Inhibition of RNA polymerase III transcription by a ribosome-associated kinase activity.
Ribosomes prepared from somatic tissue of Xenopus laevis inhibit transcription by RNA polymerase III. This observation parallels an earlier report that a high speed fraction from activated egg extract, which is enrichedin ribosomes, inhibits RNA polymerase III activityand destabilizes putative transcription complexes assembled on oocyte 5S rRNA genes. Transcription of somatic- and oocyte-type 5S rRNA genes and a tRNA gene are all repressed in the present experiments. We find that 5S rRNA genes incubated in S150 extract prepared from immature oocytes exhibit an extensive DNase I protection pattern that is nearly identical to that of the ternary complex of TFIIIA and TFIIIC bound to a somatic 5S rRNA gene. The complexes formed in this extract are stable at concentrations of ribosomes that completely repress transcription, indicating that formation of the TFIII(A+C) complex is not the target of inhibition. Ribosomes taken through a high salt treatment no longer repress transcription of class III genes, establishing that the inhibition is due to an associated factor and not the particle itself. The inhibitory activity released from ribosomes is inactivated by treatment with proteinase K, but not micrococcal nuclease. Preincubation of ribosomes with a general protein kinase inhibitor, 6-dimethylaminopurine, eliminates repression of transcription. Western blot analysis demonstrates that p34(cdc2), which is known to mediate repression of transcription by RNA polymerase III, is present in these preparations of ribosomes and can be released from the particles upon extraction with high salt. These results establish that a kinase activity, possibly p34(cdc2), is the actual agent responsible for the observed inhibition of transcription by ribosomes.
Westmark
C J
CJ
Department of Chemistry and Biochemistry, University of Notre Dame, 251 Nieuwland Science Hall, Notre Dame,IN 46556-5670, USA.
Ghose
R
R
Huber
P W
PW
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002457:Cell Extracts; D018983:DNA Footprinting; D019286:Endopeptidase K / Q000037:antagonists & inhibitors / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D020459:Genes, rRNA; D008836:Micrococcal Nuclease / Q000037:antagonists & inhibitors / Q000494:pharmacology; D009865:Oocytes / Q000201:enzymology; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism*; D012320:RNA Polymerase III / Q000037:antagonists & inhibitors* / Q000378:metabolism; D012341:RNA, Ribosomal, 5S / Q000235:genetics; D012343:RNA, Transfer / Q000235:genetics; D012270:Ribosomes / Q000201:enzymology / Q000378:metabolism*; D012492:Salts / Q000494:pharmacology; D019474:Transcription Factors, TFIII / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014982:Xenopus laevis
D002457:Cell Extracts; D004791:Enzyme Inhibitors; D047428:Protein Kinase Inhibitors; D012341:RNA, Ribosomal, 5S; D012492:Salts; D019474:Transcription Factors, TFIII; C001020:N(6),N(6)-dimethyladenine; D012343:RNA, Transfer; D011494:Protein Kinases; D012320:RNA Polymerase III; D008836:Micrococcal Nuclease; D019286:Endopeptidase K; D000225:Adenine