TRANSLOCATION AND TRANSCRIPTIONAL ARREST DURING TRANSCRIPT ELONGATIONBY RNA-POLYMERASE-II

RNA polymerase II may stop transcription, or arrest, while transcribin g certain DNA sequences. The molecular basis for arrest is not well un derstood, but a connection has been suggested between arrest and a tra nsient failure of the polymerase to translocate along the template. We have investigated this question by monitoring the movement of RNA pol ymerase II along a number of templates, using exonuclease III protecti on as our assay. We found that normal transcription is accompanied by essentially coordinate movement of the active site and both the leadin g and trailing edges of the polymerase. However, as polymerase approac hes an arrest site, translocation of the body of the polymerase stops while transcription continues, leading to an arrested complex in which the 3' end of the transcript is located much closer than normal to th e front edge of the polymerase. Surprisingly, mutated arrest sites tha t no longer block transcription continue to direct the transient failu re of polymerase translocation. As transcription proceeds through thes e sequences, the initially stationary polymerase moves forward 10-15 b ases along the template in response to the addition of only 3 bases to the nascent RNA. Mutagenesis studies indicate that the sequences resp onsible for the transient block to polymerase movement are located dow nstream of the T-rich motif required for arrest, Our results indicate that blocking translocation is not sufficient to cause arrest.