1,721,026 research outputs found
patologia del metabolismo lipidico
No full textStruttura e metabolismo delle lipoproteine plasmatiche, Enzimi e trasportatori chiave del metabolismo delle lipoproteine, Recettori delle lipoproteine, Metabolismo delle lipoproteine contenenti apoB, Biogenesi delle HDL e trasporto inverso del colesterolo, Principali malattie del metabolismo lipidico
A new pathway activated by the unfolded protein response: Involvement of the TGF-beta/Smad pathway in the dedifferentiating effect of ER stress in thyroid cells
No full textConditions perturbing the homeostasis of the endoplasmic reticulum (ER) cause accumulation of unfolded proteins and trigger ER stress. In PC Cl3 thyroid cells, thapsigargin and tunicamycin interfered with the folding of thyroglobulin causing accumulation of this very large secretory glycoprotein in the ER. Consequently, BiP and XBP-1 mRNAs were induced and spliced, respectively. In the absence of apoptosis, differentiation of PC Cl3 cells was inhibited. mRNA and protein levels of thyroid specific genes, thyroglobulin, thyroperoxidase and sodium/iodide symporter, and of thyroid transcription factors, TTF-1, TTF-2 and Pax-8, were dramatically downregulated. These effects were, at least in part, transcriptional. Moreover, they were selective and temporally distinct from the general and transient PERK-dependent translational inhibition. Thyroid dedifferentiation was accompanied by changes in the organization of polarized epithelial monolayer. E-cadherin mRNA downregulation, vimentin, α-smooth muscle actin, α(1)(I) collagen, and SNAI1/SIP1 mRNAs up-regulation, actin stress fibers formation, and loss of transepithelial resistance were found, confirming an epithelial-mesenchymal transition (EMT). The thyroid-specific and epithelial dedifferentiation by thapsigargin/tunicamycin were completely prevented by the Src-family kinases inhibitor PP2 and by stable expression of a dominant negative Src. Together, these data indicate that ER stress induces dedifferentiation and an EMT-like phenotype in thyroid cells via a Src-mediated signaling pathway
Thyroglobulin from Molecular and Cellular Biology to Clinical Endocrinology
No full textThyroglobulin (Tg) is a vertebrate secretory protein synthesized in the thyrocyte endoplasmic reticulum (ER) where it acquires N-linked glycosylation and conformational maturation (including formation of many disulfide bonds), leading to homodimerization. Its primary functions include iodide storage and thyroid hormonogenesis. Tg consists largely of repeating domains, and many tyrosyl residues in these domains become iodinated to form monoiodo- and diiodotyrosine, whereas only a small portion of Tg structure is dedicated to hormone formation. Interestingly, evolutionary ancestors, dependent upon thyroid hormone for development, synthesize thyroid hormones without the complete Tg protein architecture. Nevertheless, in all vertebrates, Tg follows a strict pattern of region I, II-III, and the Cholinesterase-Like (ChEL) domain. In vertebrates, Tg first undergoes intracellular transport through the secretory pathway, which requires the assistance of thyrocyte ER chaperones and oxidoreductases, as well as coordination of distinct regions of Tg, to achieve a native conformation. Curiously, regions II-III and ChEL behave as fully independent folding units that could function as successful secretory proteins by themselves. However, the large Tg region I (bearing the primary thyroxine-forming site) is incompetent by itself for intracellular transport, requiring the downstream regions II-III and ChEL to complete its folding. A combination of nonsense mutations, frameshift mutations, splice site mutations, and missense mutations in Tg occur spontaneously to cause congenital hypothyroidism and thyroidal ER stress. These Tg mutants are unable to achieve a native conformation within the ER, interfering with the efficiency of Tg maturation and export to the thyroid follicle lumen for iodide storage and hormonogenesis
Maturation of thyroglobulin protein region I.
No full textIn vertebrates, the thyroglobulin (Tg) gene product must be exported to the lumen of thyroid follicles for thyroid hormone synthesis. In toto, Tg is composed of multiple type-1 repeats connected by linker and hinge (altogether considered as "region I," nearly 1,200 residues); regions II-III (~720 residues); and cholinesterase-like (ChEL) domain (~570 residues). Regions II-III and ChEL rapidly acquire competence for secretion, yet regions I-II-III require 20 min to become a partially mature disulfide isomer; stabilization of a fully oxidized form requires ChEL. Transition from partially mature to mature Tg occurs as a discrete "jump" in mobility by nonreducing SDS-PAGE, suggesting formation of at most a few final pairings of Cys residues that may be separated by significant intervening primary sequence. Using two independent approaches, we have investigated which portion of Tg is engaged in this late stage of its maturation. First, we demonstrate that this event is linked to oxidation involving region I. Introduction of the Tg-C1245R mutation in the hinge (identical to that causing human goitrous hypothyroidism) inhibits this maturation, although the Cys-1245 partner remains unidentified. Second, we find that Tg truncated after its fourth type-1 repeat is a fully independent secretory protein. Together, the data indicate that final acquisition of secretory competence includes conformational maturation in the interval between linker and hinge segments of region I
The cholinesterase-like domain of thyroglobulin functions as an intramolecular chaperone.
No full textThyroid 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
TSH-induced galactose incorporation at the NH2 terminus of thyroglobulin secreted by FRTL-5 cells.
No full textThe cholinesterase-like domain, essential in thyroglobulin trafficking for thyroid hormone synthesis, is required for protein dimerization.
No full textThe carboxyl-terminal cholinesterase-like (ChEL) domain of thyroglobulin (Tg) has been identified as critically important in Tg export from the endoplasmic reticulum. In a number of human kindreds suffering from congenital hypothyroidism, and in the cog congenital goiter mouse and rdw rat dwarf models, thyroid hormone synthesis is inhibited because of mutations in the ChEL domain that block protein export from the endoplasmic reticulum. We hypothesize that Tg forms homodimers through noncovalent interactions involving two predicted alpha-helices in each ChEL domain that are homologous to the dimerization helices of acetylcholinesterase. This has been explored through selective epitope tagging of dimerization partners and by inserting an extra, unpaired Cys residue to create an opportunity for intermolecular disulfide pairing. We show that the ChEL domain is necessary and sufficient for Tg dimerization; specifically, the isolated ChEL domain can dimerize with full-length Tg or with itself. Insertion of an N-linked glycan into the putative upstream dimerization helix inhibits homodimerization of the isolated ChEL domain. However, interestingly, co-expression of upstream Tg domains, either in cis or in trans, overrides the dimerization defect of such a mutant. Thus, although the ChEL domain provides a nidus for Tg dimerization, interactions of upstream Tg regions with the ChEL domain actively stabilizes the Tg dimer complex for intracellular transport
Thyroglobulin structure, function, and biosynthesis
No full textThe chapter includes phylogeny and ontogeny of thyroglobulin, the thyroglobulin gene and its mRNA, thyroglobulin folding, congenital goitrous hypothyroidism due to thyroglobulin misfolding, posttranslational modifications of thyroglobulin in later steps of the secretory pathway, iodination and iodotyrosyl coupling within thyroglobulin, thyroglobulin binding and internalization, proteolitic digestion of thyroglobulin in the endosome-lysosome system
Demonstration of a Ca++ requirement for thyroglobulin dimerization and export to the Golgi complex
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