Mechanisms of Polycomb group protein function in cancer

Abstract

Cancer arises from a multitude of disorders resulting in loss of differentiation and a stem cell-like phenotype characterized by uncontrolled growth. Polycomb Group (PcG) proteins are members of multiprotein complexes that are highly conserved throughout evolution. Historically, they have been described as essential for maintaining epigenetic cellular memory by locking homeotic genes in a transcriptionally repressed state. What was initially thought to be a function restricted to a few target genes, subsequently turned out to be of much broader relevance, since the main role of PcG complexes is to ensure a dynamically choregraphed spatio-temporal regulation of their numerous target genes during development. Their ability to modify chromatin landscapes and refine the expression of master genes controlling major switches in cellular decisions under physiological conditions is often misregulated in tumors. Surprisingly, their functional implication in the initiation and progression of cancer may be either dependent on Polycomb complexes, or specific for a subunit that acts independently of other PcG members. In this review, we describe how misregulated Polycomb proteins play a pleiotropic role in cancer by altering a broad spectrum of biological processes such as the proliferation-differentiation balance, metabolism and the immune response, all of which are crucial in tumor progression. We also illustrate how interfering with PcG functions can provide a powerful strategy to counter tumor progression.

Fig. 1. Composition of PcG proteins in mammals.

a PRC2 can be sub-divided into PRC2.1 and PRC2.2. The core PRC2 with PCL1/2/3 and the PALI1/2 or EPOP subunits compose the PRC2.1 complex. The association between the core PRC2, JARID2 and AEBP2 constitute PRC2.2. b PRC1 complex can be sub-divided into two groups of complexes, namely cPRC1 and ncPRC1. cPRC1 is composed of RING1A/B associated with PCGF2/4 and CBX2/4/6–8. ncPRC1 is composed of either RYBP or YAF2 associated with one of the six PCGF proteins. The graphical representation of each complex is schematic and not aimed to represent the size, shape and relative position of the various subunits. AEBP2, Adipocyte enhancer-binding protein 2; AUTS2, Autism susceptibility candidate 2; BCOR, BCL6 corepressor; CBX2/4/6–8, Chromobox 2/4/6–8; CK2, Casein kinase 2; DCAF7, DDB1 and CUL4 associated factor 7; DP1, E2F dimerization partner 2; E2F6, E2F transcription factor 6; EED, Embryonic ectoderm development; EPOP, Elongin B/C and PRC2-associated protein; EZH1/2, Enhancer of zeste homolog 1/2; FBRS, Fibrosin; HDAC1/2, Histone deacetylase 1/2; HP1, Heterochromatin protein 1 gamma (here labeled as HP1, also named CBX3); JARID2, Jumonji AT rich interactive domain 2; KDM2B, Lysine demethylase 2B; L3MBTL2, Lethal(3) malignant brain tumor-like protein 2; MAX, Myc associated factor X; MGA, MAX gene associated protein; PALI1/2, PRC2 associated LCOR isoform 1/2; PCGF1–6, Polycomb group finger 1–6; PCL1–3, Polycomb like protein 1–3; PHC1–3, Polyhomeotic-like protein 1–3; RBBP4/7, Retinoblastoma binding protein 4/7; RING1A/B, Really interesting new gene 1B/A; RYBP, RING1 and YY1 binding protein; SCMH1/2, Sex comb on midleg homolog 1/2; SKP1, S-phase kinase associated protein 1; SUZ12, Suppressor of zeste 12 protein homolog; USP7, Ubiquitin specific peptidase 7; YAF2, YY1-associated factor 2; WDR5, WD repeat domain 5.

Fig. 2. Polycomb recruitment and action on target genes.

a First described in Drosophila melanogaster, the original pathway of PcG recruitment relies on two sequential steps. First, PRC2 is recruited to chromatin and deposits the repressive H3K27me3 mark via its EZH1/2 subunit. The repressive mark is then recognized by the CBX2/4/6–8 chromodomain, a subunit of cPRC1. Lastly, RING1A/B deposit the ubiquitination on H2AK119 (in Drosophila, H2AK118). While in Drosophila melanogaster PRC2 recruitment depends on specific transcription factors binding to PREs, in mammals PRC2 recruitment can occur at CGIs or depends on transcription factors or lncRNAs. More recent data suggest an alternative recruitment pathway, in which ncPRC1 complexes are recruited in a KDM2B-dependent manner which deposits the H2AK119 ubiquitination mark. In turn, this mark is recognized by the JARID2 subunit of PRC2.2. Furthermore, PRC2.1 binds the same targets via PCL1/2/3 proteins. Finally, cPRC1 is recruited via CBX2/4/6–8-mediated recognition of H3K27me3. Moreover, the new PcG proteins BAHD1 and BAHD2 have also been found to recognize the H3K27me3 repressive mark. Their interactions with HDACs generate a hypoacetylated chromatin state which participates in transcriptional silencing. b Chromatin compaction impairs the transcription of target genes. c PcG-mediated silencing depends on the inhibition of the transcriptional machinery while the repressive PRC2 and PRC1 marks are necessary to inhibit the deposition of active histone marks. d PRC1 participates in the higher-order 3D chromatin organization via its PHC subunits. The SAM domain of PHC-PRC1 is able to oligomerize which results in the maintenance of the transcriptionally repressed state. BAHD1, Bromo adjacent homology domain containing 1; CBP, CREB binding protein; HDAC, Histone deacetylase; RNA Pol II, RNA Polymerase II; SWI/SNF, Switch/sucrose non-fermentable.

Fig. 3. Multifaceted roles of PRC2 in tumorigenesis.

a Upregulation of PRC2 components results in H3K27 hypermethylation, which, if present in tumor suppressor genes, induces their downregulation. In contrast, downregulation of PRC2 components at oncogenes leads to H3K27 hypomethylation and a switch to acetylation, contributing to the overexpression of specific oncogenes. b GOF mutations (indicated by a star) affecting the SET-domain of EZH2 can lead to overactivation of its H3K27 methyltransferase catalytic activity and to the silencing of tumor suppressor genes. c PTMs of EZH2 participate in tumorigenesis. Left: methylation of K307 of EZH2 by SMYD2 enhances its stability, resulting in a H3K27 hypermethylated state of tumor suppressor genes. Right: on the other hand, methylation of its K735 causes EZH2 degradation. The loss of EZH2 induces the replacement of H3K27me3 by H3K27ac, leading to the transcriptional expression of oncogenes. d Polycomb-independent roles of EZH2 in transcriptional activation. The gene encoding the AR is a direct target of EZH2-mediated transcriptional activation in Androgen-Dependent and Castration-Resistant Prostate Cancers (ADPC and CRPC, respectively). This mechanism is methylation-independent and escapes EZH2 inhibitors. In CRPC, EZH2 acts as a co-factor of AR. This functional transition of EZH2 from a role of repression to a role of activation of transcription depends on its phosphorylation at the level of Ser21. EZH2 and AR directly interact. This interaction inhibits the degradation of the AR and causes the overexpression of the AR target genes. e Under physiological conditions, PRC2 participates in the transcriptional repression of its HOX target genes throughout development. However, oncogenic transformation can redirect PRC2 to new target genes. This PRC2 redistribution, in particular at differentiation-related genes, induces a loss of differentiation and participates in the generation of a pluripotent stem cell-like phenotype. AR, Androgen Receptor; CBP, CREB binding protein; PSA, Prostate-Specific Antigen; SETD2, SET domain-containing 2 (a histone lysine methyltransferase); SMYD2, SET and MYND domain-containing 2.

Fig. 4. Multifaceted roles of PRC1 in tumorigenesis.

a PCGF2 inhibits the transcription of c-myc. Loss of c-Myc results in the decrease of PCGF4 expression, and in the derepression of PCG4 target genes, such as the INK4a-ARF locus. p19 and p16 participate in proliferation control, respectively, by inhibiting MDM2-mediated degradation of p53 and inhibiting CycD/CDK4-mediated phosphorylation of pRb. b PRC1 oncogenic activity may also be PRC2-independent. PRC1 is found on specific targets lacking the H3K27me3 repressive mark. Surprisingly, these genes exhibit active marks such as H3K27Ac and H3K4me1/3. Gene ontology analysis characterized these cancer-related genes as components of cell signaling, like the Notch and JAK/STAT signaling pathways. c PRC1 mutations are rarely found in cancer, although some mutations have been found to impact variant PRC1. Indeed, mutations (indicated by a star) in BCOR, a scaffold protein involved in ncPRC1.1, are found in SHH-driven medulloblastoma. The presence of these mutations promotes a neoplastic state of cancer cells by preventing Polycomb recruitment to its target genes. d PTM of PRC1 subunits can promote tumorigenesis. The deposition of O-GlcNAcylation on PCGF4 (BMI-1) inhibits its degradation. PCGF4 protein levels are increased and participate in the transcriptional silencing of downstream target genes such as the INK4a-ARF locus, thus promoting oncogenic cell proliferation. e In hormone-dependent cancers, PRC1 genes are often amplified. Top: in prostate cancer, the AR promotes the expression of PCGF4. Additionally, it can interact with the PCGF4 protein, resulting in inhibition of AR degradation and transcriptional activation of its downstream target genes. Bottom: cPRC1 can also interact with the ER and its pioneer factor FOXA1 in ER⁺ breast cancer cells and bind to enhancers that stimulate transcription of cancer-related genes decorated with active histone marks. AR, Androgen Receptor; Cdk4, 6, Cyclin Dependent Kinase 4, 6; ER, Estrogen Receptor; FOXA1, Forkhead Box A1; Igf2, Insulin-like growth factor 2; MDM2, Murine Double Minute 2; PSA, Prostate Specific Antigen; Rb, Retinoblastoma.

Fig. 5. Environment-dependent oncogenic activities of PcG proteins.

a Left: in a physiological condition, the membrane transporter LAT1 participates in the transport of methionine which reacts with ATP to produce SAM. SAM can in turn be used by PRC2 to induce trimethylation of H3K27, resulting in a PcG-mediated silencing of its targets genes. Lat1 expression depends on RXRα. Right: in cancer cells, Lat1 is overexpressed, enhancing SAM production and inducing H3K27 hypermethylation of the chromatin landscape. The Lat1 negative regulator, RXRα, is thus repressed resulting in a positive feedback loop whereby EAF2 transcriptional silencing dependent on PRC2 results in overexpression of HIF1, which can in turn stimulate Lat1 expression. Therefore, an excess of LAT1 at the cellular membrane increases the transport of BCAAs, thereby enhancing protein synthesis. b Controlling the immune system is of a major importance in cancer. Cancer cells use different mechanisms to do this. First, PRC1 is able to increase the transcriptional expression of CCL2, which will dampen Treg immune response. In addition, PRC2-mediated silencing of the MHC-I antigen processing pathway results in MHC-I absence at the cell membrane, concealing cancer cells from cytotoxic T cells. Finally, PCGF4 overexpression in cancer cells stimulates the expression of GATA2, which will inhibit MICA/B transcription and reduces its presence at the membrane. This prevents the recognition of cancer cells by NK cells. These mechanisms enhance the immunosuppressive response and inhibit the cytotoxic response that would otherwise kill the cancer cells. c Oncohistones are a new line of research, analyzing the effect of mutations on histone genes that could have an impact on tumorigenesis. H3K27M has a dominant negative effect on EZH2 catalytic activity. Left: in a wild-type condition, PRC2 is recruited to nucleation sites that present unmethylated CGIs. Trimethylation of H3K27 occurs and spreads around the nucleation site. The boundaries of Polycomb domains are decorated with H3K36me2. Right: in the presence of the H3K27M oncohistone, that represents 10% of all H3, an epigenetic remodeling occurs. The spreading of H3K27me3 is inhibited and active histone marks, such as H3K27ac, are present on the oncohistone. BCAAs, Branched-chain amino acids; CCL2, C-C motif chemokine ligand 2; CCR2, C-C motif chemokine receptor; EAF2, ELL associated factor 2; GATA2, GATA binding protein 2; HIF1, Hypoxia inducible factor 1; IDH1, Isocitrate dehydrogenase 1; KDM6A/B, Lysine demethylase 6A/B; LAT1, L-type amino acid transporter 1; MHC-I, Major histocompatibility complex I; MHC-I APP, Major histocompatibility complex I antigen processing pathway; MICA/B, MHC I polypeptide-related sequence A/B; RXRα, Retinoid X receptor-alpha; SAM, S-adenosylmethionine; SAH, S-adenosylhomocysteine; TCR, T-cell receptor; SETD2, SET domain containing 2 (histone lysine methyltransferase).