Rare and common genetic variations in the Keap1/Nrf2 antioxidant response pathway impact thyroglobulin gene expression and circulating levels, respectively

Abstract

Nuclear factor, erythroid 2-like 2 (Nrf2) is a transcription factor that has been gaining attention in the field of pharmacology and especially in the chemoprevention of diseases such as cancer, metabolic and neurodegenerative diseases, etc. This is because natural compounds such as sulforaphane, which is found in broccoli sprout extracts, can activate Nrf2. The repertoire of the roles of Nrf2 is ever increasing; besides its traditional antioxidant and cytoprotective effects, Nrf2 can have other functions as a transcription factor. We have recently shown that Nrf2 directly regulates the expression of thyroglobulin (Tg), which is the most abundant thyroidal protein and the precursor of thyroid hormones. Two functional binding sites for Nrf2 (antioxidant response elements, AREs) were identified in the regulatory region of the TG gene. Interestingly, we then observed that one of these AREs harbors a rare single-nucleotide polymorphism (SNP). Also recently, we performed the first genome-wide association study (GWAS) for common SNPs that impact the circulating levels of Tg. Based on these investigations, we were triggered (i) to investigate whether common SNPs in the Nrf2 pathway correlate with circulating Tg levels; and (ii) to examine whether the rare SNP in one of the TG regulatory AREs may affect gene expression. To address the first question, we analyzed GWAS data from a general population and its two subpopulations, one with thyroid disease and/or abnormal thyroid function tests and the other without, in which circulating Tg levels had been measured. Statistically significant associations with Tg levels were observed in the genes encoding Nrf2 and Keap1, including, notably, a known functional SNP in the promoter of the gene encoding Nrf2. Regarding the rare SNP (rs778940395) in the proximal ARE of the TG enhancer, luciferase reporter gene expression studies in PCCL3 rat thyroid follicular cells showed that this SNP abrogated the basal and sulforaphane- or TSH-induced luciferase activity, behaving as a complete loss-of-function mutation. Thus, both rare and common genetic variation in the Keap1/Nrf2 pathway can impact TG expression and Tg circulating levels, respectively.

Keywords: Keap1; Nrf2; SNP; Thyroglobulin; Thyroid.