Flow cytometric quantification and characterization of intracellular protein aggregates in yeast.

ABSTRACT: The sequestration of misfolded proteins into aggregates is an integral pathway of the protein quality control network that becomes particularly prominent during proteotoxic stress and in various pathologies. Methods for systematic analysis of cellular aggregate content are still largely limited to fluorescence microscopy and to separation by biochemical techniques. Here, we describe an alternative approach, using flow cytometric analysis, applied to protein aggregates released from their intracellular milieu by mild lysis of yeast cells. Protein aggregates were induced in yeast by heat shock or by chaperone deprivation and labeled using GFP- or mCherry-tagged quality control substrate proteins and chaperones. The fluorescence-labeled aggregate particles were distinguishable from cell debri

SUBMITTER: Shiber A 

PROVIDER: S-EPMC4601222 | biostudies-literature | 2014 May-Jun

REPOSITORIES: biostudies-literature