In vitro data suggest that the human RbAp46 and RbAp48 genes encode proteins involved in multiple chromatin remodeling complexes and are likely to play important roles in development and tumor suppression. However, to date, our understanding of the role of RbAp46/RbAp48 and its homologs in metazoan development and disease has been hampered by a lack of insect and mammalian mutant models, as well as redundancy due to multiple orthologs in most organisms studied. Here, we report the first mutations in the single Drosophila RbAp46/RbAp48 homolog Caf1, identified as strong suppressors of a senseless overexpression phenotype. Reduced levels of Caf1 expression result in flies with phenotypes reminiscent of Hox gene misregulation. Additionally, analysis of Caf1 mutant tissue suggests that Caf1 plays important roles in cell survival and segment identity, and loss of Caf1 is associated with a reduction in the Polycomb Repressive Complex 2 (PRC2)-specific histone methylation mark H3K27me3. Taken together, our results suggest suppression of senseless overexpression by mutations in Caf1 is mediated by participation of Caf1 in PRC2-mediated silencing. More importantly, our mutant phenotypes confirm that Caf1-mediated silencing is vital to Drosophila development. These studies underscore the importance of Caf1 and its mammalian homologs in development and disease.
"(A) A schematic representation of a PRC2 component, CAF1p55, containing seven WD40 domains, is shown. The Caf1p55short allele carries a point mutation that changes Gly to a stop codon at AA position 79, whereas the Caf1p55long alleles carries a point mutation that changes Gly to Asp at AA position 375 (Anderson et al. 2011). (B) To determine the effect of altering Caf1p55 activity in Psc-induced rough eye phenotype, a single copy of either Caf1p55short mutant or Caf1p55long mutant chromosome was introduced in GMR-Gal4;f00391 flies. "
[Show abstract][Hide abstract] ABSTRACT: A Polycomb group protein, Posterior sex combs (Psc), was identified in a genetic screen designed to find factors that can specifically induce morphological defects in rbf1 mutant eyes. We discovered that rbf1 mutations enhance developmental phenotypes caused by Psc overexpression such as ectopic cell death and disorganized ommatidia. Our genetic analysis revealed that Psc-induced developmental defects are strongly influenced by Caf1p55, which is a shared component of several chromatin-associated complexes including a histone chaperone complex dCAF-1. Interestingly, the expression levels of dCAF-1 components, CAF1p105 and CAF1p180, are increased in rbf1 mutants, while the expression level of CAF1p55 itself remains relatively unchanged. We demonstrated that the increased levels of CAF1p105 and CAF1p180 are required for the hypersensitivity of rbf1 mutant cells to Psc-induced cell death and for the developmentally regulated cell death normally observed in rbf1 mutant eyes. We propose that Caf1p105 and Caf1p180 are important determinants of cell death sensitivity in rbf1 mutant cells and contribute to the genetic interaction between Psc and rbf1.
"In Drosophila larvae, Pcl facilitates the recruitment of PRC2 to chromosomes (Savla, Benes, Zhang, & Jones, 2008). Classical Polycomb phenotypes related to the misregulation of Hox genes have been described for all PRC2 members and with the exception of esc and escl, all homozygous null alleles show larval lethality (Anderson et al., 2011; Birve et al., 2001; Duncan, 1982; Phillips & Shearn, 1990; Struhl & Brower, 1982). According to the Drosophila developmental transcriptome project (Gelbart & Emmert, 2011; McQuilton, St Pierre, & Thurmond, 2012), mRNA levels for all PRC2 genes peak in early embryos, then decline at larval stages and increase in female but not male adults. "
[Show abstract][Hide abstract] ABSTRACT: Proper development of an embryo requires tightly controlled expression of specific sets of genes. In order to generate all the lineages of the adult, populations of pluripotent embryonic stem cells differentiate and activate specific transcriptional programs whereas others are shutdown. The role of transcription factors is obvious in promoting expression of such developmental genes; however maintenance of specific states throughout cell division needs additional mechanisms. Indeed, the nucleoprotein complex of DNA and histones, the chromatin, can act as a facilitator or barrier to transcription depending on its configuration. Chromatin-modifying enzymes regulate accessibility of DNA by establishing specific sets of chromatin, which will be either permissive or repressive to transcription. In this review, we will describe the H3K9/HP1 and Polycomb pathways, which mediate transcriptional repression by modifying chromatin. We discuss how these two major epigenetic silencing modes are dynamically regulated and how they contribute to the early steps of embryo development.
Current Topics in Developmental Biology 04/2013; 104:243-91. DOI:10.1016/B978-0-12-416027-9.00008-5 · 4.68 Impact Factor
"SUZ12 directly binds to the promoters of the PRC2 target genes probably through its Zinc-finger domains, and Suz12 expression is increased in human colon tumors . In the mammalian PRC2 complex, there is a pair of highly homologous WD40 proteins RbAP46/48, which is shown to be essential for cell survival and patterning in Drosophila development, and knockdown of the Drosophila RbAP46/48 P55 causes severe reduction in histone H3K27me3 methylation level . Although AEBP2 is not essential for PRC2 activity, it is required for optimal enzymatic activity . "
[Show abstract][Hide abstract] ABSTRACT: PRC2 is the major H3K27 methyltransferase and is responsible for maintaining repressed gene expression patterns throughout development. It contains four core components: EZH2, EED, SUZ12 and RbAp46/48 and some cell-type specific components. In this study, we focused on characterizing the histone binding domains of PHF1 and PHF19, and found that the Tudor domain of PHF1 and PHF19 selectively binds to histone H3K36me3. Structural analysis of these Tudor domains also shed light on how these Tudor domains selectively binds to H3K36me3. The Tudor domain binding of H3K36me3 of PHF1, PHF19 and likely MTF2 provide another recruitment and regulatory mechanism for the PRC2 complex. The first PHD domains of PHF1 and PHF19 do not exhibit histone H3K4 binding ability, nor do they affect the Tudor domain binding to histones.
Biochemical and Biophysical Research Communications 12/2012; 430(2). DOI:10.1016/j.bbrc.2012.11.116 · 2.30 Impact Factor
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