Enhanced binding to the molecular chaperone BiP slows thyroglobulin export from the endoplasmic reticulum.
To examine how binding of BiP (a molecular chaperone of the hsp70 family that resides in the endoplasmic reticulum) influences the conformational maturation of thyroglobulin (Tg, the precursor for thyroid hormone synthesis), we have developed a system of recombinant Tg stably expressed in wild-type Chinese hamster ovary (CHO) cells and CHO-B cells genetically manipulated for selectively increased BiP expression. The elevation of immunoreactive BiP in CHO-B cells is comparable to that seen during the unfolded protein response in the thyrocytes of certain human patients and animals suffering from congenital hypothyroid goiter with defective Tg. However, in CHO-B cells, we expressed Tg containing no mutations that induce misfolding (i.e. no unfolded protein response), so that levels of all other endoplasmic reticulum chaperones were normal. Increased availability of BiP did not accelerate Tg secretion; rather, the export of newly synthesized Tg was delayed. Tg detained intracellularly was concentrated in the endoplasmic reticulum. By coimmunoprecipitation, BiP exhibited enhanced binding to Tg in CHO-B cells. Moreover, two-dimensional gel analysis showed that BiP associated especially well with intracellular Tg containing mispaired disulfide bonds, thought to represent early Tg folding intermediates. An endoplasmic reticulum chaperone of the hsp90 family, GRP94, was also associated in Tg-chaperone complexes. The results suggest that increased binding of BiP to Tg leads to its delayed conformational maturation in the endoplasmic reticulum.
Muresan
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Z
Program in Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts 02215, USA.
Arvan
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P
D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002352:Carrier Proteins / Q000378:metabolism*; D006224:Cricetinae; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D006031:Glycosylation; D006360:Heat-Shock Proteins; D006596:Hexosaminidases / Q000378:metabolism; D015151:Immunoblotting; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones / Q000378:metabolism*; D017510:Protein Folding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013954:Thyroglobulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D002352:Carrier Proteins; D003432:Cross-Linking Reagents; D000091342:Endoplasmic Reticulum Chaperone BiP; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D011994:Recombinant Proteins; D013954:Thyroglobulin; D006596:Hexosaminidases