Identification of Moira homologs as components of yeast SWI/SNF complexes

The Saccharomyces cerevisiae Swi/Snf complex has a role in remodeling chromatin structure to facilitate transcriptional activation. The complex has 11 components, including Swi1/Adr6, Swi2/Snf2, Swi3, Snf5, Snf6, Snf11, Swp73/Snf12, and Tfg3. Mammalian homologs of these proteins have been shown to form multiple Swi/Snf-related complexes. An S. cerevisiae Swi3 homolog (Swh3) has been characterized and evidence is presented that it associates in a complex with a Snf2 homolog, Sthl. Swh3 has been identifed as a protein that interacts with the N terminus of Snf2 in the two-hybrid system. Swh3 and Swi3 are functionally distinct, and overexpression of one does not compensate for loss of the other. Swh3 is essential for viability and does not activate transcription of reporters. The Snf2 sequence that interacts with Swh3 was mapped to a region conserved in Sth1. Swh3 and Sth1 fusion proteins interact in the two-hybrid system and coimmunoprecipitate from yeast cell extracts. Interactions between Swh3 and Sth1 were mapped and the role of a leucine zipper motif was examined in self-association of Swh3. These findings, together with previous analysis of Sth1, indicate that Swh3 and Sth1 are associated in a complex that is functionally distinct from the Swi/Snf complex and essential for viability (Treich, 1997).

Identification of Moira homologs as components of mammalian SWI/SNF complexes

A novel 15-subunit complex with the capacity to remodel the structure of chromatin, termed RSC, has been isolated from S. cerevisiae on the basis of homology to the SWI/SNF complex. At least three RSC subunits are related to SWI/SNF polypeptides: Sth1p, Rsc6p, and Rsc8p are significantly similar to Swi2/Snf2p, Swp73p, and Swi3p, respectively, and were identified by mass spectrometric and sequence analysis of peptide fragments. Like SWI/SNF, RSC exhibits a DNA-dependent ATPase activity stimulated by both free and nucleosomal DNA and a capacity to perturb nucleosome structure. RSC is, however, at least 10-fold more abundant than SWI/SNF complex and is essential for mitotic growth. Contrary to a report for SWII/SNF complex, no association of RSC (nor of SWI/SNF complex) with RNA polymerase II holoenzyme was detected (Cairns, 1996).

The SWI/SNF complex in yeast facilitates the function of transcriptional activators by opposing chromatin-dependent repression of transcription. In mammals SWI/SNF complexes are present in multiple forms made up of 9-12 proteins that are referred to as BRG1-associated factors (BAFs) ranging from 47 to 250 kD. cDNAs were isolated for human BAF155, BAF170, and BAF60. BAF155 and BAF170 are encoded by separate genes that are both homologs of yeast SWI3. Both contain a region of similarity to the DNA binding domain of myb, but lack the basic residues known to be necessary for interaction with DNA. The two SWI3 homologs copurify on antibody columns specific for either BAF155 or BAF170, indicating that they are in the same complex. BAF60 is encoded by a novel gene family. An open reading frame from yeast, which is highly homologous, encodes the previously uncharacterized 73-kD subunit of the yeast SWI/SNF complex required for transcriptional activation by the glucocorticoid receptor. BAF60a is expressed in all tissues examined, whereas BAF60b and BAF60c are expressed preferentially in muscle and pancreas, respectively. BAF60a is present within the 2000-kD BRG1 complex, whereas BAF60b is in a distinct complex that shares some but not all subunits with the BRG1 complex. The observed similarity between mammalian BAF190, BAF170, BAF155, BAF60, and BAF47 and yeast SNF2/SWI2, SWI3, SWI3, SWP73, and SNF5, respectively, underscores the similarity of the mammalian and yeast complexes. However, the complexes in mammals are more diverse than the SWI/SNF complex in yeast and are likely dedicated to developmentally distinct functions (Wang, 1996).

The RSC complex of Saccharomyces cerevisiae is closely related to the SWI/SNF complex. Both complexes are involved in remodeling chromatin structure and they share conserved components. The RSC proteins Sth1, Rsc8/Swh3, Sfh1 and Rsc6 are homologs of the SWI/SNF proteins Swi2/Snf2, Swi3, Snf5 and Swp73 respectively. To investigate the RSC complex, a temperature-sensitive swh3 allele was isolated. A screen for multicopy suppressors yielded plasmids carrying the RSC6 and MAK31 loci. RSC6 also suppresses the formamide sensitivity of a strain with a C-terminal truncation of SWH3. Swh3 and Rsc6 fusion proteins interact in the two-hybrid system and the swh3-ts mutation impairs this interaction. Finally, bacterially produced Swh3 and Rsc6 fusion proteins interact in vitro, supporting the genetic evidence for direct interaction between Swh3 and Rsc6 in vivo. Swh3 also interacts with Sth1. These findings, together with the conservation of these proteins in the SWI/SNF complex and in mammalian SWI/SNF-related complexes, strongly suggest that these proteins form a structural core for the complex (Treich, 1998).

Moira homologs and nucleosome remodeling

Protein complexes of the SWI/SNF family remodel nucleosome structure in an ATP-dependent manner. Each complex contains between 8 and 15 subunits, several of which are highly conserved between yeast, Drosophila, and humans. An ATP-dependent chromatin remodeling complex has been reconstituted using a subset of conserved subunits. Unexpectedly, both BRG1 and hBRM, the ATPase subunits of human SWI/SNF complexes, are capable of remodeling mono-nucleosomes and nucleosomal arrays as purified proteins. The addition of INI1, BAF155, and BAF170 to BRG1 increases remodeling activity to a level comparable to that of the whole hSWI/SNF complex. These data define the functional core of the hSWI/SNF complex (Phelan, 1999).

Moira homologs are modified by phosphorylation dependent on the cell cycle

SWI-SNF complexes have been implicated in transcriptional regulation by chromatin remodeling. An interaction has been identified between two components of the mammalian SWI-SNF complex and cyclin E, an essential cell cycle regulatory protein required for G1/S transition. BRG1 and BAF155, mammalian homologs of yeast SWI2 and SWI3, respectively, are found in cyclin E complexes and are phosphorylated by cyclin E-associated kinase activity. Overexpression of BRG1 causes growth arrest and induction of senescence-associated beta-galactosidase activity, which can be overcome by cyclin E. These results suggest that cyclin E may modulate the activity of the SWI-SNF apparatus to maintain the chromatin in a transcriptionally permissive state (Shanahan, 1999).

During mitosis, chromatin is condensed into mitotic chromosomes and transcription is inhibited, processes that might be opposed by the chromatin remodeling activity of the SWI/SNF complexes. Brg1 and hBrm, which are components of human SWI/SNF (hSWI/SNF) complexes, are phosphorylated during mitosis. This suggests that phosphorylation might be used as a switch to modulate SWI/SNF activity. Using an epitope-tag strategy, hSWI/SNF complexes were purified at different stages of the cell cycle, hSWI/SNF is found to be inactive in cells blocked in G2-M. Mitotic hSWI/SNF contains Brg1 but not hBrm, and is phosphorylated on at least two subunits, hSWI3 and Brg1. In vitro, active hSWI/SNF from asynchronous cells can be phosphorylated and inactivated by ERK1, and reactivated by dephosphorylation. hSWI/SNF isolated as cells traverse mitosis regain activity when its subunits are dephosphorylated either in vitro or in vivo. It is proposed that this transitional inactivation and reactivation of hSWI/SNF is required for formation of a repressed chromatin structure during mitosis and reformation of an active chromatin structure as cells leave mitosis (Sif, 1998).

Biological functions of Moira homologs

A new mouse gene has been isolated that is highly expressed in thymocytes, testis, and brain. This gene, SRG3, shows a significant sequence homology to SWI3, a yeast transcriptional activator, and its human homolog BAF155. SRG3 encodes 1,100 amino acids and has 33%-47% identity with SWI3 protein over three regions. The SRG3 protein contains an acidic NH2 terminus, a myb-like DNA binding domain, a leucine-zipper motif, and a proline- and glutamine-rich region at its COOH terminus. Rabbit antiserum raised against a COOH-terminal polypeptide of the SRG3 recognized a protein with an apparent molecular mass of 155 kD. The serum also detected a 170-kD protein that seems to be a mouse homolog of human BAF170. Immunoprecipitation of cell extract with the antiserum against the mouse SRG3 also brings down a 195-kD protein that can be recognized by an antiserum raised against human SWI2 protein. The results suggest that the SRG3 protein associates with a mouse SWI2. The SRG3 protein is expressed about three times higher in thymocytes than in peripheral lymphocytes. The expression of anti-sense RNA to SRG3 mRNA in a thymoma cell line, S49.1, reduces the expression level of the SRG3 protein, and decreases the apoptotic cell death induced by glucocorticoids. These results suggest that the SRG3 protein is involved in the glucocorticoid-induced apoptosis in the thymoma cell line. The implication is that the SRG3 may play an important regulatory role during T cell development in thymus (Jeon, 1997).

SWI/SNF complexes in yeast and higher eukaryotes are thought to facilitate gene activation and transcription factor binding by disrupting repressive chromatin structures. Little is known, however, about how these complexes target specific genes for activation. A specialized SWI/SNF-related complex (PYR complex) has been purified from murine erythroleukemia (MEL) cell nuclear extract that binds pyrimidine-rich elements at the human and murine beta-globin loci. PYR complex DNA-binding activity is restricted to definitive hematopoietic cells and is both DNA sequence- and length-dependent. Mass spectrometric identification of purified peptides and antibody supershift assays indicate that PYR complex contains at least four known mammalian SWI/SNF subunits: BAF57, INI1, BAF60a, and BAF170. PYR complex broadly footprints a 250-bp pyrimidine-rich element between the human fetal and adult beta-globin genes. A short intergenic deletion that removes this element from a human globin locus cosmid construct results in delayed human fetal-to-adult globin gene switching in transgenic mice. Taken together, the data suggest that PYR complex may act through this intergenic element to facilitate human fetal-to-adult globin gene switching, presumably by opening the locus in the region of the adult genes to permit the binding of beta-globin transcriptional activators (O'Neill, 1999).

Erythroid Kruppel-like factor (EKLF) is necessary for stage-specific expression of the human beta-globin gene. EKLF requires a SWI/SNF-related chromatin remodeling complex, EKLF coactivator-remodeling complex 1 (E-RC1), to generate a DNase I hypersensitive, transcriptionally active beta-globin promoter on chromatin templates in vitro. E-RC1 contains BRG1, BAF170, BAF155, and INI1 (BAF47) homologs of yeast SWI/SNF subunits, as well as a subunit unique to higher eukaryotes, BAF57, which is critical for chromatin remodeling and transcription with EKLF. E-RC1 displays functional selectivity toward transcription factors, since it cannot activate expression of chromatin-assembled HIV-1 templates with the E box-binding protein TFE-3. Thus, a member of the SWI/SNF family acts directly in transcriptional activation and may regulate subsets of genes by selectively interacting with specific DNA-binding proteins (Armstrong, 1998).

Biological Overview | Regulation | Developmental Biology | Effects of Mutation | References

Home page: The Interactive Fly © 1995, 1996 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.