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Title: Chromatin structure of the pea plastocyanin gene
Author: Chua, Y. L.
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2001
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The pea plastocyanin gene (PetE) is a single-copy, nuclear photosynthesis gene. Pea PetE is flanked by an enhancer/5' matrix attachment region (MAR) and a 3' MAR. When linked upstream to uidA directed by the CaMV 35S promoter, the enhancer/5' MAR increased reporter gene expression in transgenic tobacco plants. In contrast, the 3' MAR increased expression only when linked downstream of the reporter gene. The 3' MAR, but not the 5' MAR, decreased variation in reporter gene expression. These results indicate that the two MARs surrounding PetE have different effects on transgene expression. The chromatin structure of PetE was examined at three different transcriptional states by investigating the nuclease accessibility of the gene in pea roots, etiolated shoots and green shoots. Time-course digestions of nuclei with micrococcal nuclease and DNaseI indicated that the enhancer/5' MAR and promoter regions were more resistant to digestion in the inactive gene in pea roots than the same regions in the active gene in shoots, whereas the transcribed region of PetE was digested similarly amongst the tissues. PetE transcription is hence accompanied by changes in the nuclease accessibility of the enhancer/5' MAR and promoter regions only. The acetylation states of histone H3 and H4 proteins associated with PetE were analysed by chromatin immunoprecipitation with antibodies specific for acetylated or non-acetylated histone tails followed by polymerase chain reaction quantification. Comparison of pea tissue indicated that histone acetylation was associated with increased PetE transcription in green shoots. Moreover, acetylation of both histone H3 and H4 proteins was targeted to the enhancer/5' MAR and promoter regions in green shoots, suggesting that only specific nucleosomes along the gene were modified.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available