Comparison of the proteome and of the phosphoproteome of S. cerevisiae cells harvested with different strategies

ABSTRACT: The budding yeast Saccharomyces cerevisiae is a powerful model system that is widely used to investigate many cellular processes. The harvesting of yeast cells is the first step in almost every experimental procedure. Here, yeast cells are isolated from their growth medium, collected, and used for successive experiments or analysis. The two most common methods to harvest S. cerevisiae are centrifugation and filtration. Understanding, if and how centrifugation and filtration affect yeast physiology is essential with respect to downstream data interpretation. Here, we profile and compare the proteomes and the phosphoproteomes, using isobaric label-based quantitative mass spectrometry, of three common methods used to harvest S. cerevisiae cells: low-speed centrifugation, high-speed centrifugation, and filtration. Our data suggest that while the proteome was stable across the tested conditions, hundreds of phosphorylation events were different between centrifugation and filtration. Our analysis shows that under our experimental conditions, filtration may cause both cell wall and osmotic stress at higher levels compared to centrifugation, and implying harvesting method-specific stresses. Thus, considering that the basal activation levels of specific stresses may differ under certain harvesting conditions is an important, but often overlooked aspect of experimental design.

INSTRUMENT(S): Orbitrap Eclipse, Orbitrap Fusion Lumos

ORGANISM(S): Saccharomyces Cerevisiae (baker's Yeast)

SUBMITTER: Joao Paulo  

LAB HEAD: Joao A. Paulo

PROVIDER: PXD044387 | Pride | 2023-12-28

REPOSITORIES: Pride