Abstract
The 137 ribosomal protein genes (RPGs) of Saccharomyces provide a model for gene coregulation. We examined the positional and functional organization of their regulators (Rap1 [repressor activator protein 1], Fhl1, Ifh1, Sfp1, and Hmo1), the transcription machinery (TFIIB, TFIID, and RNA polymerase II), and chromatin at near-base-pair resolution using ChIP-exo, as RPGs are coordinately reprogrammed. Where Hmo1 is enriched, Fhl1, Ifh1, Sfp1, and Hmo1 cross-linked broadly to promoter DNA in an RPG-specific manner and demarcated by general minor groove widening. Importantly, Hmo1 extended 20-50 base pairs (bp) downstream from Fhl1. Upon RPG repression, Fhl1 remained in place. Hmo1 dissociated, which was coupled to an upstream shift of the +1 nucleosome, as reflected by the Hmo1 extension and core promoter region. Fhl1 and Hmo1 may create two regulatable and positionally distinct barriers, against which chromatin remodelers position the +1 nucleosome into either an activating or a repressive state. Consistent with in vitro studies, we found that specific TFIID subunits, in addition to cross-linking at the core promoter, made precise cross-links at Rap1 sites, which we interpret to reflect native Rap1-TFIID interactions. Our findings suggest how sequence-specific DNA binding regulates nucleosome positioning and transcription complex assembly >300 bp away and how coregulation coevolved with coding sequences.
Highlights
Saccharomyces contains 78 distinct ribosomal proteins (RPs) encoded by 137 genes (RPGs): 19 single genes and 59 paralogous gene pairs
The strongest positional reinforcement of ribosomal protein genes (RPGs)-specific factor patterning across all RPGs occurred when aligned by their Rap1 sites rather than their transcriptional start site (TSS) (Fig. 1B, left vs. right graphs)
preinitiation complex (PIC) occupancy at RPGs that lacked Hmo1 was unaffected by Hmo1 loss, indicating that they are Hmo1-independent. These results demonstrate that the breadth of Hmo1 binding helps restrict PIC placement to RPG core promoters downstream from Rap1/FIS/Hmo1, as observed in single-gene tests (Kasahara et al 2008, 2011)
Summary
Saccharomyces contains 78 distinct ribosomal proteins (RPs) encoded by 137 genes (RPGs): 19 single genes and 59 paralogous gene pairs. Fhl interacts with Ifh to activate RPG transcription, which is stimulated by the TOR and PKA signaling pathways, Sirtuins, and Sfp (Jorgensen et al 2004; Marion et al 2004; Martin et al 2004; Schawalder et al 2004; Wade et al 2004; Rudra et al 2005; Hall et al 2006; Zhao et al 2006; Berger et al 2007; Xiao et al 2011; Downey et al 2013). Rap acts as a repressor instead of an activator, but its mechanism remains unclear
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