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Related Experiment Video

Updated: Dec 12, 2025

Methods to Classify Cytoplasmic Foci as Mammalian Stress Granules

Methods to Classify Cytoplasmic Foci as Mammalian Stress Granules

Published on: May 12, 2017

An In Vitro Assembly System Identifies Roles for RNA Nucleation and ATP in Yeast Stress Granule Formation.

Kyle Begovich¹, James E Wilhelm¹

¹Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA; Howard Hughes Medical Institute (HHMI) Summer Institute Marine Biological Laboratory, Woods Hole, MA 02543, USA.

|August 12, 2020

Summary

Stress granules (SGs) form via RNA interactions. This study shows distinct RNA networks are crucial for canonical SG assembly, highlighting the interplay between transcriptome and ATP-dependent remodeling.

Keywords:

ATP RNA condensation RNA-RNA interactions reconstitution stress granules

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Monitoring Protein-RNA Interaction Dynamics In Vivo at High Temporal Resolution Using χCRAC

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Published on: May 9, 2020

Area of Science:

Cell Biology
Molecular Biology
Biochemistry

Background:

Stress granules (SGs) are dynamic cellular condensates formed from messenger ribonucleoprotein (mRNP) complexes.
SG assembly is driven by multivalent interactions, including protein-protein, protein-RNA, and RNA-RNA interactions.
The specific role of RNA-RNA interactions in SG formation is not well understood.

Purpose of the Study:

To investigate the role of RNA-RNA interactions in stress granule assembly.
To establish a yeast reconstitution system for studying SG formation triggered by specific RNAs.
To explore the contribution of distinct transcripts and ATP-dependent remodeling in canonical SG formation.

Main Methods:

Development of a yeast-based in vitro system to reconstitute stress granules.
Utilizing stem-loop RNA molecules as triggers for SG assembly.
Analyzing the ATP-sensitivity, helicase/chaperone regulation, and maturation of reconstituted SGs.
Investigating the phase-separation behavior of total RNA and specific transcripts like NFT1 mRNA.

Main Results:

Reconstituted SGs triggered by stem-loop RNAs exhibit ATP-sensitivity and hallmarks of maturation.
The phase-separated fraction of total cellular RNA is sufficient to initiate SG formation.
Condensation of a single transcript (NFT1 mRNA) results in an incomplete SG.
Distinct transcript networks appear necessary for canonical SG assembly.

Conclusions:

RNA-RNA interactions are critical for the proper assembly of canonical stress granules.
A network of diverse transcripts, not just individual ones, is required for complete SG formation.
The study provides a platform to dissect the interplay between the transcriptome and ATP-dependent remodeling in SG biogenesis.