Transcription of spacer sequences in genes coding for ribosomal RNA in Xenopus cells.

Abstract

Untreated Xenopus cells synthesize RNA molecules (SS-23S) containing sequences complementary to the external "nontranscribed" spacer of the genes coding for rRNA. In cells incubated in the col maturation cleavages of the rRNA precursors are inhibited. Concomitantly, the relative proportion of spacer transcripts detectable in newly synthesized RNA is increased and some sequences complementary to those of the nontranscribed spacer are found in the heavy shoulder of the 40S precursor rRNA peak. In such events transcription initiation seems to take place within the spacer because its middle and right BamHI endonuclease fragments are preferentially transcribed, whereas only few RNA sequences complementary to the left spacer fragment are found. It is concluded that at least some spacer regions contain promoters for transcription and can be transcribed either into a special class of "spacer transcripts" or into molecules covalently linked to rRNA precursor.It is still unclear which sequences of eukaryotic genes coding for pre-rRNA or proteins are actually transcribed and whether transcription initiation starts only from a single defined initiation site. The rRNA genes (rDNA) of Xenopus, which are located in the nucleolus, are among the best-studied eukaryotic genes (for reviews see refs. 1-4). Their first stable transcript is a rRNA molecule (pre-rRNA) of Mr 2.6 X 106 and a sedimentation coefficient of 40 S. After prolonged labeling times no molecules larger than 40S have been, found hybridizing to rDNA (5). However, no 5'-terminal polyphosphates were detected on the 40S pre-rRNA fron cultured Xenopus cells (6). On the other hand, it has recently been shown that the 40S pre-rRNA isolated from oocytes may be capped by an enzyme preparation from vaccinia virus, and thus seems to carry a polyphosphate at its 5' end (7). These results do not, however, exclude other initiation events elsewhere in rDNA, especially if the resulting transcripts are highly unstable.There are various observations in the literature (for reviews see refs. 4, 8, and 9) indicating that the first stable pre-rRNA is not necessarily the only transcript of rDNA. In the electron microscope, the spacer intercepts between the 40S matrix units of Xenopus and Triturus are often found associated with isolated lateral fibrils or even with dense transcriptional complexes forming the so-called prelude regions (10, 11). In isolated nuclei of Rana, methylated, G+C-rich RNA molecules larger than the 40S pre-rRNA have been observed (12). Moreover, Xenopus cells grown in culture treated with fluorouridine synthesize large RNA molecules that seem to contain sequences of both the 40S pre-rRNA and the external "nontranscribed" spacer (ENTS) of rDNA (13). Accordingly, changes in the ultrastructure of rDNA transcription units such as the presence of transcriptional complexes along nearly the entire deoxynucleoprotein axis have been observed after fluorouridine treatment (14).In the present study we ,uCi/ml (1 Ci = 3.7 X 1010 becquerels). Incubation was for eith...

Additional RNA polymerase I initiation site within the nontranscribed spacer region of the rat rRNA gene.

“…The Bam islands represent imperfect copies of the promoter (nucleotides -145 to +4) (1); and the 60/81-bp repeats are homologous to the sequences between -72 and -114 of the promoter. It is clear from electron microscopic (31) and hybridization studies (33,36) that the Bam islands are capable of directing initiation, albeit at a very low efficiency. Maximally efficient utilization of the downstream promoter occurs when upstream elements are colinear with it.…”

Transcription of spacer sequences flanking the rat 45S ribosomal DNA gene.

“…Although spacer transcription 5' to the precursor rRNA promoter has been documented in Xenopus spp. (18,34,35,36,40) and D. melanogaster (14,26,32), the analogous spacer regions in mammals have thought to be transcriptionally inert. Wood et al (50) were unable to detect transcription of the analogous mouse spacer region containing the internally repetitious DNA (-1894 to -169).…”

“…Serial repeats of a 240-bp element in Drosophila melanogaster also contain sequence homology to the pol I promoter and can support in vitro transcription (14,26). Low-level transcription from these regions in both frogs (18,34,36,40) and D. melanogaster (32) can be detected in vivo.…”

“…Serial repeats of a 240-bp element in Drosophila melanogaster also contain sequence homology to the pol I promoter and can support in vitro transcription (14,26). Low-level transcription from these regions in both frogs (18,34,36,40) Moss (35) has suggested that transcription in the Xenopus NTS is functionally related to precursor rRNA synthesis. In his model, ribosomal spacer serves as a loading site for pol I molecules that are then delivered to the rDNA gene promoter by the process of transcription.…”

Inhibitor of ribosomal RNA synthesis in Xenopus laevis embryos. VII. Inhibition of 40S pre-rRNA synthesis in Xenopus neurula cells.

“…1 and 2 reveals that the effect of blastula PCA-extract differed markedly from that of tubercidin; the former inhibited the labeling of 40S pre-rRNA, whereas the latter enhanced the accumulation of 40S pre-rRNA. For Xenopus, 40S pre-rRNA is accepted to be the primary transcript (2,4,7,8,16) although there have been some reports suggesting the existence of a larger precursor (1,11). Thus, there is the implication that the PCA-extract suppresses formation of the primary transcript.…”