The cholinesterase-like domain of thyroglobulin functions as an intramolecular chaperone
Jaemin Lee, … , Bruno Di Jeso, Peter Arvan
Jaemin Lee, … , Bruno Di Jeso, Peter Arvan
Published July 1, 2008
Citation Information: J Clin Invest. 2008;118(8):2950-2958. https://doi.org/10.1172/JCI35164.
View: Text | PDF
Research Article Endocrinology

The cholinesterase-like domain of thyroglobulin functions as an intramolecular chaperone

Abstract

Thyroid hormonogenesis requires secretion of thyroglobulin, a protein comprising Cys-rich regions I, II, and III (referred to collectively as region I-II-III) followed by a cholinesterase-like (ChEL) domain. Secretion of mature thyroglobulin requires extensive folding and glycosylation in the ER. Multiple reports have linked mutations in the ChEL domain to congenital hypothyroidism in humans and rodents; these mutations block thyroglobulin from exiting the ER and induce ER stress. We report that, in a cell-based system, mutations in the ChEL domain impaired folding of thyroglobulin region I-II-III. Truncated thyroglobulin devoid of the ChEL domain was incompetent for cellular export; however, a recombinant ChEL protein (“secretory ChEL”) was secreted efficiently. Coexpression of secretory ChEL with truncated thyroglobulin increased intracellular folding, promoted oxidative maturation, and facilitated secretion of region I-II-III, indicating that the ChEL domain may function as an intramolecular chaperone. Additionally, we found that the I-II-III peptide was cosecreted and physically associated with secretory ChEL. A functional ChEL domain engineered to be retained intracellularly triggered oxidative maturation of I-II-III but coretained I-II-III, indicating that the ChEL domain may also function as a molecular escort. These insights into the role of the ChEL domain may represent potential therapeutic targets in the treatment of congenital hypothyroidism.

Authors

Jaemin Lee, Bruno Di Jeso, Peter Arvan

×

Figure 9

Oxidation state of secreted I-II-III.

Options: View larger image (or click on image) Download as PowerPoint
ChEL functions as a molecular chaperone for I-II-III. (A) Cells were eit...
293 cells were transfected either with empty vector (control) or with secretory ChEL bearing or lacking the KDEL appendage. The cells were pulse labeled for 30 min with 35S-labeled amino acids and then chased for the times indicated (control cells were chased for 2 hours). The cells were lysed, and both lysates and chase medium immunoprecipitated with anti-Tg and subjected to PNGase F digestion to remove N-glycans. The samples were then analyzed by nonreducing 5.5% SDS-PAGE. Under these conditions, it is apparent that secreted I-II-III closely migrates with the intracellular E isomer of I-II-III. A black line has been added to distinguish the medium from the cell lysate. The position of a 176-kDa molecular mass marker is shown at left.