brahma
SWI/SNF complex and nucleosome remodeling (part 2/2) Chromatin presents a significant obstacle to transcription. This paper proposes two means of overcoming its repressive effects: histone acetylation and the activities
of the Swi-Snf complex. Histone acetylation and Swi-Snf activity have been shown to be crucial for transcriptional induction
and to facilitate binding of transcription factors to DNA. By regulating the activity of the Swi-Snf complex in vivo, active
transcription is found to require continuous Swi-Snf function, demonstrating a role for this complex beyond the induction of transcription. Despite the
presumably generalized packaging of genes into chromatin, previous studies have indicated that the transcriptional requirements for the histone
acetyltransferase, Gcn5, and the Swi-Snf complex are limited to a handful of genes. However, inactivating Swi-Snf function in cells also lacking
GCN5 reveals defects in transcription for several genes previously thought to be SWI-SNF- and GCN5-independent. These findings suggest that
chromatin remodeling plays a widespread role in gene expression and that these two chromatin remodeling activities perform independent and
overlapping functions during transcriptional activation (Biggar, 1999).
The SWI-SNF genes are crucial for initiating transcription of inducible genes such as HO, SUC2 and INO1. In vitro studies suggest that the Swi-Snf complex stably disrupts nucleosomes and provides transcription factors with opportunities to bind nucleosomal DNA. However, inactivating Swi2 in vivo rapidly silences on-going transcription from the SUC2 promoter and a SWI-SNF-dependent variant of the GAL1 promoter, GALp*. These results demonstrate that Swi-Snf function is not limited to the initiation of gene expression and suggest that transcription continuously requires chromatin remodeling. If the biochemical results demonstrating stable nucleosome disruption by Swi-Snf reflect the physiological activity of this complex, then Swi-Snf must perform additional, transitory functions during transcriptional activation. Alternatively, yeast may possess additional activities capable of rapidly restoring nucleosomes to their base states in the absence of Swi-Snf activity (Biggar, 1999).
The continuous requirement for Swi2 activity indicates that Swi-Snf accomplishes more than just providing transcriptional activators with access to their binding sites on promoters. Once bound to DNA, transcription factors stimulate transcription even in the presence of histones, suggesting that, once histones are disrupted and activators bind DNA, further chromatin remodeling should be unnecessary for transcription. Consistent with a role for Swi-Snf beyond activator binding, swi-snf mutations minimally diminish Gal4 binding at SWI-SNF-dependent promoters, such as GALp* (Biggar, 1999 and references).
Components of the Swi-Snf complex have been found associated with the RNA polymerase holoenzyme. Though this interaction is a controversial one, other components of the holoenzyme are also crucial for on-going transcription. The continuous requirement for Swi-Snf activity could be explained if, as a component of the holoenzyme, Swi-Snf acts during each round of transcriptional initiation to disrupt nucleosomes and facilitate transcriptional initiation by RNA polymerase. Alternatively, Swi-Snf function may be essential for transcriptional elongation. In some studies, RNA polymerases appear to proceed unimpeded through nucleosomes, but physiological chromatin structures may pose more substantial challenges to polymerase elongation in vivo. Also, other studies have identified activities necessary for polymerases to elongate through nucleosomes (Biggar, 1999 and references).
Both Swi-Snf and histone acetyltransferases have been proposed as means of remodeling nucleosomes. However, the functional relationship between these activities had not been addressed. By comparing expression from several distinct promoters in cells lacking the activities of one or both complexes, inactivating Swi2 in cells lacking GCN5 is found to significantly reduced transcription. These results show that Swi-Snf plays a role in transcription that is independent of GCN5, thereby demonstrating that the two complexes act in distinct pathways to activate transcription. Thus, histone acetylation by Gcn5 does not simply serve to stabilize nucleosomes that have been disrupted by Swi-Snf, or to label nucleosomes as future targets of Swi-Snf activity. These data do not exclude a role for Gcn5 in Swi-Snf function, but clearly Swi-Snf retains significant transcriptional activity in the absence of GCN5 (Bigger, 1999).
These results further suggest that Swi-Snf and Gcn5-Ada perform redundant functions during transcription. The transcriptional requirements for Swi2 function depend on the GCN5 background of the yeast: the absence of GCN5 reveals novel functions for Swi2. Inactivating Swi2 in a wild-type GCN5 background produces minimal defects in GAL1, PGK1 and ACT1 transcription (1.0- to 1.7-fold), but inactivating Swi2 in a gcn5Delta background reduces transcription of these genes 3.9- to 4.7-fold. These findings indicate that Swi-Snf and Gcn5-Ada perform overlapping functions, since one complex is dispensable for transcription unless cells lack the other complex. Thus, the Swi-Snf and Gcn5-Ada complexes represent two independent means of accomplishing the same step during transcription, namely chromatin remodeling (Biggar, 1999).
DNA-binding transcriptional activators often employ additional mechanisms to stimulate gene expression. The results presented here suggest that Gal4, a prototypical transcriptional activator, utilizes the Swi-Snf and Gcn5-Ada complexes as co-activators of transcription. The induction of GAL1 transcription depends on Gal4 and involves changes in nucleosome positions on the promoter. Consistent with a role for chromatin remodeling during GAL1 transcription, cells lacking the activity of both Swi-Snf and Gcn5-Ada show profound defects in GAL1 expression. A model of GAL1 transcriptional activation is proposed in which Gal4 engages the functions of both Swi-Snf and Gcn5-Ada to disrupt nucleosomes and activate transcription (Biggar, 1999 and references).
GAL1 expression involves separable events of derepression and transcriptional activation. Repression at the GAL1 promoters depends on the transcriptional co-repressor composed of Ssn6 and Tup1. Ssn6-Tup1 positions nucleosomes on target promoters, and the disruption of these structures accompanies transcriptional activation. Although these data suggest a possible functional interaction between Ssn6-Tup1 and Swi-Snf, Swi-Snf function at the GAL1 promoter is found to be dedicated to transcriptional activation by Gal4 instead of counteracting repression by Ssn6-Tup1. Transcriptional activation by Gal4 under derepressing conditions requires continuous Swi-Snf activity, and derepression prior to Swi2 inactivation fails to restore transcription from GALp*. Consistent with these results, chromatin remodeling and transcription from the SUC2 promoter require SWI1, even in strains lacking SSN6 or TUP1 (Biggar, 1999 and references).
These results reveal that, in the absence of GCN5, Swi-Snf activity is required for transcription of multiple, distinctly regulated genes. The general packaging of genes into chromatin, along with the demonstration that histone depletion appears to activate promoters non-specifically, suggests that transcription of many, if not all, genes should require chromatin remodeling. Until now, however, the roles for chromatin remodeling complexes, including the Swi-Snf, Gcn5-Ada and Rsc complexes, were thought to be limited to a handful of promoters. The finding that swi2-6.3;gcn5Delta double mutants display widespread transcriptional defects substantiates the notion that chromatin remodeling is a general requirement for gene expression, and establishes the Swi-Snf and Gcn5-Ada complexes as major activities fulfilling this role (Biggar, 1999 and references).
Defined here are three general classes of genes based on their requirements for Swi-Snf and Gcn5-Ada functions. Expression of one class, including HO and SUC2, depends on both complexes. A second category consists of genes whose expression depends on either the Swi-Snf or the Gcn5-Ada complex. Transcription of these genes persists unless both complexes are inactivated. Examples of this class include genes whose expression is regulated (GAL1) and genes that are constitutively expressed (PGK1 and ACT1). The third category consists of genes whose expression is affected minimally or indirectly in strains lacking both Swi-Snf and Gcn5-Ada (e.g., TCM1). This class of genes may not require chromatin remodeling activities due to the presence of sequences in the promoters, such as nucleosome positioning elements, that prevent nucleosomes from repressing transcription. Alternatively, these promoters may use chromatin remodeling activities other than Swi-Snf and Gcn5-Ada to disrupt nucleosomes and activate transcription. The Rsc complex, for example, contains many Swi-Snf-related proteins and remodels nucleosomes in vitro, but its role in transcription awaits further study (Biggar, 1999 and references).
In summary, a conditional allele of Swi2 was used to regulate the function of the Swi-Snf complex in vivo and explore the physiological function of this chromatin remodeling complex. Swi-Snf is found to play a continuous role in transcription; Swi-Snf and the histone acetyltransferase, Gcn5-Ada, function in independent and redundant pathways to stimulate transcription. Furthermore, in the absence of GCN5, Swi-Snf activates transcription of several distinctly regulated genes, indicating that these chromatin remodeling activities play widespread roles in the expression of many yeast genes (Biggar, 1999).
The generation of a local chromatin topology conducive to transcription is a key step in gene regulation. The yeast SWI/SNF
complex is the founding member of a family of ATP-dependent remodelling activities capable of altering chromatin structure both
in vitro and in vivo. Despite its importance, the pathway by which the SWI/SNF complex disrupts chromatin structure is unknown.
A model system has been used to demonstrate that the yeast SWI/SNF complex can reposition nucleosomes in an ATP-dependent
reaction that favors attachment of the histone octamer to an acceptor site on the same molecule of DNA (in cis).
SWI/SNF-mediated displacement of the histone octamer is effectively blocked by a barrier introduced into the DNA, suggesting
that this redistribution involves sliding or tracking of nucleosomes along DNA, and that it is achieved by a catalytic mechanism. It is concluded that SWI/SNF catalyses the redistribution of nucleosomes along DNA in cis, which may represent a general mechanism by which ATP-dependent chromatin remodelling occurs (Whitehouse, 1999).
SWI/SNF regulates growth control, differentiation and tumor suppression, yet few direct targets of this chromatin-remodeling complex
have been identified in mammalian cells. SWI/SNF is required for interferon (IFN)-gamma induction of CIITA,
the master regulator of major histocompatibility complex class II expression. Despite the presence of functional STAT1, IRF-1 and
USF-1, activators implicated in CIITA expression, IFN-gamma did not induce CIITA in cells lacking BRG1 and hBRM, the
ATPase subunits of SWI/SNF. Reconstitution with BRG1, but not an ATPase-deficient version of this protein (K798R), rescues CIITA induction, and enhances the
rate of induction of the IFN-gamma-responsive GBP-1 gene. Notably, BRG1 inhibits the CIITA promoter in transient transfection assays, underscoring
the importance of an appropriate chromosomal environment. Chromatin immunoprecipitation has revealed that BRG1 interacts directly with the endogenous CIITA
promoter in an IFN-gamma-inducible fashion, while in vivo DNase I footprinting and restriction enzyme accessibility assays have shown that chromatin
remodeling at this locus requires functional BRG1. These data provide the first link between a cytokine pathway and SWI/SNF, and suggest a novel role for this
chromatin-remodeling complex in immune surveillance (Pattenden, 2002).
The 12-subunit Swi/Snf chromatin remodeling complex is conserved from yeast to humans. It functions to alter nucleosome positions by either sliding nucleosomes on DNA or evicting histones. Interestingly, 20% of all human cancers carry mutations in subunits of the Swi/Snf complex. Many of these mutations cause protein instability and loss, resulting in partial Swi/Snf complexes. Although several studies have shown that histone acetylation and activator-dependent recruitment of Swi/Snf regulate its function, it is less well understood how subunits regulate stability and function of the complex. Using functional proteomic and genomic approaches, this study has assembled the network architecture of yeast Swi/Snf. In addition, it was found that subunits of the Swi/Snf complex regulate occupancy of the catalytic subunit Snf2 (see Drosophila Brahma), thereby modulating gene transcription. These findings have direct bearing on how cancer-causing mutations in orthologous subunits of human Swi/Snf may lead to aberrant regulation of gene expression by this complex (Dutta, 2017).
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