Kinetic assay of starvation sensitivity in yeast autophagy mutants allows for the identification of intermediary phenotypes

BMC Research Notes 12, Article number: 505 (2019) | Download Citation

Abstract

Objective

A classical method to quantitatively determine the starvation sensitivity phenotype of autophagy mutant budding yeast strains is to starve them for a period of time and then to assess the proportion of cells that retain the ability to form colonies when the availability of nutrients is restored. The readout of this colony-formation assay is generally evaluated after a fixed period of time following the restoration of nutrients, so that it can be considered an endpoint assay. One drawback we have identified is the inability to characterize subtle intermediary phenotypes that are detectable at the molecular level but fail to reach statistical significance in the colony formation experiment. We set out to determine whether a more dynamic measurement of growth during recovery after starvation would increase the sensitivity with which we are able to detect partial loss-of-function phenotypes.

Results

We describe a 96-well plate-based assay to kinetically assess starvation sensitivity in budding yeast that allows for the quantitative detection of very modest starvation sensitivity phenotypes with statistical significance in autophagy mutant yeast strains lacking the ATG27 gene.