A Phos-tag-based approach reveals the extent of physiological endoplasmic reticulum stress

PLoS One. 2010 Jul 16;5(7):e11621. doi: 10.1371/journal.pone.0011621.

Abstract

Cellular response to endoplasmic reticulum (ER) stress or unfolded protein response (UPR) is a key defense mechanism associated with many human diseases. Despite its basic and clinical importance, the extent of ER stress inflicted by physiological and pathophysiological conditions remains difficult to quantitate, posing a huge obstacle that has hindered our further understanding of physiological UPR and its future therapeutic potential. Here we have optimized a Phos-tag-based system to detect the activation status of two proximal UPR sensors at the ER membrane. This method allowed for a quantitative assessment of the level of stress in the ER. Our data revealed quantitatively the extent of tissue-specific basal ER stress as well as ER stress caused by the accumulation of misfolded proteins and the fasting-refeeding cycle. Our study may pave the foundation for future studies on physiological UPR, aid in the diagnosis of ER-associated diseases and improve and facilitate therapeutic strategies targeting UPR in vivo.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blotting, Western
  • Cell Line
  • Endoplasmic Reticulum / metabolism*
  • Endoribonucleases / metabolism
  • Humans
  • Mice
  • Phosphorylation
  • Polymerase Chain Reaction
  • Protein Folding
  • Protein-Serine-Threonine Kinases / metabolism
  • Pyridines / chemistry*
  • Stress, Physiological / genetics
  • Stress, Physiological / physiology*
  • eIF-2 Kinase / metabolism

Substances

  • 1,3-bis(bis(pyridin-2-ylmethyl)amino)propan-2-ol
  • Pyridines
  • ERN1 protein, human
  • PERK kinase
  • Protein-Serine-Threonine Kinases
  • eIF-2 Kinase
  • Endoribonucleases