Revealing Protein Aggregates under Thapsigargin-Induced ER Stress Using an ER-Targeted Thioflavin

Peter Verwilst, Kyutae Kim, Kyoung Sunwoo, Hye Ri Kim, Chulhun Kang, Jong Seung Kim

Research output: Contribution to journalArticle

Abstract

Endoplasmic reticulum-thioflavin T (ER-ThT), a thioflavin T-based fluorescent chemosensor, was developed to detect protein aggregates in the endoplasmic reticulum (ER) and was applied to live cells under various forms of ER stress. Upon dithiothreitol (DTT)-induced reductive denaturation of lysozyme and albumin, the intensity was increased in a protein concentration-dependent way, following a nonfluorescent lag phase. ER-ThT detects protein aggregates rather than unfolded proteins in solution, and the protein aggregation can be visualized in the presence of lipid membranes or native proteins. Within live HeLa cells, ER-ThT is localized in the ER and its fluorescence was dramatically increased upon ER stress induction by DTT, Thapsigargin, or Brefeldin A. Moreover, in the presence of ER stress modulators (tauroursodeoxycholic acid, trimethylamine N-oxide, or 4-phenylbutyric acid), also known as chemical chaperones, the fluorescence under Thapsigargin treatment was suppressed to the level of the control group. Thus, ER-ThT is capable of detecting the accumulation of protein aggregates under ER stress in living cells and acts as an in vitro screening tool for ER stress modulators, putative prodrugs against ER-related proteopathy. Overall, the results strongly suggest that protein aggregation is intricately involved in the activation of the unfolded protein response following ER stress.

Original languageEnglish
Pages (from-to)2858-2863
Number of pages6
JournalACS Sensors
Volume4
Issue number11
DOIs
Publication statusPublished - 2019 Nov 22

Fingerprint

endoplasmic reticulum
Thapsigargin
proteins
Proteins
Dithiothreitol
Modulators
Agglomeration
Fluorescence
Brefeldin A
Denaturation
Prodrugs
Membrane Lipids
Muramidase
Protein Aggregates
Albumins
Screening
modulators
Acids
Chemical activation
Cells

Keywords

  • chemical chaperones
  • ER stress
  • fluorescence
  • protein aggregation
  • thioflavin

ASJC Scopus subject areas

  • Bioengineering
  • Instrumentation
  • Process Chemistry and Technology
  • Fluid Flow and Transfer Processes

Cite this

Revealing Protein Aggregates under Thapsigargin-Induced ER Stress Using an ER-Targeted Thioflavin. / Verwilst, Peter; Kim, Kyutae; Sunwoo, Kyoung; Kim, Hye Ri; Kang, Chulhun; Kim, Jong Seung.

In: ACS Sensors, Vol. 4, No. 11, 22.11.2019, p. 2858-2863.

Research output: Contribution to journalArticle

Verwilst, Peter ; Kim, Kyutae ; Sunwoo, Kyoung ; Kim, Hye Ri ; Kang, Chulhun ; Kim, Jong Seung. / Revealing Protein Aggregates under Thapsigargin-Induced ER Stress Using an ER-Targeted Thioflavin. In: ACS Sensors. 2019 ; Vol. 4, No. 11. pp. 2858-2863.
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