Effections Of Histone Special Sites Methylation Changes On Transcription Functions Of Complete And The Second Structure Domain Flaw ELP3

AIM: The yeast elongator complex is composed of six subunits, as a complex possessing a activity of histone acetyltransferase (histone acetyltransferase, HAT), facilitates hyperphosphorylated RNAP II to involve in the process of SSA3 and GAL1 transcription elongation. Elp3 is the catalytic subunit of highly conserved histone acetyltransferase Elongator complex, The subunit possesses a C-terminal histone acetyltransferase (HAT) domain, has a second catalytic domain in the N-terminus—might be involved in catalyzing histone demethylation. Most previous studies have indicated that Elp3 as a histone acetyltransferase, facilitates hyperphosphorylated RNAP II to involve in the process of SSA3 and GAL1 transcription elongation.Previous studies have indicated that there is probably a relationship between Radical SAM superfamily in the N-terminus of Elp3 and methylation. However, some studies indicated that this domain related to complex formation had no catalytic activity. Now we are not sure Elp3 has demethylation activity that bioinformatics speculate. It is not clear that what is the relationship between the transcriptional functions of elongator and histone methylation.In order to understand whether complete and second structure domain flaw Elp3 participate in transcription elongation of genes when histone methylation, make sure the relationship between transcription function of Elp3 and histone Specific sites methylation. In this thesis, to confirm the effection of the transcription function of complete and the second structure domain flaw Elp3 through chromatin immune precipitation experiment when particular histone methylation site (H4K20,H3K79,H4K20/H3K79) was mutated. These studies are important to Elongator for establishing yeast epigenetic-based pertaining to participate in regulating gene expression model, further research on the function of elongator, explore its function abnormalities related disease occurrence mechanism.METHODS: Based on Elp3 antibodies, perform chromatin immunoprecipitation on Elp3 Series plasmid shift to histone mutations elp3Δstrains, PCR of special primer made by Input DNA and precipitation DNA template, confirm the effection of the transcription function of complete and second structure domain flaw Elp3 when particular histone methylation site was mutated.CONCLUSIONS: 1. Chromatin immunoprecipitation experiments indicates: In SSA3 gene transcription process, effections of histone special sites methylation changes on transcription functions of complete and the second structure domain flaw Elp3 is different, the recruiting qualitity of the second structure domain flaw Elp3 is less than that of complete Elp3, it shows that the second structure domain is necessary for the regualar function of Elp3.2. Chromatin immunoprecipitation experiments indicates: In GAL1 gene transcription process, effections of histone special sites methylation changes on transcription functions of complete and the second structure domain flaw Elp3 is different, although yElp3 and hElp3 are highly conservative on the functions, still have differences, the recruiting qualitity of the second structure domain flaw Elp3 is less than that of complete Elp3, it shows that the second structure domain is necessary for the regualar function of Elp3.