Chromatin dynamics and genomic instability during replicative aging

Histones are the fundamental protein subunit of chromatin and have a profound impact on processes such as chromatin assembly/disassembly and diseases such as aging. I examined the functional importance of mutating the histone H4 C-terminal tail, which is normally involved in chromatin assembly and disassembly by studying the in vivo and in vitro consequences of mutating a critical residue, histone H4 gycine 94 to a proline residue (H4 G94P) in budding yeast. Substitution of the gycine residue for a proline was used to restrict the conformational flexibility of the C-terminal tail of histone H4. The H4 C-terminal tail makes direct contacts with the histone chaperone Anti-silencing function 1 (Asf1) and undergoes a dramatic 180° conformational change in the opposite direction of the contacts it makes with histones in the nucleosome. I found that in addition to the role the H4 C-terminal tail has in chromatin assembly, it also has additional roles in maintaining octamer and nucleosome stability as well as yeast viability.;The loss of chromatin integrity and the histone density play an important role in the aging process. A more open chromatin structure during aging is caused by the loss of histone density from the DNA in yeast. I examined whether there was increased genomic instability during replicative aging as well as the types of damage that might contribute to the aging process. I found that the DNA damage checkpoint was activated during aging and that most of the damage detected via gamma-H2A ChIP-sequencing, accumulates at the rDNA locus and the mitochondria during replicative aging. Additionally, I found that Ty retrotransposons, which are normally silenced by chromatin have increased transcription and increased Ty genomic DNA during aging. The increase in Ty gene transcription and genomic DNA coincided with an increase in Ty retrotransposition, which contributes to genomic instability during aging. In summary, genomic instability increases during replicative aging and is mediated in part via the integration of Ty retrotansposons, which are normally silenced by chromatin.