Regulation of p53 and Rb Links the Alternative NF-κB Pathway to EZH2 Expression and Cell Senescence

Abstract

There are two major pathways leading to induction of NF-κB subunits. The classical (or canonical) pathway typically leads to the induction of RelA or c-Rel containing complexes, and involves the degradation of IκBα in a manner dependent on IκB kinase (IKK) β and the IKK regulatory subunit NEMO. The alternative (or non-canonical) pathway, involves the inducible processing of p100 to p52, leading to the induction of NF-κB2(p52)/RelB containing complexes, and is dependent on IKKα and NF-κB inducing kinase (NIK). Here we demonstrate that in primary human fibroblasts, the alternative NF-κB pathway subunits NF-κB2 and RelB have multiple, but distinct, effects on the expression of key regulators of the cell cycle, reactive oxygen species (ROS) generation and protein stability. Specifically, following siRNA knockdown, quantitative PCR, western blot analyses and chromatin immunoprecipitation (ChIP) show that NF-κB2 regulates the expression of CDK4 and CDK6, while RelB, through the regulation of genes such as PSMA5 and ANAPC1, regulates the stability of p21WAF1 and the tumour suppressor p53. These combine to regulate the activity of the retinoblastoma protein, Rb, leading to induction of polycomb protein EZH2 expression. Moreover, our ChIP analysis demonstrates that EZH2 is also a direct NF-κB target gene. Microarray analysis revealed that in fibroblasts, EZH2 antagonizes a subset of p53 target genes previously associated with the senescent cell phenotype, including DEK and RacGAP1. We show that this pathway provides the major route of crosstalk between the alternative NF-κB pathway and p53, a consequence of which is to suppress cell senescence. Importantly, we find that activation of NF-κB also induces EZH2 expression in CD40L stimulated cells from Chronic Lymphocytic Leukemia patients. We therefore propose that this pathway provides a mechanism through which microenvironment induced NF-κB can inhibit tumor suppressor function and promote tumorigenesis.

The Ubiquitin-like with PHD and Ring Finger Domains 1 (UHRF1)/DNA Methyltransferase 1 (DNMT1) Axis Is a Primary Regulator of Cell Senescence

“…EZH2 reportedly recruits DNMT1 to target genes, thereby mediating promoter methylation (23). However, EZH2 also antagonizes a subset of p53-regulated genes associated with cell senescence (24), implying its irrelevance to direct senescence induction. Chk1 recruits DNMT1 to DNA damage sites (25) and directs both senescence and mitotic catastrophe in HCT116 human colorectal cancer cells (26).…”

Nicotinamide metabolism regulates glioblastoma stem cell maintenance

“…Taken together, core regulators in NAM metabolism, NNMT and NAMPT, are negative prognostic factors for human glioma patients. (91,92); NF-κB complex ChIP-seq data from previously published studies (93,94). (C) Motif analysis of enhancer regions of upregulated genes of the nicotinamide and nicotinate metabolism pathway (NNMT, NAMPT, MAT2A, AHCY, BST1, and BST2).…”

Mutation Spectra of Histone Methyltransferases With Canonical SET Domains and EZH2-Targeted Therapy

“…The regulation and function of EZH2 as well as the development of EZH2 inhibitors and EZH2-targeted therapy based on synthetic lethal strategy will be described below, and perspectives on EZH2-related pathophysiology and EZH2-targeted therapy will also be provided. 10 [59][60][61] (Figure 2A). EZH2 mRNA is transcriptionally repressed directly by NFIB [62] and indirectly via the TP53-CDKN1A axis [63] and is also post-transcriptionally repressed by miRNAs, such as miR-25, miR-26a, miR-30d, miR-98, miR-101 and miR-214 (Figure 2A) [64][65][66].…”