Introduction. Congenital hypothyroidism (CH) is the most common congenital endocrine disease, resulting from a partial or complete loss of thyroid function. To date, the molecular mechanisms underlying CH pathogenesis remain largely unknown. This is partly due to limited knowledge regarding the cross-talk of intrinsic (thyroid-specific genes) and extrinsic factors (growth factors, morphogens) in the regulation of normal thyroid organogenesis. Thyroid development and function appear highly conserved in vertebrates; similar to mammals, the zebrafish thyroid gland is composed of endoderm- derived follicles, filled with colloid and producing thyroid hormones. A better comprehension of the precise role of different pathways during thyroid development is expected to offer novel working hypotheses on the molecular and genetic basis of human CH. Methods. To investigate the pathways involved in thyroid formation, we have generated transgenic zebrafish lines expressing in vivo mCherry or GFP under the control of a thyroglobulin (TG) promoter, using the Multisite Gateway-based Tol2 technology. Then, we have outcrossed these transgenic thyroid lines with signaling pathway reporter lines of potential interest for thyroid formation, physiology and pathogenesis (Bmp, Shh, Fgf, Wnt, Notch, cAMP/CREB, Hypoxia, Stat3, Glucocorticoid). Results. We have found that cAMP/CREB and TGFbeta signaling pathways are activated in TG-expressing cells. In particular, TGFbeta signaling is activated during early stages of thyroid formation. The cAMP/CREB reporter signal is activated in the thyroid follicles after 3 dpf, following an antero- posterior gradient, with higher expression in cells with lower levels of thyroglobulin. This suggests a specific role for CREB signaling in the initial differentiation of the gland. Other investigated pathways did not shown co-localization with TG--positive cells. However, Bmp, Notch and Wnt signals appear activated in cells surrounding the gland. Conclusions. Double transgenic pathway/thyroid-specific lines point to cAMP/CREB and TGFbeta as the main signaling pathways involved during thyroid formation in zebrafish. We are now planning to perform a chemical screen in order to verify the role of candidate signaling pathways during normal thyroid development. This work is supported by Italian Ministry of Health (RF-2010-2309484) and EU ZF-HEALTH. B.G. is a UniPD Junior Post-doc Fellow (CPDR124317/12).

CAMP/CREB and TGFbeta signaling pathways are activated during zebrafish thyroid development

BUSOLIN, GIORGIA;EK, OLIVIER FREDERIC D;VETTORI, ANDREA;SCHIAVONE, MARCO;FACCHINELLO, NICOLA;CASARI, ALESSANDRO;ARGENTON, FRANCESCO;TISO, NATASCIA
2015

Abstract

Introduction. Congenital hypothyroidism (CH) is the most common congenital endocrine disease, resulting from a partial or complete loss of thyroid function. To date, the molecular mechanisms underlying CH pathogenesis remain largely unknown. This is partly due to limited knowledge regarding the cross-talk of intrinsic (thyroid-specific genes) and extrinsic factors (growth factors, morphogens) in the regulation of normal thyroid organogenesis. Thyroid development and function appear highly conserved in vertebrates; similar to mammals, the zebrafish thyroid gland is composed of endoderm- derived follicles, filled with colloid and producing thyroid hormones. A better comprehension of the precise role of different pathways during thyroid development is expected to offer novel working hypotheses on the molecular and genetic basis of human CH. Methods. To investigate the pathways involved in thyroid formation, we have generated transgenic zebrafish lines expressing in vivo mCherry or GFP under the control of a thyroglobulin (TG) promoter, using the Multisite Gateway-based Tol2 technology. Then, we have outcrossed these transgenic thyroid lines with signaling pathway reporter lines of potential interest for thyroid formation, physiology and pathogenesis (Bmp, Shh, Fgf, Wnt, Notch, cAMP/CREB, Hypoxia, Stat3, Glucocorticoid). Results. We have found that cAMP/CREB and TGFbeta signaling pathways are activated in TG-expressing cells. In particular, TGFbeta signaling is activated during early stages of thyroid formation. The cAMP/CREB reporter signal is activated in the thyroid follicles after 3 dpf, following an antero- posterior gradient, with higher expression in cells with lower levels of thyroglobulin. This suggests a specific role for CREB signaling in the initial differentiation of the gland. Other investigated pathways did not shown co-localization with TG--positive cells. However, Bmp, Notch and Wnt signals appear activated in cells surrounding the gland. Conclusions. Double transgenic pathway/thyroid-specific lines point to cAMP/CREB and TGFbeta as the main signaling pathways involved during thyroid formation in zebrafish. We are now planning to perform a chemical screen in order to verify the role of candidate signaling pathways during normal thyroid development. This work is supported by Italian Ministry of Health (RF-2010-2309484) and EU ZF-HEALTH. B.G. is a UniPD Junior Post-doc Fellow (CPDR124317/12).
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11577/3194976
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