HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates

McLoughlin, Fionn; Kim, Minsoo; Marshall, Richard S.; Vierstra, Richard D.; Vierling, Elizabeth

doi:10.1104/pp.19.00263

Title: HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates

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Abstract

Stressful environments often lead to protein unfolding and the formation of cytotoxic aggregates that can compromise cell survival. The molecular chaperone heat shock protein (HSP) 101 is a protein disaggregase that co-operates with the small HSP (sHSP) and HSP70 chaperones to facilitate removal of such aggregates and is essential for surviving severe heat stress. To better define how HSP101 protects plants, we investigated the localization and targets of this chaperone in Arabidopsis (Arabidopsis thaliana). By following HSP101 tagged with GFP, we discovered that its intracellular distribution is highly dynamic and includes a robust, reversible sequestration into cytoplasmic foci that vary in number and size among cell types and are potentially enriched in aggregated proteins. Affinity isolation of HSP101 recovered multiple proteasome subunits, suggesting a functional interaction. Consistent with this, the GFP-tagged 26S proteasome regulatory particle non-ATPase (RPN) 1a transiently colocalized with HSP101 in cytoplasmic foci during recovery. In addition, analysis of aggregated (insoluble) proteins showed they are extensively ubiquitylated during heat stress, especially in plants deficient in HSP101 or class I sHSPs, implying that protein disaggregation is important for optimal proteasomal degradation. Many potential HSP101 clients, identified by mass spectrometry of insoluble proteins, overlapped with known stress granule constituents andmore »« less

Authors:

^[1]; Kim, Minsoo ^[2];

^[3];

^[2]

Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, Massachusetts 01009, Department of Biology, Washington University in St. Louis, St. Louis, Missouri 63130
Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, Massachusetts 01009
Department of Biology, Washington University in St. Louis, St. Louis, Missouri 63130

Publication Date:: Tue May 21 00:00:00 EDT 2019

Research Org.:: Univ. of Massachusetts, Amherst, MA (United States)

Sponsoring Org.:: USDOE Office of Science (SC); National Institutes of Health (NIH); National Science Foundation (NSF)

OSTI Identifier:: 1515079

Alternate Identifier(s):: OSTI ID: 1610879

Grant/Contract Number:: SC0006646; GM-124452; IOS-1339325

Resource Type:: Published Article

Journal Name:: Plant Physiology (Bethesda)

Additional Journal Information:: Journal Name: Plant Physiology (Bethesda) Journal Volume: 180 Journal Issue: 4; Journal ID: ISSN 0032-0889

Publisher:: American Society of Plant Biologists

Country of Publication:: United States

Language:: English

Subject:: 59 BASIC BIOLOGICAL SCIENCES; Plant Sciences

Citation Formats


                    McLoughlin, Fionn, Kim, Minsoo, Marshall, Richard S., Vierstra, Richard D., and Vierling, Elizabeth. HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates.  United States: N. p., 2019. 
Web.  doi:10.1104/pp.19.00263.

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                    McLoughlin, Fionn, Kim, Minsoo, Marshall, Richard S., Vierstra, Richard D., & Vierling, Elizabeth. HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates.  United States.  https://doi.org/10.1104/pp.19.00263

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                    McLoughlin, Fionn, Kim, Minsoo, Marshall, Richard S., Vierstra, Richard D., and Vierling, Elizabeth. Tue .  
"HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates".  United States.  https://doi.org/10.1104/pp.19.00263.

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@article{osti_1515079,

  title        = {HSP101 Interacts with the Proteasome and Promotes the Clearance of Ubiquitylated Protein Aggregates},

  author       = {McLoughlin, Fionn and Kim, Minsoo and Marshall, Richard S. and Vierstra, Richard D. and Vierling, Elizabeth},

  abstractNote = {Stressful environments often lead to protein unfolding and the formation of cytotoxic aggregates that can compromise cell survival. The molecular chaperone heat shock protein (HSP) 101 is a protein disaggregase that co-operates with the small HSP (sHSP) and HSP70 chaperones to facilitate removal of such aggregates and is essential for surviving severe heat stress. To better define how HSP101 protects plants, we investigated the localization and targets of this chaperone in Arabidopsis (Arabidopsis thaliana). By following HSP101 tagged with GFP, we discovered that its intracellular distribution is highly dynamic and includes a robust, reversible sequestration into cytoplasmic foci that vary in number and size among cell types and are potentially enriched in aggregated proteins. Affinity isolation of HSP101 recovered multiple proteasome subunits, suggesting a functional interaction. Consistent with this, the GFP-tagged 26S proteasome regulatory particle non-ATPase (RPN) 1a transiently colocalized with HSP101 in cytoplasmic foci during recovery. In addition, analysis of aggregated (insoluble) proteins showed they are extensively ubiquitylated during heat stress, especially in plants deficient in HSP101 or class I sHSPs, implying that protein disaggregation is important for optimal proteasomal degradation. Many potential HSP101 clients, identified by mass spectrometry of insoluble proteins, overlapped with known stress granule constituents and sHSPinteracting proteins, confirming a role for HSP101 in stress granule function. Connections between HSP101, stress granules, proteasomes, and ubiquitylation imply that dynamic coordination between protein disaggregation and proteolysis is required to survive proteotoxic stress caused by protein aggregation at high temperatures.},

  doi          = {10.1104/pp.19.00263},

  journal      = {Plant Physiology (Bethesda)},

  number       = 4,

  volume       = 180,

  place        = {United States},

  year         = {Tue May 21 00:00:00 EDT 2019},

  month        = {Tue May 21 00:00:00 EDT 2019}

}

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