Function of the MYND Domain and C-Terminal Region in Regulating the Subcellular Localization and Catalytic Activity of the SMYD Family Lysine Methyltransferase Set5

ABSTRACT

SMYD lysine methyltransferases target histones and nonhistone proteins for methylation and are critical regulators of muscle development and implicated in neoplastic transformation. They are characterized by a split catalytic SET domain and an intervening MYND zinc finger domain, as well as an extended C-terminal domain. Saccharomyces cerevisiae contains two SMYD proteins, Set5 and Set6, which share structural elements with the mammalian SMYD enzymes. Set5 is a histone H4 lysine 5, 8, and 12 methyltransferase, implicated in the regulation of stress responses and genome stability. While the SMYD proteins have diverse roles in cells, there are many gaps in our understanding of how these enzymes are regulated. Here, we performed mutational analysis of Set5, combined with phosphoproteomics, to identify regulatory mechanisms for its enzymatic activity and subcellular localization. Our results indicate that the MYND domain promotes Set5 chromatin association in cells and is required for its role in repressing subtelomeric genes. Phosphoproteomics revealed extensive phosphorylation of Set5, and phosphomimetic mutations enhance Set5 catalytic activity but diminish its ability to interact with chromatin in cells. These studies uncover multiple regions within Set5 that regulate its localization and activity and highlight potential avenues for understanding mechanisms controlling the diverse roles of SMYD enzymes.

KEYWORDS:

• chromatin
• histone methylation
• protein methylation
• protein phosphorylation
• yeast

ACKNOWLEDGMENTS

We acknowledge members of the Green lab and the UMBC Applied Molecular Biology Master’s Program for technical assistance and helpful discussions and comments on the manuscript.

This study was supported in part by National Institutes of Health (NIH) grants R01GM124342 and R21AG064507 to E.M.G. and 8 P41 GM103533 to J.R.Y.

The content of this study is solely the responsibility of the authors and does not necessarily represent the official views of the NIH.