Abstract
The order of genes in a eukaryotic genome has classically been considered random. However, large-scale genomics and transcriptomics studies as well as the frequent discovery of gene clusters have established that eukaryotic gene order is far from random. Indeed, there is increasing realisation that the positioning of genes influences expression patterns. For example, a characteristic feature of gene clusters, groups of neighbouring and functionally related genes, is the co-ordinate regulation of the clustered genes.Here, using the model organism Arabidopsis thaliana and its specialised metabolic gene clusters we have investigated the importance of genomic integrity in gene cluster regulation and aimed to identify key regulatory factors that lead to its co-regulation. By exploiting T-DNA mutant lines with disrupted linear and, putatively, the 3D cluster
organisation, we have analysed how these disruptions change the expression profile of an exemplar gene cluster. Furthermore, we have applied genome editing technologies to re-localise a cluster gene to assess if the native genomic context is important for gene regulation. We have applied bioinformatic data analyses, carried out mutant screens and quantitative gene expression analyses to unveil novel chromatin regulatory enzymes of gene clusters. Moreover, we have investigated sequence and expression conservation of metabolic gene clusters across A. thaliana ecotypes.
Our results show that the genomic integrity of the entire cluster space as well as its neighbouring genomic sites is important for cluster regulation. We present preliminary evidence that changing the genomic environment of a gene cluster results in the drastic misregulation of the gene. In addition, we have identified the chromatin marks H3K4me3, H3K18ac, H3K36me3 and H3K9me2 as putative epigenetic regulators of metabolic gene clusters in A. thaliana and discovered novel chromatin mutant lines with a role in gene cluster regulation. Our pilot evolutionary analyses indicates that the expression of one gene within the cluster can alter the expression profile of the remaining genes. Collectively, the work presented here has provided novel insights into the coordinate regulation of metabolic gene clusters and the importance of genomic integrity and chromatin environment. It forms a foundation for future research that will design synthetic gene clusters, further advancing our understanding of transgenesis, and unveiling the evolutionary principles of gene cluster formation.
| Date of Award | 26 Jul 2023 |
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| Original language | English |
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| Supervisor | Hans-Wilhelm Nützmann (Supervisor), Volkan Cevik (Supervisor) & Adele Murrell (Supervisor) |