Mechanisms of thyrotropin receptor–mediated phenotype variability deciphered by gene mutations and M453T-knockin model
Kristiina Makkonen, Meeri Jännäri, Luís Crisóstomo, Matilda Kuusi, Konrad Patyra, Vladyslav Melnyk, Veli Linnossuo, Johanna Ojala, Rowmika Ravi, Christoffer Löf, Juho-Antti Mäkelä, Päivi Miettinen, Saila Laakso, Marja Ojaniemi, Jarmo Jääskeläinen, Markku Laakso, Filip Bossowski, Beata Sawicka, Karolina Stożek, Artur Bossowski, Gunnar Kleinau, Patrick Scheerer, FinnGen FinnGen, Mary Pat Reeve, Jukka Kero
Kristiina Makkonen, Meeri Jännäri, Luís Crisóstomo, Matilda Kuusi, Konrad Patyra, Vladyslav Melnyk, Veli Linnossuo, Johanna Ojala, Rowmika Ravi, Christoffer Löf, Juho-Antti Mäkelä, Päivi Miettinen, Saila Laakso, Marja Ojaniemi, Jarmo Jääskeläinen, Markku Laakso, Filip Bossowski, Beata Sawicka, Karolina Stożek, Artur Bossowski, Gunnar Kleinau, Patrick Scheerer, FinnGen FinnGen, Mary Pat Reeve, Jukka Kero
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Research Article Endocrinology Genetics

Mechanisms of thyrotropin receptor–mediated phenotype variability deciphered by gene mutations and M453T-knockin model

Abstract

The clinical spectrum of thyrotropin receptor–mediated (TSHR-mediated) diseases varies from loss-of-function mutations causing congenital hypothyroidism to constitutively active mutations (CAMs) leading to nonautoimmune hyperthyroidism (NAH). Variation at the TSHR locus has also been associated with altered lipid and bone metabolism and autoimmune thyroid diseases. However, the extrathyroidal roles of TSHR and the mechanisms underlying phenotypic variability among TSHR-mediated diseases remain unclear. Here we identified and characterized TSHR variants and factors involved in phenotypic variability in different patient cohorts, the FinnGen database, and a mouse model. TSHR CAMs were found in all 16 patients with NAH, with 1 CAM in an unexpected location in the extracellular leucine-rich repeat domain (p.S237N) and another in the transmembrane domain (p.I640V) in 2 families with distinct hyperthyroid phenotypes. In addition, screening of the FinnGen database revealed rare functional variants as well as distinct common noncoding TSHR SNPs significantly associated with thyroid phenotypes, but there was no other significant association between TSHR variants and more than 2,000 nonthyroid disease endpoints. Finally, our TSHR M453T–knockin model revealed that the phenotype was dependent on the mutation’s signaling properties and was ameliorated by increased iodine intake. In summary, our data show that TSHR-mediated disease risk can be modified by variants at the TSHR locus both inside and outside the coding region as well as by altered TSHR-signaling and dietary iodine, supporting the need for personalized treatment strategies.

Authors

Kristiina Makkonen, Meeri Jännäri, Luís Crisóstomo, Matilda Kuusi, Konrad Patyra, Vladyslav Melnyk, Veli Linnossuo, Johanna Ojala, Rowmika Ravi, Christoffer Löf, Juho-Antti Mäkelä, Päivi Miettinen, Saila Laakso, Marja Ojaniemi, Jarmo Jääskeläinen, Markku Laakso, Filip Bossowski, Beata Sawicka, Karolina Stożek, Artur Bossowski, Gunnar Kleinau, Patrick Scheerer, FinnGen FinnGen, Mary Pat Reeve, Jukka Kero

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Figure 4

Generation and characterization of the thyroid phenotype in knockin mice carrying a constitutively active TSHR M453T mutation.

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Generation and characterization of the thyroid phenotype in knockin mice...
(A) Illustration of the location of TSHR M453T mutation and its G protein signaling preferences compared with WT TSHR. Below is a representative genotyping chromatogram of WT, heterozygous (HET), and homozygous (HOM) TSHR M453T mice, from left to right. The generation of the model using CRISPR/Cas9 and homology-directed repair technique, genomic location, and designed guide RNA (gRNA) construct and genotyping protocol is described in Supplemental Figure 7. On the right, representative thyroid images of WT, HET, and HOM TSHR female mice. (B–D) Thyroid weight (B) and serum TSH and T4 concentrations of 1 and >6-month-old WT, HET, and HOM TSHR mutant mice under high-iodine (C) and sufficient-iodine (D) diets. n = 3–17 male and female mice per genotype. Box-and-whisker plot represents median, 25th, and 75th percentile, with whiskers from minimum to maximum with all points shown. Statistical analysis was carried out using the 1-way ANOVA with Bonferroni’s multiple-comparisons test. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.