Effect of iodide on nicotinamide adenine dinucleotide phosphate oxidase activity and Duox2 protein expression in isolated porcine thyroid follicles.
Thyroperoxidase requires H(2)O(2) to catalyze the biosynthesis of thyroxine residues on thyroglobulin. Iodide inhibits the generation of H(2)O(2), and consequently the synthesis of thyroid hormones (Wolff-Chaikoff effect). The H(2)O(2) generator is a calcium-dependent nicotinamide adenine dinucleotide phosphate (NADPH) oxidase involving the flavoprotein Duox2. NADPH oxidase activity and Duox2 require cAMP to be expressed in pig thyrocytes. We studied the effect of iodide on NADPH oxidase activity, the DUOX2 gene, and Duox2 protein expression in pig thyroid follicles cultured for 48 h with forskolin or a cAMP analog. Iodide inhibited the cellular release of H(2)O(2) and NADPH oxidase activity, effects prevented by methimazole. Northern blot studies showed that iodide did not reduce DUOX2 mRNA levels but did reduce those of TPO and NIS. Western blot analyses using a Duox2-specific antipeptide showed that Duox2 has two N-glycosylation states, which have oligosaccharide motifs accounting for about 15 kDa and 25 kDa, respectively, of the apparent molecular mass. Cyclic AMP increased the amount of the highly glycosylated form of Duox2, an effect antagonized by iodide in a methimazole-dependent manner. These data suggest that an oxidized form of iodide inhibits the H(2)O(2) generator at a posttranscriptional level by reducing the availability of the mature Duox2 protein.
Morand
Stanislas
S
Unité 486, Institut National de la Santé et de la Recherche Médicale, Université Paris 11, Faculté de Pharmacie, 92296 Châtenay-Malabry Cedex, France.
Chaaraoui
Mokhtar
M
Kaniewski
Jacques
J
Dème
Danielle
D
Ohayon
Renée
R
Noel-Hudson
Marie Sophie
MS
Virion
Alain
A
Dupuy
Corinne
C
D000818:Animals; D015153:Blotting, Western; D002458:Cell Fractionation; D002462:Cell Membrane / Q000201:enzymology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D000074623:Dual Oxidases; D004789:Enzyme Activation / Q000187:drug effects; D005420:Flavoproteins; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000378:metabolism; D066298:In Vitro Techniques; D007454:Iodides / Q000494:pharmacology*; D009249:NADP / Q000378:metabolism*; D019255:NADPH Oxidases / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D013552:Swine; D013961:Thyroid Gland / Q000187:drug effects* / Q000201:enzymology*
D005420:Flavoproteins; D007454:Iodides; D012333:RNA, Messenger; D005576:Colforsin; D009249:NADP; D006861:Hydrogen Peroxide; D000242:Cyclic AMP; D000074623:Dual Oxidases; D019255:NADPH Oxidases; C431513:DUOX1 protein, human