Related Experiment Videos
Functional significance of the thyrotropin receptor germline polymorphism D727E
Gerasimos P Sykiotis1, Susanne Neumann, Neoklis A Georgopoulos
1Department of Biochemistry, School of Medicine, University of Patras, 26110, Patras, Greece.
Abstract:
In a toxic thyroid adenoma we identified a novel somatic mutation that constitutively activates the thyrotropin receptor (TSHR). Two heterozygous point mutations at adjacent nucleotides led to a substitution of alanine with asparagine at codon 593 (A593N) in the fifth transmembrane helix of TSHR. This somatic mutation resided on the same TSHR allele with the germline polymorphism D727E. The functional characteristics of the single TSHR mutants A593N and D727E and of the double mutant A593N/D727E were studied in transiently transfected COS-7 cells. The TSHR mutants A593N and A593N/D727E constitutively activated the cAMP cascade, whereas the D727E mutant did not differ from the wild-type TSHR. Surprisingly, the double mutant's specific constitutive activity was 2.3-fold lower than the A593N mutant. Thus, the polymorphism significantly ameliorates G(alphas) protein activation in the presence of the gain-of-function mutation A593N, although it is functionally inert in the context of the wild-type TSHR.
Insights
A novel mutation in the thyrotropin receptor (TSHR) was found in toxic thyroid adenoma, causing constitutive activation. A co-occurring germline polymorphism surprisingly reduced this activation, impacting G-alpha protein signaling.
Area of Science:
- Endocrinology
- Molecular Biology
- Genetics
Background:
- Toxic thyroid adenomas are often caused by activating mutations in the thyrotropin receptor (TSHR).
- Understanding these mutations is key to diagnosing and treating hyperthyroidism.
Purpose of the Study:
- To identify and characterize novel somatic mutations in TSHR associated with toxic thyroid adenoma.
- To investigate the functional impact of a specific TSHR mutation (A593N) and its interaction with a germline polymorphism (D727E).
Main Methods:
- Somatic mutation identification in toxic thyroid adenoma tissue.
- Site-directed mutagenesis to create TSHR variants (A593N, D727E, A593N/D727E).
- Functional analysis of TSHR mutants in transiently transfected COS-7 cells, measuring cAMP cascade activation.
Main Results:
- A novel somatic mutation, A593N, was identified in TSHR, leading to constitutive activation of the cAMP cascade.
- The germline polymorphism D727E alone did not affect TSHR activity.
- The double mutant (A593N/D727E) showed reduced constitutive activity compared to A593N alone, indicating amelioration of G-alpha protein activation.
Conclusions:
- The A593N mutation is a gain-of-function mutation causing constitutive TSHR activation in toxic thyroid adenoma.
- The D727E polymorphism modulates the effect of the A593N mutation, reducing its signaling output.
- This interaction highlights the complex genetic factors influencing thyroid hormone regulation.