Grant: Discovery Grants Program - Individual

Background: The genetic information, which is carried by our genes and their regulatory elements, contributes to the proper development of all living organisms. Despite the importance of this genetic code, an additional layer of information is also known to be vital. Epigenetic information, defined as heritable information that does not depend on the underlying DNA sequence, has recently surfaced as a master regulator of many biological processes. The discoveries that DNA scaffolding proteins, such as histones, and DNA itself could be chemically modified and that such covalent alterations could trigger specific molecular signals have been the cornerstone in unravelling a complex, intricate and new code: the histone code. Single or a combination of modifications is now known to carry critical information vital for many molecular events, including gene transcription, protein translation, DNA repair and many others. Histone lysine methylation was first characterized by Allfrey and colleagues in the 1960s. Since their seminal work, several methyl-lysine residues have characterized on histone proteins and many of the enzyme depositing this postranslational modification (PTM) are composed of Suvar (3-9), Enhancer of Zeste, Trithorax domain (SET) domain. Despite their critical roles in the nucleus, the mechanisms controlling histone substrates recognition and catalysis by SET domain lysine methyltransferases (PKMTs) require further investigation. In plants, Arabidopsis Trithorax-Related proteins 5/6 (ATXR5/6) monomethylate H3.1K27 and this PTM is linked to repression of gene expression and is enriched in constitutive heterochromatin in pericentromeric regions. In addition to their SET domains, ATXR5/6 share a plant homeodomain (PHD), a PCNA-interacting peptide (PIP) motif, and a nuclear localization signal (NLS). Published and preliminary data from our laboratory show that all these regions are critical to maintaining the catalytic activity of the enzymes; however, the structural underpinnings underlying their roles remain to be elucidated. Moreover, akin to several other SET domain PKMTs, ATXR5/6 preferentially methylates histone H3.1 when embedded in a nucleosome core particle (NCP), yet how these enzymes recognize the NCP is unknown. We will use ATXR5 as a model enzyme to shed some light on NCP recognition by SET domain PKMTs. The research supported by this grant will be pivotal in elucidating the molecular details eliciting the writing of the epigenome. In addition, this work will also expand our bases on how enzymes recognize their histone substrates and how enzymatic reactions are performed. This project is in line with our long-term goals of elucidating the molecular determinants controlling the covalent modification of histone proteins and the writing of the epigenome.