The SAGA coactivator complex acts on the whole transcribed genome and is required for RNA polymerase II transcription

Abstract

The SAGA (Spt-Ada-Gcn5 acetyltransferase) coactivator complex contains distinct chromatin-modifying activities and is recruited by DNA-bound activators to regulate the expression of a subset of genes. Surprisingly, recent studies revealed little overlap between genome-wide SAGA-binding profiles and changes in gene expression upon depletion of subunits of the complex. As indicators of SAGA recruitment on chromatin, we monitored in yeast and human cells the genome-wide distribution of histone H3K9 acetylation and H2B ubiquitination, which are respectively deposited or removed by SAGA. Changes in these modifications after inactivation of the corresponding enzyme revealed that SAGA acetylates the promoters and deubiquitinates the transcribed region of all expressed genes. In agreement with this broad distribution, we show that SAGA plays a critical role for RNA polymerase II recruitment at all expressed genes. In addition, through quantification of newly synthesized RNA, we demonstrated that SAGA inactivation induced a strong decrease of mRNA synthesis at all tested genes. Analysis of the SAGA deubiquitination activity further revealed that SAGA acts on the whole transcribed genome in a very fast manner, indicating a highly dynamic association of the complex with chromatin. Thus, our study uncovers a new function for SAGA as a bone fide cofactor for all RNA polymerase II transcription.

Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription

“…Our results suggest that in the case of complexes, which interact transiently with chromatin, monitoring their genomic sites of action by ChIP may not reveal their complete functional range. In addition, the rapid fluorescence recovery/loss described in FRAP/FLIP experiments is consistent with our previous observation that the genome‐wide DUB activity of SAGA can be performed in < 10 min and in the absence of active transcription (Bonnet et al , 2014). Only complexes with high mobility and transient interactions with their substrate could act globally in such a short time.…”

“…Consequently, we conclude that the recruitment and function of the complexes on their chromatin substrate do not depend on direct and stable interactions with transcribing Pol II. This finding is of particular interest for the SAGA complex, which is known to exhibit its DUB activity on the gene body of actively transcribed genes (Bonnet et al , 2014). Our observations suggest that the removal of the ubiquitin moiety from H2Bub1 does not require SAGA to travel along the gene body together with elongating Pol II.…”

“…Those findings together with the fact that in yeast there was only a very weak overlap between SAGA‐bound and SAGA‐regulated genes, when analysed at the steady state mRNA level, raised questions about the detectability of these coactivators by ChIP (Lenstra & Holstege, 2012 and refs therein). Indeed, when the genome‐wide distribution of the SAGA co‐activator complex was tracked by following its two chromatin modifying activities (H3K9 acetylation or H2Bub1 deubiquitination), it was discovered that SAGA acetylates the promoters, and deubiquitinates the transcribed regions, of all expressed genes both in S. cerevisiae and in human cells (Bonnet et al , 2014). The latter study provided data suggesting that SAGA DUB interacts with chromatin in a Pol II transcription‐independent manner.…”

“…Studies in different eukaryotes have suggested that the presence of SAGA at the promoters of genes is needed to facilitate Pol II recruitment and pre‐initiation complex (PIC) formation (Wyce et al , 2007; Nagy et al , 2009; Helmlinger et al , 2011; Lang et al , 2011). The recruitment of SAGA and ATAC coactivator complexes to genomic loci has been suggested to take place by several distinct mechanisms: (i) by activator mediated recruitment (McMahon et al , 1998; Brown et al , 2001; Fishburn et al , 2005; Reeves & Hahn, 2005), (ii) by interactions with basal transcription machinery components (Larschan & Winston, 2001; Laprade et al , 2007; Mohibullah & Hahn, 2008) and (iii) through chromatin‐interacting domains of SAGA and ATAC subunits (Hassan et al , 2002; Vermeulen et al , 2010; Bian et al , 2011; Bonnet et al , 2014). Nevertheless, the dynamics of SAGA and ATAC interactions with chromatin are not yet well understood, as there has been no direct and systematic monitoring of the nuclear mobility of these two co‐activator complexes in live cells.…”

A role for histone acetylation in regulating transcription elongation

“…Two functionally distinct NuA3 complexes have been identified, NuA3a and b, which contain domains that have affinities for histone modifications found at gene promoters and ORFs respectively [20]. The SAGA complex has been mapped to the upstream activating sequences (UASs) of most Pol II transcribed genes, and has been proposed to act as a general cofactor for all Pol II transcription [21,22]. Importantly, SAGA occupancy does not necessarily correlate with Gcn5 activity, suggesting that Gcn5 can either act at a distance from where it is located, or is resistant to detection at some regions due to its highly dynamic interaction with the chromatin fibre [23,24].…”