Myb-binding Protein 1a (Mybbp1a) Regulates Levels and Processing of Pre-ribosomal RNA

Attila Németh,Heinrich Leonhardt,Axel Imhof,Gernot Längst,Michael Hölzel,Dirk Eick,Rebecca Keough,Michaela Rohrmoser,Julia Hochstatter,Thomas J. Gonda,Lothar Schermelleh

doi:10.1074/jbc.m111.303719

Abstract

Ribosomal RNA gene transcription, co-transcriptional processing, and ribosome biogenesis are highly coordinated processes that are tightly regulated during cell growth. In this study we discovered that Mybbp1a is associated with both the RNA polymerase I complex and the ribosome biogenesis machinery. Using a reporter assay that uncouples transcription and RNA processing, we show that Mybbp1a represses rRNA gene transcription. In addition, overexpression of the protein reduces RNA polymerase I loading on endogenous rRNA genes as revealed by chromatin immunoprecipitation experiments. Accordingly, depletion of Mybbp1a results in an accumulation of the rRNA precursor in vivo but surprisingly also causes growth arrest of the cells. This effect can be explained by the observation that the modulation of Mybbp1a protein levels results in defects in pre-rRNA processing within the cell. Therefore, the protein may play a dual role in the rRNA metabolism, potentially linking and coordinating ribosomal DNA transcription and pre-rRNA processing to allow for the efficient synthesis of ribosomes.

Highlights

RRNA gene transcription and processing have to be coordinated, as they are intimately linked
RNA Polymerase I Is Associated with Myb-binding protein 1a (Mybbp1a)—To reveal potential regulators that act at the level of transcription initiation and elongation, we generated a murine MB III cell line stably expressing FLAG-tagged RPA116 (FLAG-RPA116MBIII), the second-largest subunit of RNA Pol I [30]
We identified Mybbp1a association with RNA Pol I and demonstrated that Mybbp1a serves both as a negative regulator of rRNA gene transcription and as a functional part of the ribosome biogenesis machinery

Summary

Coordination of rRNA Gene Transcription and Processing

RNA and associated Utps (U three proteins) increased in size with transcript length, indicating that rRNA processing, RNA folding, RNA modification, and ribosomal protein assembly occur co-transcriptionally [12,13,14,15]. Erk-dependent phosphorylation of UBF directly influences RNA Pol I elongation rates by inducing an UBF-dependent rearrangement of the chromatin environment. This effect could serve as a mechanism to coordinate transcription and the assembly of pre-ribosomal complexes on nascent rRNA by modulating the Pol I elongation rate. The 160-kDa Mybbp1a protein had originally been described as an interacting partner of the proto-oncogene c-Myb and since it has been shown to interact with and to modulate the activity of several regulators of Pol II-dependent transcription. The data shown here demonstrate two new functions for Mybbp1a in the nucleolus It associates with the RNA Pol I machinery and is able to repress its transcriptional activity

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION