Elevated expression of the c-myc oncogene is a frequent finding in tumors and cell lines derived from carcinomas of the colon and rectum. In a previous study we demonstrated that the differentiation agent sodium butyrate causes a rapid reduction in the expression of c-myc RNA in the rectal carcinoma cell line SW837. This effect was blocked by inhibitors of protein synthesis, suggesting that butyrate causes the induction of an activity that has a negative effect on c-myc expression. In the present work we demonstrate that the rapid decrease in the level of c-myc RNA, upon treatment of SW837 cells with 2 mM butyrate, is followed by a slower decrease in the level of p53 RNA and an increase in the RNA levels for fibronectin and a placental type alkaline phosphatase. Using in vitro elongation of nascent transcripts to measure transcription and actinomycin D chase experiments to measure RNA stability, we show that the reduction in expression of c-myc RNA is due to an increase in the block to transcriptional elongation, rather than a decrease in transcriptional initiation or an increase in degradation of the RNA. We conclude that sodium butyrate induces an activity that increases the transcriptional block in SW837 cells, and that regulation of transcriptional elongation is an important mechanism for regulating c-myc expression in this cell type. A shift in relative usage of the two major promoters in the c-myc gene accompanies the reduction in expression. The potential significance of this finding with respect to transcriptional elongation is discussed. Mutations in the exon 1/intron 1 boundary region of the c-myc gene cause an increase in transcriptional elongation in Burkitt lymphoma. We sequenced this region in a series of cell lines derived from colorectal carcinomas, all of which had an elevated level of c-myc expression, to determine if a similar mutational mechanism is at work in this disease. All of the lines examined had a normal c-myc DNA sequence, suggesting that the deregulation of c-myc expression in colon cancer is not due to a cis mutation in this region.