Abstract

This work identifies central components of a feedback mechanism for the expression of mouse poly(ADP-ribose) polymerase-1 (PARP-1). Using the stress-induced duplex destabilization algorithm, multiple base-unpairing regions (BURs) could be localized in the 5' region of the mouse PARP-1 gene (muPARP-1). Some of these could be identified as scaffold/matrix-attachment regions (S/MARs), suggesting an S/MAR-mediated transcriptional regulation. PARP-1 binding to the most proximal element, S/MAR 1, and to three consensus motifs, AGGCC, in its own promoter (basepairs -956 to +100), could be traced by electrophoretic mobility-shift assay. The AGGCC-complementary GGCCT motif was detected by cis-diammine-dichloro platinum cross-linking and functionally characterized by the effects of site-directed mutagenesis on its performance in wild type (PARP-1(+/+)) and PARP-1 knockout cells (PARP-1(-/-)). Mutation of the central AGGCC tract at basepairs -554 to -550 prevented PARP-1/promoter interactions, whereby muPARP-1 expression became up-regulated. Transfection of a series of reporter gene constructs with or without S/MAR 1 (basepairs -1523 to -1007) and the more distant S/MAR 2 (basepairs -8373 to -6880), into PARP-1(+/+) as well as PARP-1(-/-) cells, revealed an additional, major level of muPARP-1 promoter down-regulation, triggered by PARP-1 binding to S/MAR 1. We conclude that S/MAR 1 represents an upstream control element that acts in conjunction with the muPARP-1 promoter. These interactions are part of a negative autoregulatory loop.