Functions and Interactions of Mammalian KDM5 Demethylases

Abstract

Mammalian histone demethylases of the KDM5 family are mediators of gene expression dynamics during developmental, cellular differentiation, and other nuclear processes. They belong to the large group of JmjC domain containing, 2-oxoglutarate (2-OG) dependent oxygenases and target methylated lysine 4 of histone H3 (H3K4me1/2/3), an epigenetic mark associated with active transcription. In recent years, KDM5 demethylases have gained increasing attention due to their misregulation in many cancer entities and are intensively explored as therapeutic targets. Despite these implications, the molecular basis of KDM5 function has so far remained only poorly understood. Little is known about mechanisms of nucleosome recognition, the recruitment to genomic targets, as well as the local regulation of demethylase activity. Experimental evidence suggests close physical and functional interactions with epigenetic regulators such as histone deacetylase (HDAC) containing complexes, as well as the retinoblastoma protein (RB). To understand the regulation of KDM5 proteins in the context of chromatin, these interactions have to be taken into account. Here, we review the current state of knowledge on KDM5 function, with a particular emphasis on molecular interactions and their potential implications. We will discuss and outline open questions that need to be addressed to better understand histone demethylation and potential demethylation-independent functions of KDM5s. Addressing these questions will increase our understanding of histone demethylation and allow us to develop strategies to target individual KDM5 enzymes in specific biological and disease contexts.

Keywords: JmjC oxygenases; KDM5; epigenetics; gene regulation; histone demethylation.

FIGURE 1

(A) KDM5 demethylases remove methyl groups from H3K4 in a sequential manner, using the dioxygenase activity of their catalytic JmjC domain. 2-Oxoglutarate (2-OG) is decarboxylated to succinate. Formaldehyde, one of the products of demethylation, is commonly detected in quantitative assays of JmjC demethylase activity. (B) Domain organization of the four human KDM5 demethylases and the Drosophila KDM5 homolog Lid. ZF = C5HC2 Zinc Finger. Numbers correspond to the amino acid numbering of each KDM5 protein.

FIGURE 2

Structures of KDM5B. The catalytic cores of KDM5 enzymes are structurally highly similar, therefore only structures of KDM5B are shown. (A) Atomic model of the catalytic core of KDM5B [PDB 5A1F (Johansson et al., 2016)]. The construct crystallized was composed of the JmjN and JmjC domains, as well as the α-helical domain including the C5HC2 Zn finger. The ARID and PHD1 domains were not included. The α-helical domain and the C5HC2 Zn-finger are required for demethylase activity, whereas the ARID and PHD1 domains are dispensable for the demethylation of peptide substrates by truncated KDM5s. (B) Alphafold2 prediction of full-length KDM5B (AF-Q9UGL1-F1), showing the predicted arrangement of the protein domains C-terminal of the catalytic core in an extended conformation, in agreement with experimental data (Dorosz et al., 2019). Of note, other conformations cannot be excluded due to the flexibility of the coiled-coil domain. Structural predictions of other KDM5s show a more compact orientation of the C-terminal part, with the PHD2 domain being located in close proximity to the N-terminal, catalytic core. Unstructured regions with low prediction confidence were omitted from the figure for clarity.

FIGURE 3

(A) Schematic representation of KDM5 interactions with chromatin. The catalytic composite JmjN/C domain binds the substrate H3 tail harboring methylated H3K4 (depicted as H3K4me3 for simplicity). Two of the PHD domains, PHD1 and 3, were shown to interact with unmethylated and trimethylated H3K4, respectively. The ARID domains are known DNA binding domains with a role in KDM5 chromatin targeting. How DNA binding is mediated in the context of full-length KDM5 proteins is currently unknown, since the arrangement of JmjC and ARID domains seems to be incompatible with the binding of nucleosomal DNA, according to homology models (Horton et al., 2016). The roles and potential chromatin interactions of the C5HC2 and PHD2 Zn fingers are unknown. Note that the depiction of three nucleosomes was chosen for clarity. It is not known how many nucleosomes are bound by a single KDM5 protein simultaneously. (B) Functional KDM5 interactions on chromatin. So far, only the binding of unmethylated H3K4 has been shown to regulate the demethylase activity of KDM5s. Given the potential interactions with HDACs, a direct or indirect responsiveness to other histone PTMs such as acetylated lysines, is conceivable. KDM5 proteins are recruited by transcription factors (TFs), reader domain proteins, or mediated by the association with other epigenetic regulators such as HDAC complexes or PRC2. The interaction and functional interplay of KDM5s with HDAC complexes and PRC2 suggests a potential mutual regulation of demethylase and other chromatin modifying activities. Such a direct interplay remains to be demonstrated experimentally.

FIGURE 4

KDM5 demethylases were shown to physically and functionally interact with HDAC complexes. Interactions with the NuRD and SIN3B complexes have been shown for the mammalian KDM5A and B proteins, as well as Drosophila Lid. CoREST interactions were shown for KDM5C, with implications for neuronal development. Note that, for reasons of clarity, the stoichiometry and detailed subunit composition of the complexes was neglected. For NuRD and SIN3B, composition and dynamics of subunits are subject to research and have not been definitely established. The placement of subunits and their proximity to each other and to the KDM5 proteins does not reflect experimentally verified proximity within the respective complexes.