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Title: Histone modification and its role in epigenetic regulation
Author: Amar, Sabrina
ISNI:       0000 0004 2686 1212
Awarding Body: University of Portsmouth
Current Institution: University of Portsmouth
Date of Award: 2010
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The development of genome-wide histone modification mapping technologies has provided a significant body of evidence implicating histone modifications in gene regulation, cell specialisation and differentiation processes. Monomethylation of lysine 4 of histone H3 (H3K4Mel) and the presence of the histone variant H2A.Z have previously been shown as a feature of enhancers, defined simply as points enriched in the HAT p300, but it was unclear whether these potential enhancers were linked to inactive, active or poised genes or whether developmental changes were reflected in the occupancy of H3K4Mel. The aim of this study was to document the spatio-temporal distribution of H3K4MeI at developmentally regulated genes and investigate proteins specifically binding to this modification so as to unravel possible control mechanisms. H3K4Mel distributions were determined by native chromatin immunoprecipitation (nChlP) experiments, analysed by real-time PCR, using chicken haematopoietic cell lines representing different erythroid and myeloid differentiation stages. Mapping at the lysozyme locus showed enrichments of H3K4Mel at the gene enhancers in myeloid cells independently of expression status, even in multipotent myeloid progenitor cells where the gene is not expressed. In contrast, in none of the erythroid cells studied - where the gene is never expressed - is there any H3K4Mel found at the lysozyme locus. At the β-globin locus, the β-adult enhancer is strongly enriched in H3K4mel in early erythroid progenitor cells where the globin genes are not expressed but the modification is lost from this enhancer in IS-day embryonic erythrocytes - which strongly express the adult and hatching globin genes - despite a general rise in H3K4Mel levels elsewhere in the locus, including the inactive embryonic globin genes. In none of the myeloid cells studied - where globin genes are never expressed - is there any high H3K4Mel enrichment found at the β-globin locus. At the folate receptor gene, the promoter carries a high level of H3K4Mel when the gene is inactive but this decreases when the gene becomes active. Taken together, the data suggest that H3K4Mel is a pioneer modification that participates in establishing the enhancers of genes poised for future activation. Pull-down experiments using immobilised peptides or reconstituted nucleosomes containing H3K4Mei and extracts from HeLa cell nuclei, followed by tryptic mass spectroscopy of bound proteins, were used to identify possible binding partners. Five potential binding partners were defined and have been shown to associate with chromatin.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available