The impact of cysteine on the protein folding and export of the thyroid hormone precursor.

CINTIA E. CITTERIO; CINTHIA B. FIJALKOWKY; ARVAN, PETER

Simposio; Protein Folding Diseases Initiative Young Investigators Meeting.; 2021

The Impact of Cysteine on the Protein Folding and Export of the Thyroid Hormone PrecursorThyroglobulin (Tg), the thyroid hormone precursor protein (MW ~330kDa), represents ~50% of total thyroidal protein synthesis. Tg structure is stabilized by a multitude of disulfide bonds formed within the endoplasmic reticulum (ER). Oxidation of cysteines to cystine is critical for protein folding, but in thyrocytes, little is known about cytosolic cysteine availability to support Tg protein synthesis. In cystinosis, deficiency of a functional lysosomal cystine transporter (which contributes importantly to the cytosolic supply of cysteines) is linked to hypothyroidism. To better understand the importance of cysteine availability, we examined PCCL3 thyrocytes exposed to Cys-limited medium. We noted that by 24h of Cys-deprivation, Tg protein synthesis (and secretion) was dramatically decreased.The formation of disulfide pairs in the ER is thought to be catalyzed by a large group of ER resident oxidoreductases. We found that in PCCL3 cells, ER oxidoreductin-ɑ (Ero1ɑ) is positively regulated by TSH (thyroid stimulating hormone). To test the importance of Ero1ɑ for Tg folding and export, we co-expressed recombinant Tg +/- recombinant Ero1ɑ (in 293T cells) and observed that increased Ero1ɑ expression resulted in significantly increased Tg secretion. By contrast in PCCL3 cells, siRNA knockdown of Ero1ɑ resulted in decreased Tg expression and secretion. Taken together, these results suggest that both cysteine availability, and the capacity to catalyze disulfide bond formation in the ER, impact on Tg expression and its subsequent secretion leading to thyroid hormonogenesis. These findings may contribute to an understanding of the pathogenesis of hypothyroidism in cystinosis.