Sequence requirements for premature termination of transcription in the human c-myc gene

We have used the Xenopus oocyte injection system to investigate the sequence requirements of premature termination of transcription within the human c-myc gene. We show that in the oocyte, truncated RNAs are produced by RNA polymerase II with 5' ends at the P1 and P2 promoters and 3' ends at two T stretches (sites I and II) near the exon 1/intron 1 junction. The location of these 3' ends is consistent with the site of the block to c-myc transcription identified by nuclear runoff assays in human cells and confirmed in dissected nuclei of injected oocytes. Evidence is presented that transcriptional termination rather than RNA processing produces these short c-myc RNAs. Deletion analysis of site I reveals that sequences upstream of the T stretch determine the site of 3' end formation, and that the stretch of T's on the sense DNA strand is not required for termination. The sequences specifying termination reside within a 95 base region located -130 to -35 relative to the exon 1/intron 1 boundary. The termination activity of these sequences is orientation-dependent and functions downstream of the HSV-TK promoter.