Mitotic silencing of human rRNA synthesis: inactivation of the promoter selectivity factor SL1 by cdc2/cyclin B-mediated phosphorylation

Abstract

We have used a reconstituted cell-free transcription system to investigate the molecular basis of mitotic repression of RNA polymerase I (pol I) transcription. We demonstrate that SL1, the TBP-containing promoter-binding factor, is inactivated by cdc2/cyclin B-directed phosphorylation, and reactivated by dephosphorylation. Transcriptional inactivation in vitro is accompanied by phosphorylation of two subunits, e.g. TBP and hTAF I 110. To distinguish whether transcriptional repression is due to phosphorylation of TBP, hTAF I 110 or both, SL1 was purified from two HeLa cell lines that express either full-length or the core domain of TBP only. Both TBP-TAF I complexes exhibit similar activity and both are repressed at mitosis, indicating that the variable N-terminal domain which contains multiple target sites for cdc2/cyclin B phosphorylation is dispensable for mitotic repression. Protein-protein interaction studies reveal that mitotic phosphorylation impairs the interaction of SL1 with UBF. The results suggest that phosphorylation of SL1 is used as a molecular switch to prevent pre-initiation complex formation and to shut down rDNA transcription at mitosis.

Multiple interactions between RNA polymerase I, TIF‐IA and TAF I subunits regulate preinitiation complex assembly at the ribosomal gene promoter

“…Previous studies revealed that the shutdown of rRNA synthesis during mitosis and early G 1 phase is due to inactivation of TIF-IB/SL1 and UBF (Heix et al, 1998;Klein and Grummt, 1999). Growth-dependent regulation of Pol I, on the other hand, has been attributed to alterations in the amount or activity of TIF-IA (Buttgereit et al, 1985;Schnapp et al, 1993).…”

Rio1 promotes rDNA stability and downregulates RNA polymerase I to ensure rDNA segregation

“…In prophase, these nucleoli disassemble and their transcription becomes repressed to allow the stripped rDNA to segregate with the rest of the genome. Nucleolar transcription is downregulated by the CDK1-cyclin B kinase, whose phosphorylation of promoter selectivity factor SL1 prevents the formation of the PolI pre-initiation complex [63][64][65] . Similar to yeast Rio1, RIOK1 might just as well contribute to the transcriptional repression of PolI.…”

Dynamic epigenetic states of ribosomal RNA promoters during the cell cycle

“…The transcription termination factor 1 (TTF-1) and SL-1 are phosphorylated by cdk2/Cyclin B at the G 2 /M transition. 2,48 The phosphorylation of SL-1 prevents its association with UBF which inhibits rRNA transcription, despite the fact that all factors remain associated with the chromatin. 49 UBF is also phosphorylated and inactivated in mitosis.…”

“…Thus, this process must be tightly regulated to ensure that the proper levels of transcription are congruent with the cellular requirements for protein production. The rate of rRNA transcription and elongation can be modulated by several factors, including growth factors, 1 cell cycle, 2 and cellular stress. 3 RNA polymerase I (Pol I) is the polymerase dedicated to transcribing the 47S pre-rRNA transcript.…”

“…During mitosis transcription is inhibited through the phosphorylation of the TAF 1 110 subunit of SL-1 by cdk1/cyclin B. 2 While the activity of SL-1 is restored early in G 1 , rRNA transcription remains low due to the hypophosphorylation of UBF. During G 1 , UBF becomes phosphorylated by cdk4/cyclin D1 and cdk2/cyclin E&A, 6,7 which stimulates its binding to Pol I to promote rRNA transcription.…”