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.
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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

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Figure 8

ChEL functions as a molecular chaperone for I-II-III.

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ChEL interaction improves recovery as well as secretion of Tg I-II-III. ...
(A) Cells were either untransfected (control) or transfected with a plasmid encoding I-II-III plus either empty vector or secretory ChEL-KDEL, as indicated. The transfected cells were pulse labeled for 30 minutes with 35S-labeled amino acids and chased for the times indicated. At each chase time, cells were immunoprecipitated with anti-Tg and newly synthesized I-II-III analyzed by nonreducing 4% SDS-PAGE and fluorography. Absence of recovery of I-II-III from untransfected control cells is shown at left. The band seen at time 0 and after chasing for 2 hours appears equivalent to that of the D isoform of full-length Tg. In the presence of the ChEL domain in the ER, a faster-migrating band equivalent to that of the mature E isoform of full-length Tg is detected (filled arrow). (B) Results of an experiment using the same cotransfection protocol and analysis as in A, except the second plasmid is either vector alone or secretory ChEL domain containing the cog mutation, the rdw mutation, or the KDEL appendage. Absence of recovery of I-II-III from untransfected 293 cells is shown in lane 1. The position of a 176-kDa molecular mass marker is shown at left.