Biochemical characterization of epigenomic markers

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The genetic information, which is carried by our genes and their regulatory elements, contribute 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. In fact, epigenetic information, which is defined as heritable information that do 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 cornerstone in unraveling 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. Among those covalent modifications, methylation of lysine residues is amongst the most prominent alteration of histone proteins. However, despite their critical roles in the nucleus, the mechanisms controlling histone substrates recognition and catalysis of protein lysine methyltransferases (PKMTs) require further investigation. Our recent findings that a specific plant PKMT selectively methylates histone variant histone H3.1 and not histone H3.3; two sub-classes of histone H3 known to impart specific properties to chromatin, have motivated us to further investigate the interplay between plant PKMTs and histone variants. Our endeavors to focus on plant PKMT is also motivated by recent findings showing that plant PKMTs play a pivotal role in plant floraison and growth; raising the hope that our findings may have important impact in agriculture. In this proposal, we intend to define the mechanistic basis underlying histone recognition by SET DOMAIN GROUP8 (SDG8) and SET DOMAIN GROUP2 (SDG2) using X-Ray crystallography and in vitro assays. In the next five years, we aim (Fig. 2) to 1) to determine the molecular basis controlling histone H3 recognition by SDG2 and SDG8, 2) determine the consensus amino acid sequences recognized by these proteins and 3), in collaboration with Dr. Yannick Jacob (Yale U.), test the impact of histone mutants targeting residues recognized by these enzymes in plants with the specific goal of identifying mutations conferring growth advantages, increasing floral initiation and improving defense mechanisms. 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.