Throughout the course of our lives we are continuously exposed to a variety of agents that can damage the genetic material, DNA, within our cells. It has been estimated that a typical human cell must repair over 10,000 DNA lesions per day (Lindahl, T. Nature, 1993). Failure to repair such damage can have catastrophic cellular consequences, leading to genome in-stability, cancer, or cell death (Hoeijmakers, J.H. Nature, 2001). To insure their genetic integrity cells employ a coordinated response to DNA damage that includes checkpoint mediated cell cycle arrest and damage repair (Sancar et al, Annu Rev Biochem, 2004). Though many molecular factors and elegant mechanisms of repair have been defined, an understanding as to how the DNA damage response machinery functions in the chromatin environment is lacking. The native genetic template is not a solitary unit of DNA but rather a tightly associated DNA-protein complex termed chromatin. The packaging of DNA into chromatin is is inherently inhibitory to DNA-dependent activities such as transcription and DNA repair.  The foundation of chromatin is formed from the nucleosome, an octamer of the four core histones H3, H4, H2A, and H2B wrapped by two turns of DNA (Khorasanizadeh, S. Cell, 2004). One mechanism employed by cells to overcome the chromatin barrier is to mark histones by enzymatic post-translational modification. Each of the core histones is subject to a variety of modifications, such as phosphorylation, acetylation, and methylation, and many chromatin regulatory factors contain intrinsic catalytic activities responsible for these modifications. It has been argued that the distinct modifications form a “histone code” that is read by the chromatin regulatory machinery (Strahl and Allis, Nature, 2000). This modification code appears to function as a signaling platform that extends the indexing potential of chromatin. Much of our knowledge concerning histone-modifying enzymes relates to their role in controlling gene expression. Our understanding as to how these modifiers also facilitate the repair of damaged DNA is limited (Peterson and Cote, Genes Dev, 2004). Addressing this deficiency is of vital importance as mounting evidence argues deregulation of histone modifying enzymes may play a central role during oncogenesis. 

The objective of the laboratory is to decipher how histone modifying enzymes function in DNA damage response. We are particularly interested in those factors that have been conserved throughout eukaryotic evolution from yeast to man. This allows us to utilize the genetic and biochemical power of model yeast organisms to rapidly dissect regulatory pathways that have implications for human disease. Utilizing the fission yeast Schizosacchromyces pombe we have recently identified an enzyme, Set9, that methylates lysine (K) 20 of histone H4 (Sanders et al, Cell 2004). Our results indicate that methyl H4-K20 functions as a histone mark to target the checkpoint protein Crb2 to sites of DNA damage.  Surprisingly the modification itself does not specifically mark sites of DNA damage.  Rather H4-K20 methylation appears to be present throughout the genome even in the absence of DNA damage but is inaccessible to Crb2 in "undamaged" chromatin.  Structural studies argue that the histone H4 tail, particularly residues surrounding H4-K20, plays an essential roll in mediating nucleosome-nucleosome interactions in packed chromatin (Schalch et al, Nature, 2005; Dorigo et al, Science, 2004; Dorigo et al, J Mol Biol, 2003).  This argues that Crb2 cannot access its required target as the H4-K20 methyl modification is "buried" in the context in the context of stacked chromatin.  Introduction of a double strand break (DSB) generates a region of unstacked or open chromatin, exposing the pre-existing methyl H4-K20 residue that can be recognized by Crb2. Recruitment of Crb2 to DSBs is essential to maintaining DNA damaged checkpoint induced cell cycle arrest. In the absence of H4-K20 methylation Crb2 localization to DSBs is severally impaired and cells cannot maintain cell cycle arrest after damage.  Thus the methyl histone H4-K20 modification controls a novel genome surveillance pathway. Crb2's mammalian equivalent, 53BP1, is also targeted to DSBs in a similar fashion, though a different methyl modification may be recognized (Huyen et al, Nature, 2004).