Monitoring Chaperone Engagement of Substrates in the Endoplasmic Reticulum of Live Cells

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 103; no. 17; pp. 6536 - 6541
• Main Authors: Snapp, Erik L., Sharma, Ajay, Lippincott-Schwartz, Jennifer, Hegde, Ramanujan S.
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 25-04-2006
• Subjects: Animals; Biochemistry; Biological Sciences; Calreticulin - deficiency; Calreticulin - genetics; Calreticulin - metabolism; Cell Line; Cells; Cellular biology; Cellular metabolism; Endoplasmic reticulum; Endoplasmic Reticulum - metabolism; Enzymes; Fibroblasts; Fluorescence; Fluorescence Recovery After Photobleaching; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Homeostasis; Lectins; Mice; Molecular Chaperones - genetics; Molecular Chaperones - metabolism; Protein Folding; Protein synthesis; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Substrates; Transfection; Treaties
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.0510657103

The folding environment in the endoplasmic reticulum (ER) depends on multiple abundant chaperones that function together to accommodate a range of substrates. The ways in which substrate engagement shapes either specific chaperone dynamics or general ER attributes in vivo remain unknown. In this study, we have evaluated how changes in substrate flux through the ER influence the diffusion of both the lectin chaperone calreticulin and an inert reporter of ER crowdedness. During acute changes in substrate load, the inert probe revealed no changes in ER organization, despite significant changes in calreticulin dynamics. By contrast, inhibition of the lectin chaperone system caused rapid changes in the ER environment that could be reversed over time by easing new substrate burden. Our findings provide insight into the normal organization and dynamics of an ER chaperone and characterize the capacity of the ER to maintain homeostasis during acute changes in chaperone activity and availability.