Novel mechanism of regulation of the DNA repair enzyme OGG1 in tuberin-deficient cells

Abstract

Tuberin (protein encodes by tuberous sclerosis complex 2, Tsc2) deficiency is associated with the decrease in the DNA repair enzyme 8-oxoG-DNA glycosylase (OGG1) in tumour kidney of tuberous sclerosis complex (TSC) patients. The purpose of this study was to elucidate the mechanisms by which tuberin regulates OGG1. The partial deficiency in tuberin expression that occurs in the renal proximal tubular cells and kidney cortex of the Eker rat is associated with decreased activator protein 4 (AP4) and OGG1 expression. A complete deficiency in tuberin is associated with loss of AP4 and OGG1 expression in kidney tumour from Eker rats and the accumulation of significant levels of 8-oxo-deoxyguanosine. Knockdown of tuberin expression in human renal epithelial cells (HEK293) with small interfering RNA (siRNA) also resulted in a marked decrease in the expression of AP4 and OGG1. In contrast, overexpression of tuberin in HEK293 cells increased the expression of AP4 and OGG1 proteins. Downregulation of AP4 expression using siRNA resulted in a significant decrease in the protein expression of OGG1. Immunoprecipitation studies show that AP4 is associated with tuberin in cells. Gel shift analysis and chromatin immunoprecipitation identified the transcription factor AP4 as a positive regulator of the OGG1 promoter. AP4 DNA-binding activity is significantly reduced in Tsc2(-/-) as compared with Tsc2(+/+) cells. Transcriptional activity of the OGG1 promoter is also decreased in tuberin-null cells compared with wild-type cells. These data indicate a novel role for tuberin in the regulation of OGG1 through the transcription factor AP4. This regulation may be important in the pathogenesis of kidney tumours in patients with TSC disease.