SINGLE-STRANDED DNA-BINDING PROTEINS AND NEURON-RESTRICTIVE SILENCER FACTOR PARTICIPATE IN CELL-SPECIFIC TRANSCRIPTIONAL CONTROL OF THE NMDAR1 GENE

Our previous studies revealed that a proximal region of the N-methyl-D -aspartate receptor I (NMDAR1) promoter is important for cell-type-spe cific expression. We have now explored the contributions of several re gulatory elements to this specificity, Deletion of the neuron restrict ive silencer element partially relieved the suppression of promoter ac tivity in C6 glioma and HeLa cells. An overlapping G(C/G)G/tandem Sp1- containing region crucial for both basal and nerve growth factor (NGF) -regulated promoter activity specifically bound nuclear proteins on it s purine-rich sense strand. A faster migrating complex, single-strande d binding protein complex 1 (SBPC1), was highly enriched in HeLa cells , whereas a slower migrating complex, SBPC2, was enriched in PC12 cell s. A high ratio of 2/1 complex correlated with a high level of promote r activity. NGF treatment of PC12 cells reduced SBPC1 but increased SB PC2. Competition experiments showed that the SBPC1 binding required a dG(4) sequence and the SBPC2 needed a core of TG(3)A plus a 5'-flankin g sequence. Single-stranded DNA encompassing TG(3)A and/or dG(4) speci fically suppressed cotransfected NMDAR1 promoter activity. UV cross-li nking studies indicated that a 31.5-kDa protein mainly formed SBPC1, w hereas SBPC1 contained several larger proteins, Our results suggest th at neuron-restrictive silencer factor and single-stranded DNA-binding proteins may both play a role in cell-type specificity of the NMDAR1 g ene, and the latter may also be involved in basal and NGF-regulated ac tivity.