Measurement and modification of the expression level of the chaperone protein and signaling regulator GRP78/BiP in mammalian cells

Methods Enzymol. 2011;490:217-33. doi: 10.1016/B978-0-12-385114-7.00013-1.

Abstract

GRP78/BiP is a major endoplasmic reticulum (ER) chaperone protein essential for protein quality control in the ER as well as a central regulator of unfolded protein response (UPR). The induction of GRP78 is well established as a marker for ER stress. Recently, mouse models targeting the Grp78 allele indicate that GRP78 has critical roles in cancer progression, drug resistance, angiogenesis, neurological diseases, and diabetes. The discovery of a cytosolic GRP78 isoform and cell surface GRP78 adds new insights to its function beyond the ER compartment in regulating growth factor signaling and cell viability. Here, we summarize and update several approaches for the detection and quantitation of total GRP78, cytosolic GRP78 isoform, and cell surface GRP78, and the use of small interfering RNA to knockdown GRP78 expression. These techniques can be applied to culture cells as well as tissues.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Endoplasmic Reticulum Chaperone BiP
  • Heat-Shock Proteins / genetics
  • Heat-Shock Proteins / metabolism*
  • Humans
  • Mice
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction / methods
  • Signal Transduction / physiology*

Substances

  • Endoplasmic Reticulum Chaperone BiP
  • HSPA5 protein, human
  • Heat-Shock Proteins
  • Hspa5 protein, mouse
  • Protein Isoforms
  • RNA, Small Interfering