Stress-Triggered Phase Separation Is an Adaptive, Evolutionarily Tuned Response.

Area of Science:

• Cell Biology
• Biochemistry
• Molecular Biology

Background:

• Eukaryotic cells form stress granules from RNA-binding proteins during stress.
• In vitro studies show stress-associated proteins form liquid or hydrogel assemblies, often requiring non-physiological conditions.

Purpose of the Study:

• To investigate the phase separation behavior of poly(A)-binding protein (Pab1) under physiological stress conditions.
• To determine the role of Pab1's low-complexity region (LCR) and RNA in its phase separation.
• To assess the in vivo relevance of Pab1 phase separation for organismal fitness during stress.

Main Methods:

• In vitro phase separation assays of Pab1 under various stress conditions.
• Biophysical characterization of Pab1 and its mutants.
• In vivo studies involving yeast genetics and stress response assays.
• Analysis of evolutionary patterns to guide LCR mutagenesis.

Main Results:

• Pab1 undergoes phase separation and hydrogel formation in vitro upon exposure to physiological stress.
• Unlike other RNA-binding proteins, Pab1's LCR is not essential for demixing, and RNA inhibits its phase separation.
• Mutations in Pab1's LCR that impair phase separation decrease yeast fitness during prolonged stress.

Conclusions:

• Poly(A)-binding protein functions as a physiological stress sensor, utilizing phase separation to mark stress onset.
• Pab1's unique phase separation mechanism, independent of its LCR and inhibited by RNA, is crucial for stress response.
• This mechanism of stress sensing via phase separation is potentially broadly generalizable across biological systems.