Histone methyltransferases and demethylases possess key chromatin remodeling activities that are needed to control proper gene expression in eukaryotes. Importantly, the recent discovery of histone demethylases has revealed that histone methylation is a dynamic process and is likely to be regulated. How histone demethylases are regulated is not thoroughly understood and is still currently being investigated. It has been suggested that post-translational modifications may play a role but how modifications regulate histone demethylase activity or how the balance of histone methyltransferases and demethylases is maintained have not been fully explored.¹ To address these issues, we turned to S. cerevisiae as our model organism of choice. In S. cerevisiae, it was recently discovered that deletion of NOT4 leads to decreased global histone H3 lysine 4 (H3 K4) trimethylation. However, until our study, the mechanism by which Not4 was regulating histone H3 K4 trimethylation was not clear.^2,3 We determined that expression of Jhd2, a histone H3 K4 demethylase, in wild-type yeast cells is tightly regulated by Not4, which in turns allows for proper H3 K4 trimethylation and gene expression levels (Fig. 1A).⁴ How, then, does Not4 control Jhd2 protein levels and histone methylation? We demonstrate for the first time that Not4, a known E3 ubiquitin ligase, can polyubiquitinate Jhd2 and target it for degradation by the proteasome.⁴ Furthermore, in the absence of Not4 or upon proteasome inhibition, Jhd2 protein levels increase leading to demethylation of trimethylated H3 K4 even in the presence of the H3 K4 methyltransferase, Set1.⁴ This, in turn, results in decreased gene expression of the GMP Synthetase, GUA1. We also show that the human homolog of Jhd2, JARID1C, is ubiquitinated by human NOT4 in vitro, suggesting that this may be a conserved mechanism by which histone demethylase activity is regulated.⁴ Our study also raises additional questions to be considered and addressed.

The JARID1 family of histone demethylases contains human homologs of Jhd2, including JARID1A/RBP2, JARID1B/PLU-1, JARID1C/SMCX and JARID1D/SMCY, and has been linked to a wide variety of human disorders from X-linked mental retardation to breast cancer.¹² PLU-1/JARID1B shows restricted and low expression in most human adult tissues and is found to be highly upregulated in breast cancer.¹³ The mechanism behind PLU-1 overexpression in breast cancer is not yet known, but our study may shed some light on this observation. We show that the human demethylase JARID1C/SMCX can be polyubiquitinated in vitro by human NOT4.⁴ In addition, protein levels of human LSD1, a non-JARID1 family member histone demethylase, have been shown to be influenced by the proteasome, suggesting that ubiquitination and degradation of demethylases may be a common mechanism for regulating their activity.¹⁴ Because of these observations and the conserved nature of these enzymes, we are currently determining if members of the JARID1 family are ubiquitinated in vitro and in vivo. If this is the case, we predict that defects in ubiquitination and degradation of PLU-1/JARID1B by human NOT4 and/or the proteasome could be a mechanism behind increased PLU-1 expression in breast cancer (Fig. 1C). It is also quite possible that NOT4 expression itself is affected in human cancers which could explain why PLU-1 is upregulated. Finally, if regulating histone demethylases by NOT4 is conserved and the reason for various human cancers, this could lead to a new therapeutic target.