InteractiveFly: GeneBrief

Jumonji, AT rich interactive domain 2: Biological Overview | References

Gene name - Jumonji, AT rich interactive domain 2

Synonyms -

Cytological map position - 67B9-67B10

Function - Transcription factor

Keywords - component of Polycomb repressive complex 2 which is involved in methylation of histone 3 at K27, required for transcriptional repression

Symbol - Jarid2

FlyBase ID: FBgn0036004

Genetic map position - chr3L:9,470,150-9,479,632

Classification - ARID/BRIGHT DNA binding domain, JmjC domain, hydroxylase

Cellular location - nuclear

NCBI links: EntrezGene
Recent literature
Sanulli, S., et al. (2015). Jarid2 methylation via the PRC2 complex regulates H3K27me3 deposition during cell differentiation. Mol Cell 57: 769-783. PubMed ID: 25620564
Polycomb Group (PcG) proteins maintain transcriptional repression throughout development, mostly by regulating chromatin structure. Polycomb Repressive Complex 2 (PRC2), a component of the Polycomb machinery, is responsible for the methylation of histone H3 lysine 27 (H3K27me2/3). Jarid2 was previously identified as a cofactor of PRC2, regulating PRC2 targeting to chromatin and its enzymatic activity. Deletion of Jarid2 leads to impaired orchestration of gene expression during cell lineage commitment. This study reveals an unexpected crosstalk between Jarid2 and PRC2, with Jarid2 being methylated by PRC2. This modification is recognized by the Eed (see Drosophila Extra Sexcombs) core component of PRC2 and triggers an allosteric activation of PRC2's enzymatic activity. Jarid2 methylation is shown to be important to promote PRC2 activity at a locus devoid of H3K27me3 and for the correct deposition of this mark during cell differentiation. These results uncover a regulation loop where Jarid2 methylation fine-tunes PRC2 activity depending on the chromatin context.
Goto, M., Toda, N., Shimaji, K., Suong, D. N., Vo, N., Kimura, H., Yoshida, H., Inoue, Y. H. and Yamaguchi, M. (2016). Polycomb-dependent nucleolus localization of Jumonji/Jarid2 during Drosophila spermatogenesis. Spermatogenesis 6(3): e1232023. PubMed ID: 28144496
Drosophila Jumonji/Jarid2 (dJmj) has been identified as a component of Polycomb repressive complex 2. However, it is suggested that dJmj has both PRC-dependent and -independent roles. Subcellular localization of dJmj during spermatogenesis is unknown. Immunocytochemical analyses was performed with specific antibodies to dJmj and tri-methylation at lysine 27 on histone H3 (H3K27me3). Interestingly, dJmj exclusively localizes at nucleolus in the late growth stage. Examination of the dJmj localization in various Polycomb group (PcG) mutant lines at the late growth stage allowed identification of some PcG genes, including Polycomb (Pc), to be responsible for dJmj recruitment to nucleolus. In addition, size of nucleolus was decreased in some of these mutant lines. In a mutant of testis-specific TAF homolog (tTAF) that is responsible for nucleolus localization of Pc, dJmj signals were detected not only at nucleolus but also on the condensed chromatin in the late growth stage. Duolink In situ Proximity ligation assay clarified that Pc interacts with dJmj at nucleolus in the late growth stage. Furthermore, the level of H3K27me3 decreased in nuclei at this stage. Taken together, it is concluded that tTAF is responsible for recruitments of dJmj to nucleolus in the late growth stage that appears to be mediated by Pc. Compartmentalization of dJmj in nucleolus together with some of PcG may be necessary to de-repress the expression of genes required to cellular growth and proliferation in the following meiotic divisions.
Shalaby, N. A., Sayed, R., Zhang, Q., Scoggin, S., Eliazer, S., Rothenfluh, A. and Buszczak, M. (2017). Systematic discovery of genetic modulation by Jumonji histone demethylases in Drosophila. Sci Rep 7(1): 5240. PubMed ID: 28701701
Jumonji (JmjC) domain proteins (see Jarid2) influence gene expression and chromatin organization by way of histone demethylation, which provides a means to regulate the activity of genes across the genome. JmjC proteins have been associated with many human diseases including various cancers, developmental and neurological disorders, however, the shared biology and possible common contribution to organismal development and tissue homeostasis of all JmjC proteins remains unclear. This study systematically tested the function of all 13 Drosophila JmjC genes. Generation of molecularly defined null mutants revealed that loss of 8 out of 13 JmjC genes modify position effect variegation (PEV) phenotypes, consistent with their ascribed role in regulating chromatin organization. However, most JmjC genes do not critically regulate development, as 10 members are viable and fertile with no obvious developmental defects. Rather, it was found that different JmjC mutants specifically alter the phenotypic outcomes in various sensitized genetic backgrounds. The data demonstrate that, rather than controlling essential gene expression programs, Drosophila JmjC proteins generally act to "fine-tune" different biological processes.
Suong, D. N. A., Shimaji, K., Pyo, J. H., Park, J. S., Yoshida, H., Yoo, M. A. and Yamaguchi, M. (2017). Overexpression of dJmj differentially affects intestinal stem cells and differentiated enterocytes. Cell Signal 42: 194-210. PubMed ID: 29102770
Jumonji (Jmj)/Jarid2 is a DNA-binding transcriptional repressor mediated via histone methylation. Nevertheless, the well-known function of Jmj is as a scaffold for the recruitment of various complexes including Polycomb repressive complex 2 (PRC2), and is required for mouse embryonic stem cell development. However, PRC2 independent function is suggested for Drosophila Jumonji (dJmj). To clarify the function of dJmj during cell differentiation, the Drosophila adult intestinal stem cell system was used that allows following of stem cell behaviors in vivo. Overexpression of dJmj in intestinal stem cells/enteroblasts (ISCs/EBs) induces cell-autonomous ISC proliferation followed by differentiation, that is controlled by the Notch and EGFR pathway. In contrast, overexpression of dJmj in enterocytes (ECs) resulted in activation of the JNK pathway in ECs followed by the induction of apoptosis. Activated JNK increased the level of Yorkie in ECs and induced the reduction of Upd proteins and EGFR ligands, which activated the JAK/STAT and EGFR pathway in both ISCs and EBs to promote ISC proliferation. The Notch signaling pathway appears to be highly activated to support the differentiation of EBs to ECs. Thus, the combination of these signaling pathways caused by ECs-specific dJmj-overexpression induced non-cell-autonomous ISC proliferation and differentiation. Surprisingly, these effects did not relate to H3K27me3 status, likely represented PRC2 activity, in cells that overexpressed dJmj. Instead of this, the disappearance of H3K27me3 in ISC/EB-specific overexpressed dJmj suggested a possible PRC2-independent role of dJmj in regulating chromatin structure.


Jarid2 was recently identified as an important component of the mammalian Polycomb repressive complex 2 (PRC2), where it has a major effect on PRC2 recruitment in mouse embryonic stem cells. Although Jarid2 is conserved in Drosophila, it has not previously been implicated in Polycomb (Pc) regulation. Therefore, Drosophila Jarid2 and its associated proteins were purified, and it was found that Jarid2 associates with all of the known canonical PRC2 components, demonstrating a conserved physical interaction with PRC2 in flies and mammals. Furthermore, in vivo studies with Jarid2 mutants in flies demonstrate that among several histone modifications tested, only methylation of histone 3 at K27 (H3K27), the mark implemented by PRC2, was affected. Genome-wide profiling of Jarid2, Su(z)12 (Suppressor of zeste 12), and H3K27me3 occupancy by chromatin immunoprecipitation with sequencing (ChIP-seq) indicates that Jarid2 and Su(z)12 have very similar distribution patterns on chromatin. However, Jarid2 and Su(z)12 occupancy levels at some genes are significantly different, with Jarid2 being present at relatively low levels at many Pc response elements (PREs) of certain Homeobox (Hox) genes, providing a rationale for why Jarid2 was never identified in Pc screens. Gene expression analyses show that Jarid2 and E(z) (Enhancer of zeste, a canonical PRC2 component) are not only required for transcriptional repression but might also function in active transcription. Identification of Jarid2 as a conserved PRC2 interactor in flies provides an opportunity to begin to probe some of its novel functions in Drosophila development (Herz, 2012).

Different and distinct gene expression patterns are established during development, which need to be maintained and regulated. This is important to allow for the integrity of cell identity and thus the functional preservation of tissues and organs. However, at the same time, transcribed loci must be equipped with an intrinsic flexibility to regulate these expression patterns and initiate changes if necessary. The core components that are required for the maintenance of gene expression or gene repression have been characterized quite extensively to date. Trithorax and Polycomb group genes play antagonistic roles in determining whether a gene is transcriptionally turned on or off, respectively . In Drosophila, so far four distinct complexes, pleiohomeotic repressive complex (PhoRC), Polycomb repressive complex 2 (PRC2), Polycomb repressive complex 1 (PRC1), and recently Polycomb repressive deubiquitinase (PR-DUB) have been described to play a role in Polycomb group-mediated gene repression. However, little is known about the factors involved in controlling recruitment and activity of these complexes on chromatin or about the mechanisms that drive such changes. It should be expected that quite a significant number of proteins would convey Polycomb group-mediated transcriptional changes in order to allow an uncoupling of individual gene activity from that of a group of Polycomb group-controlled genes. Functional redundancy might account for part of the problem to discover such candidates. Furthermore, biochemical approaches might be hindered by the fact that such context-specific and more gene-specific recruiters are contained in only a minor fraction of Polycomb repressive complexes (Herz, 2012).

Recently, Jarid2 the founding member of the JmjC domain-containing protein family (Klos, 2006), which plays important developmental roles in mice and Drosophila (Jung, 2005; Landeira, 2011; Sasai, 2007), has been characterized as a component of PRC2 in embryonic stem (ES) cells (Landeira, 2010; Pasini, 2011, Peng, 2009, Shen, 2009, Zhang, 2011). The consensus indicates that in ES cells, PRC2 recruitment to many of its targets requires Jarid2. However, levels of bulk histone 3 trimethylated at K27 (H3K27me3) in ES cells depleted of Jarid2 were reported to be only slightly changed at best. This also holds true when individual PRC2 target genes are analyzed. Even though core components of the PRC2 complex were lost from chromatin in the absence of Jarid2, H3K27me3 was not reproducibly affected to a similar degree. Additionally, gene expression analyses in Jarid2-/- ES cells did not confirm a genome-wide derepression of PRC2 target genes as would be expected for any core component of PRC2 (Landeira, 2010; Herz, 2012 and references therein).

To further address whether Jarid2 constitutes a core PRC2 component, is involved in recruitment of PRC2 to chromatin, and regulates H3K27 methylation in Drosophila, a Jarid2 complex was purified from flies and a global in vivo analysis of was performed of Suppressor of zeste 12 [Su(z)12] and H3K27me3 occupancy in Jarid2 mutant animals. The data confirm that Drosophila Jarid2 purifies with the core members of the PRC2 complex. In imaginal discs, global H3K27me3 levels are only weakly but reproducibly affected under Jarid2 mutant and Jarid2-overexpressing conditions. These genome-wide studies suggest that in Drosophila, under physiological conditions, Jarid2 does not appear to be a canonical component of the PRC2 complex as PRC2 recruitment is not altered on most target genes in Jarid2 mutant animals. Interestingly, overexpression of Jarid2 results in reduced Su(z)12 binding and changed chromatin compaction on polytene chromosomes, highlighting a possible role for Jarid2 in altering chromatin architecture. Genome-wide, Jarid2 and Su(z)12 binding correlate very well. However, certain loci, such as Homeobox (Hox) genes, differ significantly from this pattern. Here, Jarid2 occupancy on Polycomb response elements (PREs) is often very low where usually the highest enrichment for Su(z)12 can be observed. Gene expression analyses suggest a PRC2-dependent and -independent role for Jarid2 in transcriptional regulation. Jarid2 appears to be involved in the regulation of a certain number of PRC2 target genes and also transcriptionally controls a subset of genes independently of PRC2. These data not only imply a function for Jarid2 and PRC2 in transcriptional repression but also support a possible role for both Jarid2 and PRC2 in active transcription on genes that are occupied by these factors (Herz, 2012).

This study describes the purification of a Jarid2 complex in Drosophila. Consistent with previous results in mammalian systems, Jarid2 was found to be a component of PRC2. Evidence is provided that in imaginal discs and on polytene chromosomes, Jarid2 is required to fine-tune global H3K27me3 levels. Jarid2 might accomplish this by modulating the activity of the core complex [E(z), Su(z)12, Esc, and Caf1]. The data indicate that Jarid2 could play an inhibitory role in the implementation of H3K27me3 as Jarid2 mutant imaginal disc clones display a global increase and as overexpression of Jarid2 results in a reduction in H3K27me3. Despite having a JmjC domain, Jarid2 has been predicted (Klose, 2006) and reported (Li, 2010; Shen, 2009) to be catalytically inactive as a histone demethylase. Therefore, it is unlikely but not impossible that it could function in this manner toward H3K27me3, thereby counteracting PRC2 activity. Even if Jarid2 would be inactive as a histone demethylase, it might still be able to bind to chromatin and prevent spreading of the H3K27me3 mark, such as opposing a possible positive spreading effect of Esc (EED in mammals) (Herz, 2012).

Furthermore, even though Jarid2 could be purified with the PRC2 core members and its occupancy generally correlates very well with canonical PRC2 components such as Su(z)12, it does not appear to play a significant role in regulating PRC2 recruitment in a physiological context, as assessed by Jarid2 mutant animal studies. Apparent differences with published mammalian studies, which imply a major role for Jarid2 in recruitment of PRC2, could be explained by variation in the mechanisms employed or by the fact that the recruitment of PRC2 in ES cells generally differs from that in differentiated tissues. For example, PREs have been known to be highly effective in recruiting PRC2 to target sites in Drosophila. In mammals, attempts have been made to identify functionally analogous sequences but with only limited success. Indeed, it seems more likely that the recruitment of PRC2 in mammals not only requires specific sequences but is also more dependent on additional factors (proteins and RNA), which might explain why PRC2 recruitment is more strongly affected in Jarid2-depleted cells and why PRC1 recruitment in some instances appears to be dependent on PRC2 (H3K27me3). However, the data in Drosophila salivary glands suggest that recruitment of PRC2 (and methylation of H3K27) is not a prerequisite for targeting of PRC1, and the generality of this mechanism is also increasingly questioned in the mammalian system. Nonetheless, when Jarid2 is overexpressed in Drosophila, changes in chromosome compaction can be observed. Under these conditions, Jarid2 extensively occupies the chromosomes, and Su(z)12 localization and H3K27me3 are negatively affected). It is possible that increasing Jarid2 levels beyond a certain physiological level might interfere with PRC2 integrity. Larger amounts of Jarid2 might alter the stoichiometry of the PRC2 subunits, resulting in destabilization of the PRC2 complex on chromatin (Herz, 2012).

Jarid2 also behaves differently from other canonical PRC2 members in Drosophila, as is evident from its binding pattern on certain Hox genes. At Hox genes, occupancy of PRE sites by canonical PRC2 members is one of the highest in the whole genome. In contrast, Jarid2 displays relatively low occupancy on many of these loci, implying a minor or different function for Jarid2 in controlling transcription of these well-described PRC2 targets. It is also possible that at these loci Jarid2 has a more transient association or even that it is less accessible to interact with the antibodies that were have generated. However, these findings are also in agreement with modifier screens that have been performed in Drosophila to identify major regulators of Polycomb group-mediated phenotypes but that were unable to capture Jarid2 (Herz, 2012).

Additionally, the data suggest that Jarid2 appears to control PRC2-dependent transcription, although not necessarily in the same way as expected for canonical PRC2 members. For example, in contrast to the mammalian findings, this study observed that PRC2-mediated transcriptional regulation by Jarid2 in Drosophila is generally independent of changes in Su(z)12 occupancy and does not correlate with changes in H3K27me3 enrichment. However, it needs to be stressed that most Jarid2/PRC2 cobound genes with altered expression patterns in Jarid2 mutants and E(z)-RNAi larvae contain no or low levels of H3K27me3, which is in contrast to the mammalian system where PRC2 components are usually found only at genes with high H3K27me3 enrichment. Nonetheless, in Drosophila, genes with high H3K27me3 enrichment exist that change in transcription in Jarid2 mutants and E(z)-RNAi animals, demonstrating that H3K27me3 is not necessarily instructive for transcriptional repression per se. To date most of the evidence ascribing to H3K27me3 the role of a repressive mark is based on correlation from the observation that PRC2 components colocalize with H3K27me3 and that the respective genes seem to be transcriptionally silenced. The data imply that this might generally be the case but that there are also exceptions to the rule. That certain H3K27me3 patterns can also be connected to transcriptionally active genes in mammals has just recently been reported (Young, 2011; Herz, 2012 and references therein).

Finally, the results imply that Jarid2 and PRC2 are not only involved in maintenance of gene repression but could also function in active transcriptional processes such as transcriptional activation of elongation. This is in agreement with previous reports and demonstrates that PRC2 has cellular functions that extend beyond what was learned from its role at Hox genes. Importantly, the current studies also suggest that despite a very good correlation of Jarid2 and Su(z)12 occupancies, Jarid2 might function in transcriptional repression and activation independently of the canonical PRC2 complex [E(z)] and vice versa. This distinction in target genes between Jarid2 and canonical PRC2 components [E(z)] provides additional confirmation that Jarid2 in some respects behaves fundamentally differently than the canonical PRC2 complex. Together with the varied functions proposed for Jarid2 in mammals, these studies highlight the diverse aspects of Jarid2 function in PRC2-mediated gene regulation (Herz, 2012).

Functions of Jarid2 orthologs in other species

Jarid2 and PRC2, partners in regulating gene expression

The Polycomb group proteins foster gene repression profiles required for proper development and unimpaired adulthood, and comprise the components of the Polycomb-Repressive Complex 2 (PRC2) including the histone H3 Lys 27 (H3K27) methyltransferase Ezh2. How mammalian PRC2 accesses chromatin is unclear. This study found Jarid2 associates with PRC2 and stimulates its enzymatic activity in vitro. Jarid2 contains a Jumonji C domain, but is devoid of detectable histone demethylase activity. Instead, its artificial recruitment to a promoter in vivo resulted in corecruitment of PRC2 with resultant increased levels of di- and trimethylation of H3K27 (H3K27me2/3). Jarid2 colocalizes with Ezh2 and MTF2, a homolog of Drosophila Pcl, at endogenous genes in embryonic stem (ES) cells. Jarid2 can bind DNA and its recruitment in ES cells is interdependent with that of PRC2, as Jarid2 knockdown reduced PRC2 at its target promoters, and ES cells devoid of the PRC2 component EED are deficient in Jarid2 promoter access. In addition to the well-documented defects in embryonic viability upon down-regulation of Jarid2, ES cell differentiation is impaired, as is Oct4 silencing (Li, 2010).

Since the first characterization of the PRC2 core complex, the subsequent, persuasive evidence supports that PRC2 is actually a family of complexes whose composition varies during development, as a function of cell type, or even from one promoter to another. This study identified two new components that interact with PRC2: MTF2 and Jarid2. These analyses of the proteins that interact with the PRC2 complex initiated with transformed cells. Yet it has become clear that interactions observed using transformed cells might be specific to such cells, and not a determinant to the integrity of a normal organism. Thus, studies of a developmentally relevant process was incorporated and it was confirmed that the interactions observed between PRC2 and Jarid2 were of consequence to the developmental program (Li, 2010).

MTF2 is a paralog of Drosophila Pcl. PHF1, another mammalian paralog of Pcl, is required for efficient H3K27me3 and gene silencing in HeLa cells. Although PHF1 appears dispensable for PRC2 recruitment in HeLa cells, work in Drosophila has suggested that the absence of Pcl could impair PRC2 gene targeting. It is possible that the other paralogs of Pcl (MTF2 and PHF19) exhibit a role that is partially redundant with PHF1 function and thereby maintain PRC2 recruitment upon its knockdown. Pcl and its mammalian paralogs contain two PHD domains and a tudor domain, domains reported to potentially recognize methylated histones. Although the ability of Pcl to specifically bind modified histone has not been elucidated to date, it is tempting to speculate that the PHD and tudor domains could target Pcl to specific chromatin regions. Its presence would then stabilize PRC2 recruitment and promote its enzymatic activity. In support of this hypothesis, it was observed that, whereas Ezh2 targeting is severely impaired in Eed-/- ES cells, MTF2 recruitment is affected in a promoter-dependent manner and to a lesser extent than that of Ezh2. This observation suggests that MTF2 gene targeting could be partially independent of PRC2 (Li, 2010).

The exact function of Jarid2 is more enigmatic. Indeed, Jarid2 is a member of a family of enzymes capable of demethylating histones. However, Jarid2 is devoid of the amino acids required for iron and αKG binding, and consequently is unable to catalyze this reaction. It is considered that Jarid2 could act as a dominant negative and inhibit the activity of other histone demethylases; however, coexpression of Jarid2 with, for instance, SMCX did not affect H3K4me3 demethylation. Jarid2 has two domains that could potentially bind DNA: the ARID domain and a zinc finger. Although the ARID domain of Jarid2 was reported to bind DNA, band shift assay suggests that other parts of the Jarid2 C terminus (potentially a zinc finger) are also important for binding to DNA. The SELEX experiment performed with the full-length Jarid2 did not allow identification of any sequence-specific DNA binding, but did result in a slight enrichment of GC-rich DNA sequences. Importantly, it was found that the N-terminal part of Jarid2 could robustly stimulate PRC2-Ezh2 enzymatic activity on nucleosomes. A knockdown of Jarid2 decreased the enrichment of PRC2 at its target genes. Conversely, overexpression of a Gal4-Jarid2 chimera recruited PRC2 at a stably integrated reporter and increased PRC2 enrichment at its target genes, supporting the hypothesis that Jarid2 contributes to PRC2 recruitment (Li, 2010).

In the case of Drosophila, PRE (Polycomb group response element) sequences have been described, and PRC2 access to chromatin is expected to involve the concerted action of several distinct and specific DNA-binding proteins that interact directly or indirectly with PRC2. However, these same DNA-binding factors, or even a combination thereof, are also found at active genes devoid of PRC2. What distinguishes PRE sequences harboring PRC2 from active genes is still not clear. During the evolution from Drosophila to mammals, only a few of the DNA-binding factors that bind PREs (Dsp1 and Pho) are conserved. Either PRC2 recruitment in mammals involves other mechanisms, or distinct transcription factors have emerged to stabilize PRC2 at its target genes. A recent study has identified a presumed mammalian PRE; however, the role of this putative PRE at the endogenous locus that is enriched for PRC2 is not reproduced when the element is integrated upstream of a transgene, as PRC2 is absent. Of note, whereas DNA-binding proteins are likely to play an important role for PRC2 recruitment in mammals, some studies have now suggested that long noncoding RNA could also be involved in this process. These observations together suggest that the recruitment of PRC2 to target genes is complex and requires more than one factor. These findings suggest that the DNA-binding activity of Jarid2 is one such factor, but its affinity for DNA is low and likely requires the help of other factors (Li, 2010).

A critical issue at this juncture is whether or not the composition of PRC2 changes during development. This study reports that Jarid2 interacts with PRC2, but its expression, unlike the PRC2 core components, seems to be restricted to some cell lines. In agreement with previous gene expression profiles that monitored mRNA levels during the reprogramming of mouse embryonic fibroblast cells into ES cells, it is observed that Jarid2 expression is higher in undifferentiated ES cells and decreases upon differentiation. Polycomb target genes are enriched with the H2A variant H2A.Z in undifferentiated ES cells; furthermore, H2A.Z and PRC2 targeting are interdependent in these cells. This result suggests that PRC2 recruitment might involve distinct mechanisms in ES cells and differentiated cells. It is possible that Jarid2 somehow contributes to this specificity (Li, 2010).

Knockdown of Jarid2 in undifferentiated ES cells does not give rise to an obvious phenotype; gene expression patterns appear to be only moderately affected, and cell proliferation is unchanged. In contrast, when cells are induced to differentiate, a process that entails dramatic changes in gene expression, impairments were observed as a function of Jarid2 knockdown. Interference with Jarid2 resulted in a failure to accurately coordinate the expression of genes required for the differentiation process, consistent with the previous report on Suz12 knockout cells. Instead of the requisite silencing of OCT4 and Nanog loci that occurs upon normal differentiation, each of which become enriched in H3K27me3, Jarid2 knockdown prevented such H3K27 methylation at these genes, and this correlated with their delayed repression. Thus, the Jumonji family of proteins that usually exhibits demethylase activity that might function in opposition to the role mediated by PRC2 contains the member Jarid2 that is devoid of such activity and instead facilitates the action of PRC2 through enabling its access to chromatin (Li, 2010).

Phosphorylation of the PRC2 component Ezh2 is cell cycle-regulated and up-regulates its binding to ncRNA

Ezh2 functions as a histone H3 Lys 27 (H3K27) methyltransferase when comprising the Polycomb-Repressive Complex 2 (PRC2). Trimethylation of H3K27 (H3K27me3) correlates with transcriptionally repressed chromatin. The means by which PRC2 targets specific chromatin regions is currently unclear, but noncoding RNAs (ncRNAs) have been shown to interact with PRC2 and may facilitate its recruitment to some target genes. This study, carried out in mammalian cells, shows that Ezh2 interacts with HOTAIR and Xist. Ezh2 is phosphorylated by cyclin-dependent kinase 1 (CDK1) at threonine residues 345 and 487 in a cell cycle-dependent manner. A phospho-mimic at residue 345 increased HOTAIR ncRNA binding to Ezh2, while the phospho-mimic at residue 487 was ineffectual. An Ezh2 domain comprising T345 was found to be important for binding to HOTAIR and the 5' end of Xist (Kaneko, 2010).

The results presented here demonstrate that PRC2 binding to HOTAIR (expressed from the HOXC cluster) and RepA (repeats found in Xist) ncRNAs is mediated through its Ezh2 component, and that phosphorylation of Ezh2-T345 up-regulates HOTAIR-binding activity. Given that phosphorylation at this site is cell cycle-regulated, it is speculated that PRC2 recruitment to chromatin, mediated through Ezh2 interaction with HOTAIR or RepA ncRNAs and presumably other ncRNAs, must be restricted to a tightly defined interval during the cell cycle (G2/M). Most importantly, the results establish that there are at least two populations of PRC2 complexes during the G2-M stages of the cell cycle. This is consistent with a model whereby PRC2 is recruited to specific genes to initiate repression as a function of its Ezh2 component being phosphorylated at T345, after which other PRC2 complexes then spread the repressing signature (H3K27me2/3). Of note, a recent study has also documented that human Ezh2 is phosphorylated at Thr 350 (murine T345) by CDK1, and, in agreement with the current findings, the report shows that this modification is ineffectual with respect to the integrity of the PRC2 complex and PRC2-mediated histone lysine methyltransferase activity. Instead, this study shows that mutant Ezh2 that is not subject to T350 phosphorylation results in down-regulated PRC2 recruitment, such that appropriate gene repression is thwarted. This report demonstrates that abrogation of this phosphorylation site within Ezh2 compromises Ezh2 interaction with ncRNAs, and this may bear directly on the mechanism by which PRC2 recruitment is impaired (Kaneko, 2010).

It has been demonstrated previously that the Eed component of PRC2 binds to trimethylated histone-repressive marks, but its binding to H3K27me3 in particular results in an allosteric effect that markedly increases the histone methyltransferase activity of its partner, Ezh2. Thus, PRC2 binding to the product of its activity increases its production of this mark. It is postulated that HOTAIR and RepA ncRNAs (and other ncRNAs) recruit PRC2 to initiate repression of target genes, and that this recruitment is enhanced by Ezh2 phosphorylation at T345. This proposed mechanism is consistent with only a small percentage of Ezh2 being phosphorylated at T345. If ncRNA species are responsible for targeting PRC2 to chromatin during G2/M, it is postulated that the recruited PRC2 would set the initial H3K27me3 mark. A larger number of PRC2 complexes, independent of their Ezh2 component being phosphorylated, would then propagate this mark upon their Eed component binding to the initial H3K27me3, with resultant allosteric activation of their Ezh2 activity (Kaneko, 2010).

An important question remaining is whether Ezh2 is the only component of PRC2 that binds to ncRNAs. These studies established that a 30-amino-acid domain of the PRC2-associated protein Jarid2 also binds to ncRNAs. Additionally, a recent study suggested that the Suz12 subunit of PRC2 binds to nascent transcripts. It was postulated that this binding results in the halting of RNA polymerase II. Interestingly, it was postulated that Suz12 binding to nascent transcripts requires a unique stem-loop structure on the RNA. This structure is similar to the structure of RepA, and computer analysis of the amino acid sequence of Suz12 revealed a putative domain at its N terminus with a predicted RNA-binding domain. These observations collectively suggest that PRC2 recruitment to its target genes is mediated by ncRNA, different species of which likely bind to different PRC2 subunits. Whether specificity or affinity of PRC2 for its target genes is regulated by PRC2 binding through its component(s) to one family of ncRNA (specificity), or whether multiple subunits of PRC2 simultaneously bind different ncRNAs or domains within a ncRNA (affinity), remains to be established. Regardless, the studies described here are beginning to shed light on the role of ncRNAs in mediating the recruitment of mammalian PRC2 to its target genes (Kaneko, 2010).

Histone H2A monoubiquitination promotes histone H3 methylation in Polycomb repression

A key step in gene repression by Polycomb is trimethylation of histone H3 K27 by PCR2 to form H3K27me3. H3K27me3 provides a binding surface for PRC1. This study shows that monoubiquitination of histone H2A by PRC1-type complexes to form H2Aub creates a binding site for Jarid2-Aebp2-containing PRC2 and promotes H3K27 trimethylation on H2Aub nucleosomes. Jarid2, Aebp2 and H2Aub thus constitute components of a positive feedback loop establishing H3K27me3 chromatin domains (Kalb, 2014).

Nucleosomes constitute the building blocks of eukaryotic chromosomes. They consist of a core of histone proteins around which DNA is wrapped in two helical turns. The post-translational modification of histones is a key step for the regulation of diverse processes that occur on nucleosomal DNA. Specific histone modifications often decorate arrays of nucleosomes that comprise many kilobases of DNA, but how such extended stretches of chromatin become modified is not well understood. A paradigm for a long-range chromatin-modification mechanism is transcriptional repression by Polycomb protein complexes. The Polycomb system generates two distinct histone modifications: methylation of K27 in histone H3 and monoubiquitination of K119 in histone H2A in vertebrates and of the corresponding K118 in Drosophila H2A. Polycomb repressive complex 2 (PRC2) catalyzes mono-, di- and trimethylation at H3 K27. At inactive Polycomb-target genes, H3 K27 trimethyl marks typically decorate nucleosomes across the entire upstream, promoter and coding region and are essential for repression of these genes. The H3K27me3 modification is recognized by Polycomb, a subunit of the canonical Polycomb repressive complex 1 (PRC1), and is thought to promote PRC1 interaction with chromatin across the entire length of repressed genes. PRC1 has been proposed to repress transcription through chromatin compaction and also through its ubiquitin-ligase activity for H2Amonoubiquitination. To gain insight into the function of H2Aub, this study set out to identify interactors of this modification (Kalb, 2014).

Arrays of four nucleosomes (referred to as oligonucleosomes) were reconstituted with recombinant Drosophila or Xenopus histones and monoubiquitinated H2A in these templates, using appropriate recombinant enzymes. Drosophila monoubiquitinated H2AK118 (H2AK118ub) oligonucleosomes and the corresponding unmodified oligonucleosome control template were used for affinity purification of H2AK118ub-binding proteins from Drosophila embryo nuclear extracts. In parallel, Xenopus monoubiquitinated H2A K119 (H2AK119ub) and unmodified control oligonucleosomes were used to identify vertebrate H2AK119ub interactors in nuclear extracts from mouse embryonic stem cells. In both experiments, quantitative MS analyses identified PRC2 subunits as being among the most highly enriched H2Aub interactors. Jarid2 and Aebp2 were the PRC2 subunits showing highest enrichment in both cases (Kalb, 2014).

The identification of PRC2 as an H2Aub interactor in both flies and vertebrates prompted an analysis of PRC2 histone methyltransferase (HMTase) activity on H2Aub nucleosomes. Recombinant human PRC2 containing EED, EZH2, SUZ12 and RBBP4 (referred to as PRC2) and assemblies of the same complex that in addition contained AEBP2 (AEBP2-PRC2), JARID2 (JARID2-PRC2) or both JARID2 and AEBP2 (JARID2-AEBP2-PRC2) were reconstituted. For substrates, Xenopus mononucleosomes were used that were either unmodified or monoubiquitinated at H2A K119, and in all cases western blot analyses were used with antibodies against either monomethylated H3 K27 (H3K27me1) or H3K27me3 to monitor PRC2 activity. A time-course experiment was performed to compare the activity of PRC2 and JARID2-AEBP2-PRC2 on H2A and H2Aub nucleosomes. It was found that, consistently with earlier reports, the catalytic activity of PRC2 alone is largely unchanged on H2Aub nucleosome templates. As expected, inclusion of JARID2 and AEBP2 in PRC2 resulted in stronger activity for H3 K27 methylation on unmodified nucleosome templates. However, a much stronger increase was used in H3K27me3 formation when JARID2-AEBP2-PRC2 was used for HMTase reactions on H2Aub nucleosomes. It was estimated that JARID2-AEBP2-PRC2 trimethylates H3K27 in H2Aub nucleosomes with an efficiency 25-fold higher than that of PRC2. To assess the contributions of JARID2 and AEBP2 to this stimulation of HMTase activity, the catalytic activity was compared of all four forms of PRC2 on H2A and H2Aub nucleosome substrates. JARID2-PRC2 showed higher H3K27 methyltransferase activity than did PRC2 on unmodified nucleosomes, as previously reported, but this was not further increased on H2Aub nucleosomes. In contrast, AEBP2-PRC2 methylated H3K27 in H2Aub nucleosomes with considerably higher efficiency than in unmodified nucleosomes. This suggests that AEBP2 is critical for the specific activation of PRC2 by H2Aub, whereas JARID2 has a more general function in boosting PRC2 HMTase activity, independently of the H2A modification state (Kalb, 2014).

The work reported in this study reveals that Jarid2-Aebp2-containing PRC2 binds to H2Aub nucleosomes and demonstrates that H3K27 trimethylation by this complex is strongly enhanced on H2Aub nucleosomes. This establishes H2Aub, Aebp2 and Jarid2 as components of a positive feedback loop in which H2Aub promotes PRC2 binding and H3K27 trimethylation, and H3K27me3 in turn promotes binding of the canonical PRC1 via the chromodomain of Polycomb. It is currently not clear whether canonical PRC1 indeed has E3 ligase activity for H2Amonoubiquitination or whether this modification is generated only by forms of PRC1 lacking Polycomb. Intriguingly, in embryonic stem cells, the PRC1-type complexes PRC1.1 and PRC1.6 were also identified as H2Aub interactors, results suggesting an additional feedback loop for H2A ubiquitination in vertebrates. The positive feedback loop for H3K27me3 formation by H2Aub uncovered in this study provides a rationale for how extended domains of Polycomb-repressed chromatin could be generated in both Drosophila and vertebrates. These findings could explain why H3K27me3 levels at Polycomb-target genes are reduced in mouse embryonic stem cells in which H2AK119ub has been depleted. However, it was previously found that bulk H3K27me3 levels were undiminished in late-stage Drosophila larvae in which bulk H2Aub levels had been depleted, thus suggesting that maintenance of H3K27me3-containing chromatin domains does not strictly depend on H2Aub. The H2Aub-mediated feedback loop may thus primarily be required for the initial formation of H3K27me3 chromatin domains when Polycomb repression is first established during the early stages of embryogenesis (Kalb, 2014).

Jarid2/Jumonji coordinates control of PRC2 enzymatic activity and target gene occupancy in pluripotent cells

Polycomb Repressive Complex 2 (PRC2) regulates key developmental genes in embryonic stem (ES) cells and during development. Jarid2/Jumonji (see Drosophila Little imaginal discs), a protein enriched in pluripotent cells and a founding member of the Jumonji C (JmjC) domain protein family, is a PRC2 subunit in ES cells. Genome-wide ChIP-seq analyses of Jarid2, Ezh2, and Suz12 binding reveal that Jarid2 and PRC2 occupy the same genomic regions. Jarid2 promotes PRC2 recruitment to the target genes while inhibiting PRC2 histone methyltransferase activity, suggesting that it acts as a 'molecular rheostat' that finely calibrates PRC2 functions at developmental genes. Using Xenopus laevis as a model, Jarid2 knockdown was shown to impair the induction of gastrulation genes in blastula embryos and results in failure of differentiation. These findings illuminate a mechanism of histone methylation regulation in pluripotent cells and during early cell-fate transitions (Peng, 2009).

Jarid2 and Jarid1a regions responsible for Suz12 binding do not overlap with any discernible structural domains and display low similarity, with the exception of a highly homologous short sequence 'GSGFP.' It is hypothesized that this motif may play a role in Suz12 recognition. Indeed, mutations of GSGFP to GAGAA diminished binding of Jarid2 and Jarid1a fragments to full-length Suz12. This motif is conserved in all vertebrate Jarid2 proteins, as well as in C. elegans Jarid2, whereas D. melanogaster and other Drosophila species contain a non-conservative substitution within the motif (GYGFP). The GSGFP motif is also conserved in all four Jarid1 family proteins: Jarid1a/RBP2, Jarid1b/PLU-1, Jarid1c/SMCX and Jarid1d/SMCY, as well as in the single Jarid1 homolog in Drosophila, Lid. The presence of the GSGFP motif in metazoan Jarid proteins suggests that the association with Suz12 may be a common feature of Jarid family members. However, the possibility that additional molecular interactions control Jarid-PRC2 complex formation in vivo cannot be excluded (Peng, 2009).

Erk1/2 activity promotes chromatin features and RNAPII phosphorylation at developmental promoters in mouse ESCs

Erk1/2 activation contributes to mouse ES cell pluripotency. This study found a direct role of Erk1/2 in modulating chromatin features required for regulated developmental gene expression. Erk2 binds to specific DNA sequence motifs typically accessed by Jarid2 and PRC2. Negating Erk1/2 activation leads to increased nucleosome occupancy and decreased occupancy of PRC2 and poised RNAPII at Erk2-PRC2-targeted developmental genes. Surprisingly, Erk2-PRC2-targeted genes are specifically devoid of TFIIH, known to phosphorylate RNA polymerase II (RNAPII) at serine-5, giving rise to its initiated form. Erk2 interacts with and phosphorylates RNAPII at its serine 5 residue, which is consistent with the presence of poised RNAPII as a function of Erk1/2 activation. These findings underscore a key role for Erk1/2 activation in promoting the primed status of developmental genes in mouse ES cells and suggest that the transcription complex at developmental genes is different than the complexes formed at other genes, offering alternative pathways of regulation (Tee, 2014).

Nucleosome-binding activities within JARID2 and EZH1 regulate the function of PRC2 on chromatin

Polycomb-repressive complex 2 (PRC2) comprises specific members of the Polycomb group of epigenetic modulators. PRC2 catalyzes methylation of histone H3 at Lys 27 (H3K27me3) through its Enhancer of zeste (Ezh) constituent, of which there are two mammalian homologs: Ezh1 and Ezh2. Several ancillary factors, including Jarid2, modulate PRC2 function, with Jarid2 facilitating its recruitment to target genes. Jarid2, like Ezh2, is present in poorly differentiated and actively dividing cells, while Ezh1 associates with PRC2 in all cells, including resting cells. Jarid2 was found to exhibit nucleosome-binding activity that contributes to PRC2 stimulation. Moreover, such nucleosome-binding activity is exhibited by PRC2 comprising Ezh1 (PRC2-Ezh1), in contrast to PRC2-Ezh2. The presence of Ezh1 helps to maintain PRC2 occupancy on its target genes in myoblasts where Jarid2 is not expressed. These findings lead to a model in which PRC2-Ezh2 is important for the de novo establishment of H3K27me3 in dividing cells, whereas PRC2-Ezh1 is required for its maintenance in resting cells (Son, 2013).

KDM4A coactivates E2F1 to regulate the PDK-dependent metabolic switch between mitochondrial oxidation and glycolysis

The histone lysine demethylase KDM4A/JMJD2A (see Drosophila Jumonji) has been implicated in prostate carcinogenesis through its role in transcriptional regulation. This study describes KDM4A as a E2F1 (see Drosophila E2F1) coactivator and demonstrate a functional role for the E2F1-KDM4A complex in the control of tumor metabolism. KDM4A associates with E2F1 on target gene promoters and enhances E2F1 chromatin binding and transcriptional activity, thereby modulating the transcriptional profile essential for cancer cell proliferation and survival. The pyruvate dehydrogenase kinases (PDKs; see Drosophila Pdk) PDK1 and PDK3 are direct targets of KDM4A and E2F1 and modulate the switch between glycolytic metabolism and mitochondrial oxidation. Downregulation of KDM4A leads to elevated activity of pyruvate dehydrogenase and mitochondrial oxidation, resulting in excessive accumulation of reactive oxygen species. The altered metabolic phenotypes can be partially rescued by ectopic expression of PDK1 and PDK3, indicating a KDM4A-dependent tumor metabolic regulation via PDK. These results suggest that KDM4A is a key regulator of tumor metabolism and a potential therapeutic target for prostate cancer (Wang, 2016).

JMJD-1.2/PHF8 controls axon guidance by regulating Hedgehog-like signaling

Components of the KDM7 family of histone demethylases are implicated in neuronal development and one member, PHF8, is also found mutated in cases of X-linked mental retardation. However, how PHF8 regulates neurodevelopmental processes and contributes to the disease is still largely missing. This study shows that the catalytic activity of a PHF8 homolog in Caenorhabditis elegans, JMJD-1.2 (see Drosophila Jarid2), is required non-cell autonomously for proper axon guidance. Loss of JMJD-1.2 deregulates the transcription of the Hedgehog-related genes wrt-8 and grl-16 whose overexpression is sufficient to induce the axonal defects. Deficiency of either wrt-8 or grl-16, or reduced expression of homologs of genes promoting Hedgehog signaling restore correct axon guidance in jmjd-1.2 mutant. Genetic and overexpression data indicate that Hedgehog-related genes act on axon guidance through actin remodelers. Thus, this study highlights a novel function of jmjd-1.2 in axon guidance that may be relevant for the onset of X-linked mental retardation and provides compelling evidences of a conserved function of the Hedgehog pathway in C. elegans axon migration (Riveiro, 2017).


Search PubMed for articles about Drosophila Jarid2

Herz, H. M., Mohan, M., Garrett, A. S., Miller, C., Casto, D., Zhang, Y., Seidel, C., Haug, J. S., Florens, L., Washburn, M. P., Yamaguchi, M., Shiekhattar, R. and Shilatifard, A. (2012). Polycomb repressive complex 2-dependent and -independent functions of Jarid2 in transcriptional regulation in Drosophila. Mol Cell Biol 32: 1683-1693. PubMed ID: 22354997

Jung, J., Mysliwiec, M. R. and Lee, Y. (2005). Roles of JUMONJI in mouse embryonic development. Dev Dyn 232: 21-32. PubMed ID: 15580614

Kalb, R., Latwiel, S., Baymaz, H. I., Jansen, P. W., Muller, C. W., Vermeulen, M., Muller, J. (2014) Histone H2A monoubiquitination promotes histone H3 methylation in Polycomb repression. Nat Struct Mol Biol 21: 569-571. PubMed ID: 24837194

Kaneko, S., Li, G., Son, J., Xu, C. F., Margueron, R., Neubert, T. A. and Reinberg, D. (2010). Phosphorylation of the PRC2 component Ezh2 is cell cycle-regulated and up-regulates its binding to ncRNA. Genes Dev 24: 2615-2620. PubMed ID: 21123648

Kaneko, S., et al. (2010). Phosphorylation of the PRC2 component Ezh2 is cell cycle-regulated and up-regulates its binding to ncRNA. Genes Dev. 24(23): 2615-20. PubMed ID: 21123648

Klose, R. J., Kallin, E. M. and Zhang, Y. (2006). JmjC-domain-containing proteins and histone demethylation. Nat Rev Genet 7: 715-727. PubMed ID: 16983801

Landeira, D., et al. (2010). Jarid2 is a PRC2 component in embryonic stem cells required for multi-lineage differentiation and recruitment of PRC1 and RNA Polymerase II to developmental regulators. Nat Cell Biol 12: 618-624. PubMed ID: 20473294

Landeira, D. and Fisher, A. G. (2011). Inactive yet indispensable: the tale of Jarid2. Trends Cell Biol 21: 74-80. PubMed ID: 21074441

Li, G., Margueron, R., Ku, M., Chambon, P., Bernstein, B. E. and Reinberg, D. (2010). Jarid2 and PRC2, partners in regulating gene expression. Genes Dev 24: 368-380. PubMed ID: 20123894

Pasini, D., Cloos, P. A., Walfridsson, J., Olsson, L., Bukowski, J. P., Johansen, J. V., Bak, M., Tommerup, N., Rappsilber, J. and Helin, K. (2010). JARID2 regulates binding of the Polycomb repressive complex 2 to target genes in ES cells. Nature 464: 306-310. PubMed ID: 20075857

Peng, J. C., Valouev, A., Swigut, T., Zhang, J., Zhao, Y., Sidow, A. and Wysocka, J. (2009). Jarid2/Jumonji coordinates control of PRC2 enzymatic activity and target gene occupancy in pluripotent cells. Cell 139: 1290-1302. PubMed ID: 20064375

Riveiro, A. R., Mariani, L., Malmberg, E., Amendola, P. G., Peltonen, J., Wong, G. and Salcini, A. E. (2017). JMJD-1.2/PHF8 controls axon guidance by regulating Hedgehog-like signaling. Development [Epub ahead of print]. PubMed ID: 28126843

Sasai, N., Kato, Y., Kimura, G., Takeuchi, T. and Yamaguchi, M. (2007). The Drosophila jumonji gene encodes a JmjC-containing nuclear protein that is required for metamorphosis. FEBS J 274: 6139-6151. PubMed ID: 17970746

Shen, X., Kim, W., Fujiwara, Y., Simon, M. D., Liu, Y., Mysliwiec, M. R., Yuan, G. C., Lee, Y. and Orkin, S. H. (2009). Jumonji modulates polycomb activity and self-renewal versus differentiation of stem cells. Cell 139: 1303-1314. PubMed ID: 20064376

Son, J., Shen, S. S., Margueron, R. and Reinberg, D. (2013). Nucleosome-binding activities within JARID2 and EZH1 regulate the function of PRC2 on chromatin. Genes Dev 27: 2663-2677. PubMed ID: 24352422

Tee, W. W., Shen, S. S., Oksuz, O., Narendra, V. and Reinberg, D. (2014). Erk1/2 activity promotes chromatin features and RNAPII phosphorylation at developmental promoters in mouse ESCs. Cell 156: 678-690. PubMed ID: 24529373

Wang, L. Y., Hung, C. L., Chen, Y. R., Yang, J. C., Wang, J., Campbell, M., Izumiya, Y., Chen, H. W., Wang, W. C., Ann, D. K. and Kung, H. J. (2016). KDM4A Coactivates E2F1 to Regulate the PDK-Dependent Metabolic Switch between Mitochondrial Oxidation and Glycolysis. Cell Rep 16: 3016-3027. PubMed ID: 27626669

Young, M. D., Willson, T. A., Wakefield, M. J., Trounson, E., Hilton, D. J., Blewitt, M. E., Oshlack, A. and Majewski, I. J. (2011). ChIP-seq analysis reveals distinct H3K27me3 profiles that correlate with transcriptional activity. Nucleic Acids Res 39: 7415-7427. PubMed ID: 21652639

Zhang, Z., et al. (2011). PRC2 complexes with JARID2, MTF2, and esPRC2p48 in ES cells to modulate ES cell pluripotency and somatic cell reprogramming. Stem Cells 29: 229-240. PubMed ID: 21732481

Biological Overview

date revised: 7 April 2014

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