:metabolism*; D015825:Chromosomes, Fungal / Q000235:genetics; D004396:Coloring Agents / Q000378:metabolism; D056426:Genetic Loci / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D019426:Integrases / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism*; D057054:Molecular Imaging / Q000379:methods*; D009068:Movement; D061848:Optical Imaging; D016133:Polymerase Chain Reaction; D004718:Saccharomycetales / Q000166:cytology* / Q000187:drug effects / Q000235:genetics; D017421:Sequence Analysis; D014170:Transformation, Genetic / Q000187:drug effects
D000085:Acetates; D004396:Coloring Agents; C045073:Cre recombinase; D019426:Integrases; C488804:lithium acetate
Actin; Cytoskeleton; Deconvolution; Fluorescent proteins; Live-cell imaging; Microscopy; Mitochondria; Ratio imaging; Vital staining; Yeast