Understanding the regulatory DNA sequences are becoming increasingly important in understanding the way plants integrate signalling cues mediated through the actions of the transcription factors (TFs). This thesis presents an interdisciplinary investigations into regulatory elements found in the promoter regions of a model organism Arabidopsis thaliana. The intergenic DNA sequences are studied between sets of orthologous genes in A. thaliana and 3 other related species to uncover hundreds of evolutionary conserved noncoding sequences (CNSs). The CNSs are found to be more skewed towards the annotated transcription start sites (TSSs) and enriched in previously identified transcription factors binding motifs. Furthermore, the nucleosomes are predicted to have strong presence in the uncovered CNS than random intergenic sequences alone. Altogether the evidence presented in the thesis points to the functional nature of the CNSs. Then, the promoters of genes thought to be co-regulated together and transcriptionally active during infection with fungal pathogen Botrytis cinerea are experimentally tested for direct protein-DNA interaction using high-throughput Yeast One-Hybrid (Y1H) library screens against the TFs found in A. thaliana. The resulting predictions were further validated using pairwise Y1H screen to suggest potential common regulation by ORA59, PIF7, ESE1, At4g38900 and ERF14, and uncovering a complex gene regulatory network (GRN) associated with the tested genes. The promoter fragments together with the predictions from the Y1H screens were used in the computational analysis to establish transcription factor specific binding motifs. Some of the newly predicted motifs were mutated and tested again for altered binding of the associated TFs. Furthermore, in planta mutations of the TFs predicted to be interacting with the promoters of the genes in the Y1H screens were found to have significant impact on the susceptibility of A. thaliana to infection with B. cinerea, further informing gene regulatory network active in response to biotic stress.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:606149 |
| Date | January 2013 |
| Creators | Jironkin, Aleksey |
| Publisher | University of Warwick |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://wrap.warwick.ac.uk/60669/ |
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