Live-cell imaging of mitochondrial motility and interactions in Drosophila neurons and yeast

Mitochondria are highly dynamic organelles that undergo directed movement and anchorage, which in turn are critical for calcium buffering and energy mobilization at specific regions within cells or at sites of contact with other organelles. Physical and functional interactions between mitochondria and other organelles also impact processes, including phospholipid biogenesis and calcium homeostasis. Indeed, mitochondrial motility, localization, and interaction with other organelles are compromised in many neurodegenerative diseases. Here, we describe methods to visualize and carry out quantitative analysis of mitochondrial movement in two genetically-manipulatable, widely-used model systems: Drosophila neurons and the budding yeast, Saccharomyces cerevisiae. We also describe approaches for multi-color imaging in living yeast cells that may be used to visualize colocalization of proteins within mitochondria, as well as interactions of mitochondria with other organelles.