Characterization of a novel loss‐of‐function mutation of PAX8 associated with congenital hypothyroidism

Abstract

Summary Background Congenital hypothyroidism (CH) is a common endocrine disease that occurs in about 1:3000 newborns. In 80–85% of the cases, CH is presumably secondary to thyroid dysgenesis (TD), a defect in the organogenesis of the gland leading to an ectopic (30–45%), absent (agenesis, 35–40%) or hypoplastic (5%) thyroid gland. The pathogenesis of TD is still largely unknown. Most cases of TD are sporadic, although familial occurrences have occasionally been described. Recently, mutations in the PAX8 transcription factor have been identified in patients with TD. Objective Our aim was to identify and functionally characterize novel PAX8 mutations with autosomal dominant transmission responsible for TD. Design The PAX8 gene was sequenced in a mother and child both suffering from congenital hypothyroidism (CH) because of thyroid hypoplasia. Subsequently, expression vectors encoding the mutated PAX8 were generated, and the effects of the mutation on both the DNA‐binding capability and the transcriptional activity were evaluated. Results PAX8 gene sequencing revealed a heterozygous mutation that consists of the substitution of a histidine residue with a glutamine at position 55 of the PAX8 protein (H55Q). When tested in cotransfection experiments with a thyroglobulin promoter reporter construct, the mutant protein turned out to be still able to bind DNA in Electrophoretic Mobility Shift Assay assays but transcriptionally inactive. Conclusions Our findings confirm the important role of PAX8 in normal thyroid development and support the evidence that in humans haploinsufficiency of PAX8 is associated with TD.

Two Cases of Thyroid Dysgenesis Caused by Different Novel PAX8 Mutations in the DNA Binding Region: In vitro studies reveal different pathogenic mechanisms.

“…This mutation is located in a highly conserved area of the gene, which encodes the DNA-binding domain of the PAX8 protein. Other mutations in this area of the gene have been described before and have been characterized at the molecular level (4,5,7,(9)(10)(11)(12)(13)(14)(15)17). In agreement with previously reported mutations (R31C, R31H, Q40P, R52P, S54G, H55Q, C57Y, L62R), S54R has lost its ability to bind to TPO promoter-binding sites.…”

“…Furthermore, PAX8 and TTF1 synergistically activate the promoter of human TG (3). So far, 13 different mutations in the PAX8 gene have been reported (3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17), and the phenotypes of affected individuals vary considerably. Even within the same family, heterozygous individuals with PAX8 mutations can have no thyroid gland abnormality, have thyroid hypoplasia or TD (6), making it very difficult to identify individuals harboring a PAX8 gene mutation (1).…”

Functional characterization of four novel PAX8 mutations causing congenital hypothyroidism: new evidence for haploinsufficiency as a disease mechanism

“…The former type seems to be the predominant mechanism of PAX8 mutations: ten out of 12 previously reported mutations are amino acidaltering ones located in the paired domain (4,7,8,9,10,11,12,13). The paired domain consists of two b-turns (b1 and b2) and six a-helices (a1-a6).…”

“…In humans, a heterozygous PAX8 mutation causes congenital hypothyroidism (CH). To date, 32 mutation carriers belonging to 13 families have been described (4,5,6,7,8,9,10,11,12,13,14). Clinical phenotypes of mutation carriers are variable, ranging from overt CH with severe thyroid hypoplasia to subclinical CH with a morphologically normal gland.…”

Extreme phenotypic variability of thyroid dysgenesis in six new cases of congenital hypothyroidism due to PAX8 gene loss-of-function mutations

“…The PAX8 gene at 2q12-q14 is among the genes involved in thyroid gland development and, more specifically, in the proliferation and differentiation of thyroxine (T 4 )-producing follicular cells (4,5). PAX8 protein expression starts at the third gestational week in the human embryo and continues until adulthood (6,7,8,9,10). In thyrocytes, PAX8 contributes to proper timing of the switch from progenitor cell proliferation to the initiation of terminal differentiation programs marked by the expression of genes such as those encoding thyroglobulin, thyroperoxidase (TPO), and sodium/iodide symporter (8,11).…”