Published in Methods in Enzymology by group Reggiori

Monitoring the Formation of Autophagosomal Precursor Structures in Yeast Saccharomyces cerevisiae Chapter in Methods in Enzymology Volume 588 (Molecular Characterization of Autophagic Responses Part B, 1st Edition Jana Sánchez Wandelmer, Fulvio Reggiori & Rubén Gómez Sánchez Pdf

Abstract       

The budding  yeast  Saccharomyces  cerevisiae is a  powerful  and  versatile model  organism  for  studying multiple aspects of the biology of eukaryotic cells, including the molecular principles underlying autophagy. One of the unique advantages of this unicellular system is its amenability to genetic and biochemical approaches, which had a pivotal role in the discovery and characterization of most of the autophagy-related (Atg) proteins, the central players of autophagy. The relevance of investigating autophagy in this cell model lies in the high conservation of this pathway among eukaryotes, i.e., most of the yeast Atg proteins possess one or more mammalian orthologs. In addition to the experimental advantages, a very large collection of reagents keeps S. cerevisiae in a leading position for the study of the molecular mechanism and regulation of autophagy. In this chapter, we describe fluorescence microscopy and biochemical methods that allow to monitor in vivo the assembly the of Atg machinery, a key step of autophagy. These approaches can be very useful to those researchers that would like to assess the progression of the autophagosomal precursor structure formation under various conditions, in the presence of specific Atg protein mutants or in the absence of other factors.