New purifications reveal yeast and human stress granule cores are discrete particles with complex transcriptomes and proteomes

Abstract

Stress granules are a key cellular response, conserved across eukaryotes, to environmental insults. They arise upon stress-induced reversible stalling of protein synthesis and redistribution of mRNAs from polysomes into these large cytoplasmic ribonucleoprotein condensates. Many cellular pathways lead to formation and turnover of stress granules. Despite research spanning four decades, biochemical analysis of stress granules remains challenging because of their complexity and dynamic nature. Hitherto, stress granules have been studied cytologically, and their composition deduced from candidate protein approaches including immunoprecipitation and proximity labeling. Now, we have directly purified stress granule cores from yeast, relying on their density and size, finding that they are discrete particles with a size distribution peaking at 135 nm. Unexpectedly, we discovered that in addition to diverse proteins and RNAs, stress granule cores contain extrachromosomal circular DNA (eccDNA). These double-stranded DNAs have been found in all eukaryotes, but there is no precedent for a role in membraneless organelle biogenesis. By repurposing CRISPR technology to target DNA in the cytoplasm, we demonstrate that both presence and accessibility of eccDNA are required for stress granule formation in live yeast cells. Our finding reveals an unrecognized link between DNA metabolism and the cytoplasmic stress response, and suggests that sequence-specific eccDNA-mRNA interactions may mediate stress granule assembly.