Polycomb group (PcG) proteins function as high molecular weight complexes that maintain transcriptional repression patterns during embryogenesis. The vertebrate DNA binding protein and transcriptional repressor, YY1, shows sequence homology with the Drosophila PcG protein, pleiohomeotic (PHO). YY1 might therefore be a vertebrate PcG protein. We used Drosophila embryo and larval/imaginal disc transcriptional repression systems to determine whether YY1 repressed transcription in a manner consistent with PcG function in vivo. YY1 repressed transcription in Drosophila, and this repression was stable on a PcG-responsive promoter, but not on a PcG-nonresponsive promoter. PcG mutants ablated YY1 repression, and YY1 could substitute for PHO in repressing transcription in wing imaginal discs. YY1 functionally compensated for loss of PHO in pho mutant¯ies and partially corrected mutant phenotypes. Taken together, these results indicate that YY1 functions as a PcG protein. Finally, we found that YY1, as well as Polycomb, required the co-repressor protein CtBP for repression in vivo. These results provide a mechanism for recruitment of vertebrate PcG complexes to DNA and demonstrate new functions for YY1.

Polycomb group (PcG) proteins are responsible for maintaining transcriptional repression of developmentally important genes. However, the mechanism of PcG recruitment to specific DNA sequences is poorly understood. Transcription factor YY1 is one of the few PcG proteins with sequence-specific DNA binding activity. We previously showed that YY1 can recruit other PcG proteins to DNA, leading to histone posttranslational modifications and stable transcriptional repression. Using Drosophila transgenic approaches, we identified YY1 sequences 201-226 as necessary and sufficient for PcG transcriptional repression in vivo. When fused to a heterologous DNA-binding domain, this short 26-aa motif was sufficient for transcriptional repression, recruitment of PcG proteins to DNA, and methylation of histone H3 lysine 27. Deletion of this short YY1 motif did not affect transient transcriptional repression but ablated PcG repression, PcG protein recruitment to DNA, and methylation of H3 lysine 27. We propose that this motif be named the REPO domain for its function in recruitment of Polycomb. The REPO domain is well conserved in YY1 orthologs and in related proteins.repression ͉ transcription ͉ chromatin P olycomb group (PcG) proteins are responsible for the heritable silencing of target genes in metazoans (1, 2) and are functionally conserved in Drosophila and mammals (2, 3). Mutations in PcG genes result in misexpression of target genes with resulting homeotic transformation of body parts (4). Studies in mammals showed that PcG proteins are important for normal skeletal, muscular, and hematopoietic development (5). A variety of studies support the existence of at least two PcG complexes, each composed of several polypeptides: PRC1, containing Polycomb (Pc), polyhomeotic (Ph), dRing, and posterior sex combs (Psc); and PRC2 containing enhancer of zeste [E(z)], extra sex combs (Esc), and suppressor of zeste 12 [Su(z)12] (1, 2). These complexes are recruited to chromatin, where they maintain transcriptional silencing. An unresolved issue is the mechanism of recruitment of PcG complexes to appropriate target genes.The mechanism of PcG complex recruitment has remained elusive, especially in mammals. Drosophila Polycomb response elements (PREs) bind to PcG proteins and silence cis-linked genes in vivo (1, 2). The large size and poor sequence homology of characterized PREs has hampered the search for recruiting factors. Furthermore, nearly all characterized PcG proteins lack sequencespecific DNA-binding activity. Exceptions include the Drosophila PcG proteins Pleiohomeotic (PHO) and Pleiohomeotic-like (PHOL) and their mammalian counterpart, the vertebrate transcription factor Yin Yang 1 (YY1) (6, 7). Homology between these transcription factors is localized to the C-terminal zinc-finger DNA-binding domain (YY1 residues 298-414, 95% identical to PHO) and a short internal sequence (YY1 residues 205-226, 82% identical to PHO). We previously demonstrated that, similar to PHO, YY1 can function as a PcG protein to mediate silencing (8). Th...

The Polycomb Group (PcG) complex of transcriptional repressors is critical for the maintenance of stage-specific developmental gene expression, stem cell maintenance and for large-scale chromosomal dynamics. Functional deficiency of a single PcG gene can severely compromise PcG function, leading to developmental defects, embryonic lethality, or a number of malignancies. Despite the critical nature of PcG proteins, the mechanisms by which these complexes mediate their effects are relatively uncharacterized. Nearly all vertebrate PcG proteins lack inherent DNA binding capacity, making it unclear how they are targeted to Polycomb response element (PRE) sequences. Transcription factor YY1 is a functional ortholog of a Drosophila PcG protein, Pleiohomeotic (PHO), one of the few PcG proteins with specific DNA binding capability, and YY1 can recruit PcG proteins to specific DNA sequences. A small 25 amino acid YY1 domain (the REPO domain) is necessary and sufficient for recruitment of PcG proteins to DNA and for transcriptional repression. We show here that the YY1 REPO domain interacts with PcG protein Yaf2 and recruits Yaf2 to DNA. Interaction is lost when the YY1 REPO domain is deleted. In addition we show that Yaf2, when linked to a heterologous DNA binding domain, can recruit PcG proteins to DNA leading to transcriptional repression. When the Drosophila homolog of Yaf2 (dRYBP) is mutated, PcG recruitment to DNA is reduced. Taken together, our results suggest that Yaf2 serves as a molecular bridge between YY1 and other PcG complex proteins.

Polycomb Group (PcG) proteins are crucial for epigenetic inheritance of cell identity and are functionally conserved from Drosophila to humans. PcG proteins regulate expression of homeotic genes and are essential for axial body patterning during development. Earlier we showed that transcription factor YY1 functions as a PcG protein. YY1 also physically interacts with YAF2, a homolog of RYBP. Here we characterize the mechanism and physiologic relevance of this interaction. We found phenotypic and biochemical correction of dRYBP mutant flies by mouse YAF2 demonstrating functional conservation across species. Further biochemical analysis revealed that YAF2 bridges interaction between YY1 and the PRC1 complex. ChIP assays in HeLa cells showed that YAF2 is responsible for PcG recruitment to DNA, which is mediated by YY1 DNA binding. Knock-down of YY1 abrogated PcG recruitment, which was not compensated by exogenous YAF2 demonstrating that YY1 DNA binding is a priori necessary for Polycomb assembly on chromatin. Finally, we found that although YAF2 and RYBP regulate a similar number of Polycomb target genes, there are very few genes that are regulated by both implying functional distinction between the two proteins. We present a model of YAF2-dependent and independent PcG DNA recruitment by YY1.