The circadian clock is the endogenous mechanism by which a wide variety of biological processes are regulated in anticipation of daily changes in the external environment. In Arabidopsis thaliana, the clock comprises a number of complex gene and protein interactions, involving multiple regulatory feedback loops. The clock gene LHY has a central role in these loops, activating and repressing morning- and evening- expressed genes respectively. These clock genes in turn sequentially repress the expression of LHY throughout the day and night, restricting it to a sharp transcriptional peak at dawn. However, the molecular mechanisms of these regulatory interactions with the LHY promoter were not known. Therefore, this project first aimed to determine which promoter motifs are responsible for mediating regulation of LHY circadian expression. This was achieved through luciferase assays with mutated pLHY:LUC reporter constructs, which identified the CT-rich region as responsible for rhythmic expression of LHY, and the G-box as mediating regulation by the clock protein TOC1. Since few regulators were known to target the LHY promoter, this project also aimed to identify transcription factors binding the promoter using a Yeast One-Hybrid assay. Transcription factors with roles in a wide variety of biological pathways were identified from this screen, with abiotic stress and plant defence pathways particularly well-represented. In addition, a number of antagonistic and synergistic regulatory interactions were established as occurring between stress factors and clock proteins at specific promoter motifs. We can therefore conclude that LHY is regulated by a complex network of transcription factor interactions, enabling the rapid integration of environmental stress signals into the clock.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:582377 |
| Date | January 2013 |
| Creators | Davies, Siân Elizabeth Wynne |
| Publisher | University of Warwick |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://wrap.warwick.ac.uk/57320/ |
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