Type 2 deiodinase polymorphism causes ER stress and hypothyroidism in the brain
Sungro Jo, Tatiana L. Fonseca, Barbara M. L. C. Bocco, Gustavo W. Fernandes, Elizabeth A. McAninch, Anaysa P. Bolin, Rodrigo R. Da Conceição, Joao Pedro Werneck-de-Castro, Daniele L. Ignacio, Péter Egri, Dorottya Németh, Csaba Fekete, Maria Martha Bernardi, Victoria D. Leitch, Naila S. Mannan, Katharine F. Curry, Natalie C. Butterfield, J.H. Duncan Bassett, Graham R. Williams, Balázs Gereben, Miriam O. Ribeiro, Antonio C. Bianco
Sungro Jo, Tatiana L. Fonseca, Barbara M. L. C. Bocco, Gustavo W. Fernandes, Elizabeth A. McAninch, Anaysa P. Bolin, Rodrigo R. Da Conceição, Joao Pedro Werneck-de-Castro, Daniele L. Ignacio, Péter Egri, Dorottya Németh, Csaba Fekete, Maria Martha Bernardi, Victoria D. Leitch, Naila S. Mannan, Katharine F. Curry, Natalie C. Butterfield, J.H. Duncan Bassett, Graham R. Williams, Balázs Gereben, Miriam O. Ribeiro, Antonio C. Bianco
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Research Article Endocrinology Metabolism

Type 2 deiodinase polymorphism causes ER stress and hypothyroidism in the brain

Abstract

Levothyroxine (LT4) is a form of thyroid hormone used to treat hypothyroidism. In the brain, T4 is converted to the active form T3 by type 2 deiodinase (D2). Thus, it is intriguing that carriers of the Thr92Ala polymorphism in the D2 gene (DIO2) exhibit clinical improvement when liothyronine (LT3) is added to LT4 therapy. Here, we report that D2 is a cargo protein in ER Golgi intermediary compartment (ERGIC) vesicles, recycling between ER and Golgi. The Thr92-to-Ala substitution (Ala92-D2) caused ER stress and activated the unfolded protein response (UPR). Ala92-D2 accumulated in the trans-Golgi and generated less T3, which was restored by eliminating ER stress with the chemical chaperone 4-phenyl butyric acid (4-PBA). An Ala92-Dio2 polymorphism–carrying mouse exhibited UPR and hypothyroidism in distinct brain areas. The mouse refrained from physical activity, slept more, and required additional time to memorize objects. Enhancing T3 signaling in the brain with LT3 improved cognition, whereas restoring proteostasis with 4-PBA eliminated the Ala92-Dio2 phenotype. In contrast, primary hypothyroidism intensified the Ala92-Dio2 phenotype, with only partial response to LT4 therapy. Disruption of cellular proteostasis and reduced Ala92-D2 activity may explain the failure of LT4 therapy in carriers of Thr92Ala-DIO2.

Authors

Sungro Jo, Tatiana L. Fonseca, Barbara M. L. C. Bocco, Gustavo W. Fernandes, Elizabeth A. McAninch, Anaysa P. Bolin, Rodrigo R. Da Conceição, Joao Pedro Werneck-de-Castro, Daniele L. Ignacio, Péter Egri, Dorottya Németh, Csaba Fekete, Maria Martha Bernardi, Victoria D. Leitch, Naila S. Mannan, Katharine F. Curry, Natalie C. Butterfield, J.H. Duncan Bassett, Graham R. Williams, Balázs Gereben, Miriam O. Ribeiro, Antonio C. Bianco

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Figure 3

ERGIC53 and SCAP play a role in Ala92-D2 trafficking.

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ERGIC53 and SCAP play a role in Ala92-D2 trafficking. (A) Western blot o...
(A) Western blot of control HEK-293 and HEK-293-ERGIC–/– cells using the indicated antibodies. (B–D) Immunofluorescence of control HEK-293 cells transiently expressing Ala92-D2HY using the indicated antibodies; on the far right is the Pearson’s plot for each immunofluorescence image. The right top number is the Pearson’s coefficient for that specific cell. (E–G) Immunofluorescence of HEK-293-ERGIC–/– cells transiently expressing Ala92-D2HY; arrows point to Golgi AlaD2 staining. (H) Pearson’s coefficient between the indicated D2 proteins and cis-Golgi marker GM130. (I) FRET in HEK293 cells transiently expressing either D2T-EYFP (Thr) or D2A-EYFP (Ala) and SCAP constructs containing the YFP and CFP fluorophores at the indicated positions; cells were treated with HPβCD to cause cholesterol deprivation, as indicated. (J) Same as I, except that SCAP-ΔWD40 was used. (K–P) Immunofluorescence of Thr92-D2HY- or Ala92-D2HY-expressing cells, which were incubated overnight with 500 μM 4-PBA. (Q) Pearson’s coefficient between the indicated D2 proteins and trans-Golgi marker p230 as shown in K–P. (R–V, X) same as K–P, except the cells were incubated overnight with 10 μg/ml cholesterol. (W) Same as in Q, except that data are from R–V, X. Original magnification, APO ×60/1.40 oil objective. Values are shown in a box-and-whiskers plot indicating median and quartiles. n = 9–69/group. Statistical analysis used was the Mann-Whitney U test or the Kruskall-Wallis test, followed by Dunn’s multiple comparison test. *P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001.