Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
5589707 | Gene | 2017 | 13 Pages |
Abstract
Transcription of the ribosomal RNA genes (rDNA) by RNA polymerase I (Pol I) is a major control step for ribosome synthesis and is tightly linked to cellular growth. However, the question of whether this process is modulated primarily at the level of transcription initiation or elongation is controversial. Studies in markedly different cell types have identified either initiation or elongation as the major control point. In this study, we have re-examined this question in NIH3T3 fibroblasts using a combination of metabolic labeling of the 47S rRNA, chromatin immunoprecipitation analysis of Pol I and overexpression of the transcription initiation factor Rrn3. Acute manipulation of growth factor levels altered rRNA synthesis rates over 8-fold without changing Pol I loading onto the rDNA. In fact, robust changes in Pol I loading were only observed under conditions where inhibition of rDNA transcription was associated with chronic serum starvation or cell cycle arrest. Overexpression of the transcription initiation factor Rrn3 increased loading of Pol I on the rDNA but failed to enhance rRNA synthesis in either serum starved, serum treated or G0/G1 arrested cells. Together these data suggest that transcription elongation is rate limiting for rRNA synthesis. We propose that transcription initiation is required for rDNA transcription in response to cell cycle cues, whereas elongation controls the dynamic range of rRNA synthesis output in response to acute growth factor modulation.
Keywords
PI3KSelectivity factor 1rRNAUBFMFPRAPALY294002MifepristoneCHXECLS.E.M.ETsqRT-PCRPICDMEMFBSRDNAEXPDulbecco's modified Eagle's mediumRibosomal RNARNA polymerase Istandard deviationenhanced chemiluminescencestandard error of the meanRapamycinHourfetal bovine serumcycloheximideUpstream binding factorphosphatidylinositol-3-kinasequantitative real-time PCRPol Ipreinitiation complexribosomal RNA gene
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Authors
Sandy S Hung, Analia Lesmana, Abigail Peck, Rachel Lee, Elly Tchoubrieva, Katherine M Hannan, Jane Lin, Karen E Sheppard, Katarzyna Jastrzebski, Leonie M Quinn, Lawrence I Rothblum, Richard B Pearson, Ross D Hannan, Elaine Sanij,