Investigation of the ultrastructure of yeast using cryo-immuno-electron microscopy

Yeast has been a crucial model system for the investigation of a multitude of cellular processes and many approaches have been used to study these, including genetics, biochemistry and fluorescence microscopy approaches. The morphological analyses of these organisms by electron microscopy (EM), however, have remained limited because of the lack of techniques leading to satisfactory ultrastructural resolution. In this compendium, we will review the various applications of an immuno-electron microscopy (IEM) protocol we have recently developed from the Tokuyasu method. This novel approach, which allows excellent cell preservation and unprecedented resolution of yeast morphology, has been successfully applied not only for the study of Saccharomyces cerevisiae and Pichia pastoris but also the filamentous fungus Aspergillus nidulans. This procedure is compatible with immuno-gold labelling and allows the combination of protein localization with a fine ultrastructural resolution of protein complexes, vesicular carriers and organelles. This new IEM protocol is thus a valuable tool for the large community of scientists studying the physiology of yeast but also exploiting these organisms as model systems for the investigation of molecular principles important cellular pathways conserved among eukaryotes