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Cell Rep. 2019 Apr 30;27(5):1356-1363.e3. doi: 10.1016/j.celrep.2019.04.015.

Active Protein Neddylation or Ubiquitylation Is Dispensable for Stress Granule Dynamics.

Author information

1
Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Stem Cell Program, University of California, San Diego, La Jolla, CA 92093, USA; Institute for Genomic Medicine, University of California, San Diego, La Jolla, CA 92093, USA.
2
Cell and Developmental Biology, Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA.
3
Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Stem Cell Program, University of California, San Diego, La Jolla, CA 92093, USA; Institute for Genomic Medicine, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: geneyeo@ucsd.edu.
4
Cell and Developmental Biology, Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA. Electronic address: e1bennett@ucsd.edu.

Abstract

Stress granule (SG) formation is frequently accompanied by ubiquitin proteasome system (UPS) impairment and ubiquitylated protein accumulation. SGs, ubiquitin, and UPS components co-localize, but the relationship between the ubiquitin pathway and SGs has not been systematically characterized. We utilize pharmacological inhibition of either the ubiquitin- or NEDD8-activating enzyme (UAE or NAE) to probe whether active ubiquitylation or neddylation modulate SG dynamics. We show that UAE inhibition results in rapid loss of global protein ubiquitylation using ubiquitin-specific proteomics. Critically, inhibiting neither UAE nor NAE significantly affected SG formation or disassembly, indicating that active protein ubiquitylation or neddylation is dispensable for SG dynamics. Using antibodies with varying preference for free ubiquitin or polyubiquitin and fluorescently tagged ubiquitin variants in combination with UAE inhibition, we show that SGs co-localize primarily with unconjugated ubiquitin rather than polyubiquitylated proteins. These findings clarify the role of ubiquitin in SG biology and suggest that free ubiquitin may alter SG protein interactions.

KEYWORDS:

Nedd8; centrosome; neurodegeneration; oxidative stress; proteasome; protein aggregation; protein homeostasis; sodium arsenite; stress granule; ubiquitin

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