Purified RNA polymerase III accurately and efficiently terminates transcription of 5S RNA genes

Abstract

RNA polymerase III was purified to about 90% homogeneity from ovarian tissue of Xenopus laevis. The enzyme accurately initiates and terminates transcription of 5S RNA synthesis in isolated nuclei, but not when naked 5S DNA is used as a template. A sensitive hybridization technique was used to demonstrate that the purified polymerase, even when supplemented with a transcription factor that binds specifically to the 5S RNA gene internal control region, is unable to initiate synthesis at the start site of the 5S RNA gene. However, the polymerase alone terminates transcription at precisely the same site that is recognized in vivo and in complete transcription extracts. The purified polymerase distinguishes between weak and strong terminator sequences with the same relative efficiency as the enzyme in complete extracts. We conclude that the pure enzyme can recognize the simple consensus sequence found at the end of genes transcribed by RNA polymerase III.