UCSF

ABCG2 encodes for a multidrug efflux transporter called the mitoxantrone resistance protein (MXR, BCRP) that mediates the efflux of substrates out of the cell and is important in detoxification. The present study was focused on the expression and function of MXR amino acid variants, activity of the ABCG2 promoter and promoter variants, characterization of ABCG2 locus cis-regulatory elements and variant enhancers, and examination of DNA methylation around ABCG2. MXR expression, localization and activity were characterized using whole cells and inside-out vesicles. The Q141K variant had reduced expression. MXR I206L had increased efflux of pheophorbide A and both V12M and D620N had increased ATPase activity. The activity of ABCG2 regulatory elements was tested in in vitro and in vivo luciferase assays. Two promoter SNPs (rs76656413 and rs59370292) had decreased in vivo liver enhancer activity. Six regions with in vivo liver enhancer activity and several enhancer SNPs (rs9999111, rs12508471, rs72873421, rs149713212 and rs2725263) with altered activity in vivo were identified. Association of these SNPs with ABCG2, PPM1K or PDK2 expression in different tissues was detected. In vitro assays were used to identify nuclear receptor response elements. The ABCG2 promoter responded to multiple nuclear receptor ligands, and the promoter SNP rs66664036 had a significantly increased response to 17β-estradiol. Nine rifampin, six 17β-estradiol and three dexamethasone responsive regions were identified. Enhancer SNP rs12508471 had decreased response to 17β-estradiol and increased response to dexamethasone, while rs573519157 had an increased and rs190754327 had a decreased response to 17β-estradiol. Finally, methylation of CpG islands in the ABCG2 locus was correlated with the expression of ABCG2 in human liver and kidney tissues. There was no correlation of whole CpG island methylation with ABCG2 expression. However, a CpG site within CpG4 correlated with ABCG2 expression in the kidney, and part or all of select CpG islands had significantly lower methylation in liver than in kidney. The genetic and epigenetic regulation of the ABCG2 gene locus described in this dissertation may contribute to clinical variation in ABCG2 expression.