Starting and Stopping Sequences for the RNA Polymerase

Abstract

INTRODUCTION
The RNA polymerase transcribes DNA into RNA starting at sites called promoters. At these sites, the polymerase must recognize a DNA sequence, attach, open the DNA strands to gain access to the bases, and then initiate transcription. How can we identify the features that the polymerase detects? One approach, pioneered by Heinz Schaller and co-workers (Schaller, Gray and Herrmann 1975), is to isolate and sequence a DNA fragment protected by the RNA polymerase against digestion with pancreatic DNase. The very stable complexes of RNA polymerase and DNA that form at 37°C (and are even maintained in DNase) define a limited number of “binding sites” corresponding to initiation sites. The sequence of the protected DNA fragments from these complexes presumably contains those bases that make the critical contacts responsible for maintaining the polymerase-DNA interaction. A more basic approach, however, is to exploit the original genetic definition of a promoter, i.e., the locus of mutations that block or enhance transcription. Changes in mutant sequences should identify the essential base pairs.

PROMOTER SEQUENCES
Polymerase-protected Fragments from Promoters
Figure 1 shows the five RNA polymerase-protected fragments that have been sequenced to date. These protected fragments have common properties. They are about the same size, 41–44 base pairs long. In each case, the polymerase can initiate on the DNA contained in the DNase-resistant complex and synthesize a short RNA as it runs off the end of the DNA fragment. These RNAs are about 17–20 bases long and correspond to the 5′ initial...