High resolution live cell imaging to define ultrastructural and dynamic features of the halotolerant yeastDebaryomyces hansenii

Abstract

ABSTRACTAlthough some budding yeasts have proved tractable and intensely studied models, others are more recalcitrant.Debaryomyces hansenii, an important yeast species in food and biotechnological industries with curious physiological characteristics, has proved difficult to manipulate genetically and remains poorly defined. To remedy this, we have combined live cell fluorescent dyes with high resolution imaging techniques to define the sub-cellular features ofD. hansenii, such as the mitochondria, nuclei, vacuoles and the cell wall. Using these tools, we define biological processes like the cell cycle, organelle inheritance and different membrane trafficking pathways ofD. hanseniifor the first time. Beyond this, reagents designed to studySaccharomyces cerevisiaeproteins were used to access proteomic information aboutD. hansenii. Finally, we optimised the use of label free holotomography to image yeast, defining the physical parameters and visualising sub-cellular features like membranes and vacuoles. Not only does this work shed light onD. hanseniibut this combinatorial approach serves as a template for how other cell biological systems, which are not amenable to standard genetic procedures, can be studied.