AbstractThe primary level of chromatin organisation consists of arrays of nucleosomes that are present across the genetic template. Advances in the post genomics era have made it possible to determine the positions of nucleosomes genome-wide where it has been observed that nucleosomes adopt a distinct organisation with respect to genetic and trans-binding elements. Amongst the best studied of these is the transcription start site where it has been observed that genic nucleosome locations are well maintained with respect to promoters.
DNA and chromatin replication are coupled processes whereby chromatin is disrupted ahead of the replication fork and nucleosomes are rapidly assembled on the nascent DNA template. Classically it has been observed that nascent chromatin is more susceptible to digestion, prompting the possibility of an “immature” chromatin organisation post assembly. However it has not been investigated genome-wide if nucleosomes are initially assembled in phase or must be reorganised post assembly to canonical locations.
We have developed two methods of isolating nascent DNA fragments representative of nucleosome positions from synchronised and asynchronous populations of the budding yeast S. cerevesiae. High throughput sequencing has revealed that chromatin is assembled and organised rapidly behind the replication fork. The most nascent chromatin isolated displays typical patterns of nucleosome organisation suggesting that reorganisation of nucleosomes on the nascent template is replication coupled. Deletion of specific histone chaperones and chromatin remodelers perturbs this pathway. However, this can be compensated for by a transcription directed reorganisation of nascent chromatin. Our analysis of nascent chromatin has allowed us to investigate the mechanisms that act to direct chromatin organisation in addition to evaluation of models that describe nucleosome organisation genome-wide.
|Date of Award||2014|
|Supervisor||Tom Owen-Hughes (Supervisor)|
Chromatin Dynamics During DNA Replication
Fennessy, R. (Author). 2014
Student thesis: Doctoral Thesis › Doctor of Philosophy