KDM5B protein expressed in viable and fertile ΔARID mice exhibit no demethylase activity

Abstract

Post-translational modification of histones serve a crucial role in the control of gene transcription. Trimethylation of lysine 4 on histone 3 is associated with transcription activation. There are currently six known methylases and six known demethylases that can control the methylation status of this site. Lysine demethylase 5B (KDM5B) is one such demethylase, which can repress gene expression. In particular KDM5B has been found to be overexpressed in a number of cancer types, and small-molecular weight inhibitors of its demethylase activity have been identified. Previous characterisation of Kdm5b knock-out mice has revealed that this genotype leads to either embryonic or neonatal lethality. However, the ΔA-T rich interaction domain (ΔARID)-KDM5B strain of mice, which have the ARID domain and five amino acids within the Jumonji (Jmj)N domain spliced out from KDM5B, remain viable and fertile. In the present study, ΔARID-KDM5B was found to have no demethylase activity as determined by in vitro demethylase assays and by immunofluorescence in transfected Cos-1 cells. Furthermore, molecular dynamic simulations revealed conformational changes within the ΔARID-KDM5B structure compared with that in WT-KDM5B, particularly in the JmjC domain, which is responsible for the catalytic activity of WT-KDM5B. This supports the experimental data that shows the loss of demethylase activity. Since Kdm5b knock-out mice show varying degrees of lethality, these data suggest that KDM5B serves a crucial function in development in a manner that is independent of its demethylase activity.

Histone Methylases and Demethylases Regulating Antagonistic Methyl Marks: Changes Occurring in Cancer

“…However, there is agreement that the PHD1 domain can influence the re-modelling of the catalytic core by binding to H3K4me0 [ 15 , 19 , 161 ]. Our own data from the ΔARID mouse expressing KDM5B with a deletion of the ARID domain, together with five amino acids from the JmjN domain (but with the PHD1 intact), show that this mutant has lost demethylase activity [ 85 , 86 ]. However, the fact that the ΔARID mouse is viable and fertile indicates that domains other than the demethylase activity are important in development and viability (See Table 2 and below).…”

“…Different phenotypes have been reported for KO of KDM5B in C57Bl/6 with Catchpole et al reporting early embryonic lethality [ 85 ], and Albert et al finding perinatal death due to respiratory failure, resulting from neurological abnormalities [ 166 ]. Since the ΔARID mouse, expressing a demethylase null mutant of KDM5B, is viable and fertile [ 85 , 86 ], it follows that functions other than demethylase activity are important in development. The ΔARID mouse provides an appropriate model for examining the effect of the loss of KDM5B enzyme activity on neural activity and behaviour.…”

“…It is possible that oncogenic effects which are independent of de-methylase activity may present opportunities for selective inhibition of the individ-ual KDM5 proteins. An important question is how important the functioning of the KDM5 demethylase activity in the adult brain is, in the context of inhibiting this function in cancer patients [ 86 , 152 , 211 ]. A comparison of behaviour in WT mice and the ΔARID mice expressing a demethylase-null KDM5B could give some answers.…”

A truncated and catalytically inactive isoform of KDM5B histone demethylase accumulates in breast cancer cells and regulates H3K4 tri-methylation and gene expression

“…Since the protein is correctly localized to the nucleus, questions arise about its biological roles. Recently, Taylor-Papadimitru et al [ 44 ] studied a strain of mice expressing a KDM5B splicing variant that lacked the ARID domain and five amino acids in the JmjN domain. They demonstrated that, although the protein is catalytically inactive, the mouse is viable and fertile, contrary to the KDM5B knockout mouse, which displays an embryonic lethal phenotype [ 45 ].…”

KDM5 family as therapeutic targets in breast cancer: Pathogenesis and therapeutic opportunities and challenges

“…Compared to full-length KDM5A, ARID-deficient KDM5A exhibits reduced demethylation capacity of H3K4me3 possibly because this mutation alters spatial arrangements or global folding [ 66 ]. Molecular dynamics simulations reveal that the JmjC domain of ARID-deficient KDM5B is more flexible and can induce protein structure dynamic changes compared to full-length KDM5B [ 67 ]. Additionally, KDM5A achieves transcriptional regulation of specific genes through the binding of ARID to the CCGCCC motif [ 66 ].…”