Mapping of transcription initiation and termination signals on Xenopus laevis ribosomal DNA

We have injected cloned derivatives of Xenopus laevis ribosomal genes into X. laevis oocyte nuclei and examined the resulting transcription complexes in the electron microscope. From this work we conclude that the promoter lies somewhere within a region between -320 nucleotides upstream and +113 nucleotides downstream from the site of transcription initiation. This assignment agrees with inferences based on sequence conservation. It further suggests that the duplicated initiation region sequences located further out in the spacer ("Bam islands") are not required for the normal high densities of RNA polymerase loading seen on ribosomal genes. Concerning termination, the cluster of four Ts that forms part of the HindIII restriction site at the 3' end of the gene appears to be part of the normal termination signal. Termination still occurs when only three Ts are present, but reduction to two Ts damages termination. Because clusters of three Ts appear at several sites within the gene, it is likely that sequences adjacent to the T cluster also are required for normal termination. In addition, we present evidence for a fail-safe termination site just upstream from the site of transcription initiation.