TOHO rSNP DB2

A regulatory SNP is a variations which affect the ability of a transcription factor to bind to DNA.


Completion of the human genome project led to the post-genome era. Next important task is to apply differences of genome information of each individual to the personalized medicine. Non-synonymous single nucleotide polymorphisms (SNPs) that result in amino acid changes in proteins were extensively studied. However examples of variation in regulatory sequences are poorly reported, although there are several examples of regulatory SNPs associated with disease susceptibility related to development and differentiation.

Here, we focused on the constitutive androstane receptor (CAR) which is a transcription factor that belongs to the nuclear receptor superfamily. CAR binds as a heterodimer with the retinoid X receptor (RXR) to CAR response elements (CAREs) and regulates the expression of various drug metabolizing enzymes and transporters.To identify CAR/RXR binding sites in the human genome, we performed a modified yeast one-hybrid system [J Recept Signal Transduct Res. 2010 Apr;30(2):88-105, Methods Mol Biol. 2013;977:125-36] . The human genomic fragments were recovered from positive yeast colonies by PCR and sequenced. A motif enrichment analysis revealed that the most frequent motif was a direct repeat (DR) of RGKTCA-like core sequence spaced by 4 bp. Next, we predicted 149 putative CAR/RXR binding sites from 414 unique clones, by searching for DRs, everted repeats (ERs) and inverted repeats (IRs) of the RGKTCA-like core motif. Based on gel mobility shift assays, the CAR/RXR heterodimer could directly interact with the 108 predicted sequences, which included not only classical CAREs but also a wide variety of arrangements. Furthermore, we identified 17 regulatory polymorphisms on the CAR/RXR-binding sites that may influence individual variation in the expression of CAR-regulated genes. These results provide insights into the molecular mechanisms underlying the physiological and pathological actions of CAR/RXR heterodimers.


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 Kazuyuki Yanai

Department of Biomolecular Science, Faculty of Science, Toho University, 2-2-1 Miyama, Funabashi, Chiba 274-8510, Japan


 

  This work was supported by JSPS KAKENHI Grant Number 25450137.