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Characterization of ThOX proteins as components of the thyroid H2O2-generating system

par De Deken, Xavier ;Wang, Dantong ;Dumont, Jacques ;Miot, Françoise
Référence Experimental Cell Research, 273, 2, page (187-196)
Publication Publié, 2002

Article révisé par les pairs

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Résumé :

We have recently cloned two thyroid-specific cDNAs encoding new members of the NADPH oxidase family. ThOX1 and ThOX2 proteins are colocalized with thyroperoxidase at the apical membrane of human thyroid cells. In the present study we have determined their subcellular localization and maturation in relation to their enzymatic activity. A majority of ThOX proteins accumulated inside the cell and only a small fraction was expressed at the surface. Western blots demonstrated that ThOX's are glycoproteins of 180,000 and 190,000. When totally deglycosylated the molecular weight of both ThOX1 and ThOX2 drops to 160,000. Ca2+ stimulates the basal H2O2 generation in PC Cl3 cells at a level corresponding to 20% of the leukocyte H2O2 production stimulated by PMA. Nonthyroid cell lines transfected with ThOX1 and ThOX2 show only a single immunoreactive band in Western blot analysis, corresponding to the protein of 180,000. This "immature" protein remains exclusively intracellular and does not present any enzymatic activity. This is not modified by coexpression of thyroperoxidase and p22Phox. Transfection of ThOX cDNAs into PLB-XCGD cells does not reconstitute their NADPH oxidase activity. We conclude that (1) the thyroid contains some elements of the leukocyte H2O2-generating system but not all of them; (2) ThOX's are predominantly or exclusively located inside the cell in thyrocytes or in transfected cells, respectively, and as such they are inactive; (3) ThOX's cannot replace gp91Phox in the leukocyte; and (4) the thyroid H2O2-generating system is analogous to the leukocyte system with regard to ThOX's and gp91Phox but very different in other aspects. Additional thyroid-specific components are probably required to get complete protein processing and full enzymatic activity in the thyroid. © 2002 Elsevier Science (USA).