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Updated: Jun 13, 2026
Methods to Classify Cytoplasmic Foci as Mammalian Stress Granules
Published on: May 12, 2017
mRNA escape from stress granule sequestration is dictated by localization to the endoplasmic reticulum
H Unsworth1, S Raguz, H J Edwards
1Department of Oncology, Imperial College London, Hammersmith Hospital Campus, Du Cane Road, London W12 0NN, UK.
Abstract:
In mammalian cells, cytotoxic stress triggers several signaling cascades that converge in the phosphorylation of translation initiation factor 2alpha, shuttling of nuclear RNA-binding proteins such as TIA-1 to the cytoplasm, and aggregation of most cellular mRNAs into TIA-1-containing stress granules (SGs). As a result, protein synthesis is greatly impaired. Here we describe different dynamics of endogenous transcripts according to their cellular location, in response to stress. While cytosolic mRNAs aggregate into SGs, endoplasmic reticulum (ER) -bound transcripts escape sequestration. This has been specifically demonstrated using the multidrug resistance transporter gene (MDR1) as a model and showing that chimeric RNA constructs can be directed to the cytosol or tethered to the ER depending on the nature of the chimera, in response to stress. In addition, polysome profile analyses indicate that, on stress, ribosomes do not disengage from ER-associated transcripts (puromycin insensitive) and recover their translation status faster than SG-targeted cytosolic mRNAs once the stress is lifted. These findings have important implications for cell survival given that many membrane proteins, which are translated at the ER, have important roles in detoxification.
Insights
Cytotoxic stress causes mRNA aggregation into stress granules, impairing protein synthesis. However, ER-bound transcripts evade this, aiding cell survival by enabling rapid recovery of essential protein translation.
Area of Science:
- Cellular Biology
- Molecular Biology
- Stress Response
Background:
- Cytotoxic stress in mammalian cells activates signaling pathways leading to impaired protein synthesis.
- Stress granules (SGs) form from aggregated mRNAs and RNA-binding proteins, sequestering most cellular transcripts.
Purpose of the Study:
- To investigate the differential dynamics of endogenous transcripts based on their cellular location during stress.
- To understand how ER-bound transcripts behave differently from cytosolic transcripts under stress conditions.
Main Methods:
- Utilized the multidrug resistance transporter gene (MDR1) as a model system.
- Constructed chimeric RNAs to direct transcripts to the cytosol or ER.
- Performed polysome profile analysis to assess translation status.
Main Results:
- Cytosolic mRNAs aggregate into stress granules, while ER-bound transcripts remain accessible.
- Ribosomes remain associated with ER-bound transcripts during stress (puromycin insensitive).
- ER-associated transcripts recover translation status faster than SG-targeted cytosolic mRNAs post-stress.
Conclusions:
- Cellular location dictates mRNA fate during stress, with ER-tethered transcripts escaping SG sequestration.
- This differential mRNA handling is crucial for cell survival, particularly for membrane proteins involved in detoxification.
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