A 37-base pair element in the far upstream spacer region can enhance transcription of rat rDNA in vitro and can bind to the core promoter-binding factor(s).

Previous studies in this laboratory have demonstrated that a 174-base pair (bp) rat rDNA spacer region located more than 2 kilobase pairs upstream of the initiation site, can enhance rat rDNA transcription in vitro independent of its orientation or distance or when inserted downstream of the initiation site. Further dissection of this region showed that transcription of a rDNA fragment containing just 37 bp of the spacer sequence, located between -2.183 and -2.219 kilobase pairs upstream of the initiation site, is 8-fold greater than that of the rDNA fragment devoid of the spacer element. Electrophoretic mobility shift assay demonstrated specific interaction of the 37-bp DNA fragment with a cellular protein(s). The spacer DNA competed for essential transcription factors as demonstrated by the absence of transcription following preincubation of the extract with the 37-bp fragment. Similar competition was also observed when a 58-bp PolI promoter was substituted for the enhancer fragment. The binding of the factor(s) to the enhancer element was not altered when coding and noncoding strands of the 37-bp oligodeoxynucleotide were used separately in the competition assay. Since the 37-bp enhancer region and the core promoter do not exhibit any significant sequence homology, the factor(s) appears to interact with these cis-acting elements in a sequence-independent manner.