Site-directed mutational analysis of a U4 small nuclear RNA gene proximal sequence element. Localization and identification of functional nucleotides.

The genes that encode the small nuclear RNAs (snRNAs) are unusual RNA polymerase II transcription units in that 5'-flanking DNA sequences more than 50 base pairs upstream of snRNA genes are essential for specifying the transcription initiation site. The relevant cis-acting DNA sequence, termed the proximal sequence element (PSE), is required for both transcription initiation and 3'-end formation of snRNAs. We have used site-directed mutagenesis and expression in Xenopus oocytes to map nucleotides important for the function of the chicken U4B snRNA gene PSE. The results indicate that nucleotide sequences upstream of position -65 are not required for U4B PSE activity. However, nucleotides lying within a region 53-65 base pairs upstream of the U4B gene are essential for obtaining a detectable level of U4B gene expression. Six nucleotides between positions -53 and -59 were identified at which base substitutions reduced the transcriptional activity of the U4B gene.