Pathogenic variations inMAML2andMAMLD1contribute to congenital hypothyroidism due to dyshormonogenesis by regulating the Notch signalling pathway

Abstract

BackgroundIn several countries, thyroid dyshormonogenesis is more common than thyroid dysgenesis in patients with congenital hypothyroidism (CH). However, known pathogenic genes are limited to those directly involved in hormone biosynthesis. The aetiology and pathogenesis of thyroid dyshormonogenesis remain unknown in many patients.MethodsTo identify additional candidate pathogenetic genes, we performed next-generation sequencing in 538 patients with CH and then confirmed the functions of the identified genes in vitro using HEK293T and Nthy-ori 3.1 cells, and in vivo using zebrafish and mouse model organisms.ResultsWe identified one pathogenicMAML2variant and two pathogenicMAMLD1variants that downregulated canonical Notch signalling in three patients with CH. Zebrafish and mice treated with N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butylester, a γ-secretase inhibitor exhibited clinical manifestations of hypothyroidism and thyroid dyshormonogenesis. Through organoid culture of primary mouse thyroid cells and transcriptome sequencing, we demonstrated that Notch signalling within thyroid cells directly affects thyroid hormone biosynthesis rather than follicular formation. Additionally, these three variants blocked the expression of genes associated with thyroid hormone biosynthesis, which was restored byHES1expression. TheMAML2variant exerted a dominant-negative effect on both the canonical pathway and thyroid hormone biosynthesis.MAMLD1also regulated hormone biosynthesis through the expression ofHES3, the target gene of the non-canonical pathway.ConclusionsThis study identified three mastermind-like family gene variants in CH and revealed that both canonical and non-canonical Notch signalling affected thyroid hormone biosynthesis.