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MAMMALIAN RRN3 IS NOT REQUIRED FOR “RECRUITMENT”, BUT IS ESSENTIAL FOR THE FORMATION OF A FUNCTIONAL PREINITIATION COMPLEX
Author(s) -
Cavanaugh Alice H.,
Evans Ann,
Rothblum Lawrence Ian
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.lb31-a
Subject(s) - transcription preinitiation complex , transcription factor ii f , transcription factor ii e , transcription factor ii d , general transcription factor , rna polymerase i , rna polymerase ii , transcription factor ii b , transcription (linguistics) , polymerase , biology , rna polymerase ii holoenzyme , microbiology and biotechnology , transcription factor ii a , rna polymerase , rna , dna , genetics , rna dependent rna polymerase , promoter , gene expression , gene , linguistics , philosophy
The human homologue of yeast Rrn3 is essential for ribosomal DNA (rDNA) transcription. However, its mechanism of action is unknown. There are significant differences in the regulation of the assembly of Rrn3 with pol I in yeast and mammalian cells, and it has been suggested that the preinitiation complex that forms in the absence of yeast Rrn3 might be different than the mammalian complex. Thus, we have begun to study the mechanism by which Rrn3 functions in mammalian cells. Our data indicate that mammalian Rrn3 becomes limiting as transcription proceeds and is inactivated during the transcription reaction. We also show that this inactivated Rrn3 not only dissociates from RNA polymerase I, but is not capable of forming a stable complex with RNA polymerase I. Using immobilized template transcription assays and a novel in vitro ChIP assay, we have also demonstrated that Rrn3 is not required for recruitment of RNA polymerase I to the committed temlate. However, that complex cannot be converted to an active complex and that complex is less stable in the presence of heparin. Our results indicate that Rrn3 functions stoichiometrically in rDNA transcription, that its ability to associate with RNA polymerase I is lost upon transcription, and that in the absence of functional Rrn3, RNA polymerase I can be recruited to the committed template, but forms a “stable”, inactive complex.