Structure and function of MDM2 and MDM4 in health and disease

Abstract

Both mouse double-minute 2 (MDM2), an E3 ubiquitin ligase, and its closely related paralog, MDM4, which lacks E3 activity, play central roles in cellular homeostasis. MDM-linked dysfunction is associated with an increased risk of oncogenesis, primarily through targeting the tumor suppressor protein p53 for ubiquitination and degradation. Recent studies have revealed multifaceted roles of MDM proteins that are p53 independent with implications for their oncogenic properties. This review aims to provide an overview of MDM2 and MDM4, by assessing gene and protein structure and implications for protein-protein interactions and functions in cell and animal physiology. We also explore MDM2 and MDM4 role(s) in angiogenesis, a critical feature of solid tumor growth and progression. Finally, we discuss the current landscape in the development of MDM2 and MDM4 inhibitors for cancer therapy.

Keywords: 26S proteasome; E3 ubiquitin ligase; MDM2; MDM4; cancer; p53.

Figure 1:. Ubiquitination pathway overview.

Schematic representation of the ubiquitination pathway involving E1 ubiquitin-activating, E2 ubiquitin-conjugating and E3 ubiquitin-ligase enzymes. Ubiquitin is activated by an E1 enzyme in an ATP-dependent manner before being transferred to an E2 enzyme. Then, the ubiquitin is transferred directly onto the substrate, catalyzed by RING-type E3 ubiquitin ligase; or indirectly by other types of E3 enzymes (not shown). Ubiquitination can lead to monoubiquitination, the attachment of a single ubiquitin molecule or polyubiquitination, where a chain of ubiquitin molecules is formed (created with BioRender.com). Abbreviation: RING, Really Interesting New Gene.

Figure 2:. MDM2 and MDM4 genes and proteins.

Graphical representation of the chromosomal location, gene structure with exon organization, and protein structures of MDM2 and MDM4. The two promoters of each gene are shown as arrows. Abbreviations: NES, nuclear export signal; NLS:, nuclear localization signal; RING, Really Interesting New Gene.;NES: nuclear export signal.

Figure 3:. MDM2 and MDM4 protein isoforms.

MDM mRNA splice variants encoding protein isoforms are depicted underneath MDM2 and MDM4 genes. Numbers indicate the different exons within the MDM2 and MDM4 genes. MDM2-FL:, full-length MDM2; MDM4-FL:, full-length MDM4.

Figure 4:. Structural models of MDM2-p53 interactions.

(A) MDM2 (aquamarine) bound to the p53 peptide (pink) (PDB ID: 4HFZ)., (B) MDM4 (yellow) bound to the p53 peptide (sand) (PDB ID: 3DAB),. and (C) sSuperimposed structures. Key p53 interacting residues, Phe19, Trp23 and Leu26, isare shown and highlighted in red. Structures generated using PyMOL (www.pymol.org). MDM2, mouse double-minute 2; MDM4, mouse double-minute 4.

Figure 5:. Mechanisms of p53 regulation by MDM2 and MDM4 proteins.

MDM2 and MDM4 can directly bind and inhibit p53-mediated activation of tumor suppressor genes. MDM2 homodimers and MDM2-MDM4 heterodimers facilitate p53 proteasomal degradation and nuclear export depending on the type of ubiquitin linkage. Under genotoxic stress, MDM2 and MDM4 can act as internal ribosomal entry sites (IRES) trans-acting factors to promote p53 protein production by interaction with TP53 mRNA. p53 activation can activate the transcriptional activation of MDM2 (created using BioRender.com). MDM2, mouse double-minute 2; MDM4, mouse double-minute 4.

Figure 6:. Interaction between MDM2 and p53 in hypoxia response.

This schematic illustrates the regulatory network involving p53, MDM2 and downstream effectors during the cellular response to hypoxia. MDM2 modulates p53 stability and activity via ubiquitination. Hypoxia-inducible factor 1-alpha (HIF-1α) is stabilized under low oxygen conditions and can be up-regulated by MDM2 expression. HIF-1α suppresses MDM2-mediated inhibition of p53, thereby promoting p53 accumulation during hypoxia which in turn inhibits HIF-1α. HIF-1α promotes the expression of vascular endothelial growth factor A (VEGF-A), a key mediator in angiogenesis. Under acute hypoxia, p53 can directly increase VEGF-A levels, whereas prolonged hypoxic condition led to p53-mediated inhibition of VEGF-A via the activation of p21 (created using BioRender.com). MDM2, mouse double-minute 2.

Figure 7:. Binding of small molecule inhibitors and stapled peptides to MDM2 and MDM4.

(A) Chemical and X-ray co-crystal structure of Nutlin-3 (PDB ID: 4J3E), idasanutlin (RG7388; PDB ID: 4JRG), siremadlin (HDM201; PDB ID: 5OC8), alrizomadlin (APG-115; PDB ID: 5TRF), Nnavtemadlin (AMG-232/KRT-232; Ppredicted using CB-DOCK2 [131]), bound to MDM2. MDM2 -binding pockets are labelled by p53 side chain in yellow. (B) X-ray co-crystal structure of ALRN-6924 (PDB ID: 8GJS) bound to MDM4. MDM4 is shown in grey and ALRN-6924 is shown in aquamarine. The hydrocarbon staple is shown as sticks in pink. Structures generated using PyMOL (www.pymol.org). MDM2, mouse double-minute 2; MDM4, mouse double-minute 4.