|Institution:||University of Oslo|
|Full text PDF:||https://www.duo.uio.no/handle/10852/11465
Epigenetic gene regulation is crucial in developmental processes, controlling which genes that are expressed at what times and in what tissues. Posttranslational modifications on histone tail residues yield various outcomes depending on the modification pattern, leading to an open or more compact structure of the chromatin, and thus to activation or repression of transcription. The idea that modifications “written” by some enzymes influence each other and are “read” by other enzymes to specify distinct chromatin states, is called the histone code hypothesis. In this thesis, the three SET domain proteins SUVR1, SUVR2 and SUVR4, possessing a domain that has been found to have histone methyltransferase activity, were investigated. SUVR4 has been thoroughly examined in that regard, and found to repress transposable elements. Previous research has also indicated that SUVR4 is a H3K9 dimethylase with H3K9me1 as its preferred substrate, a modification mark abundant on transposon chromatin. To change activity from dimethylation to trimethylation, SUVR4 seems to be dependent on the signalling molecule ubiquitin in vitro. For ubiquitin binding, another domain, WIYLD, has been found important. In this master project, the WIYLD domain was mutated, reducing ubiquitin binding, and transposon H3K9 methylation levels were investigated. In lines overexpressing SUVR4 with mutated WIYLD domain, ChIP indicated that there was more monomethylation and less trimethylation compared to lines overexpressing wild type SUVR4, suggesting that ubiquitin binding is important for H3K9 trimethylation by SUVR4. DNA methylation is also linked to repression of transposable elements known to be regulated by SUVR4, involving the plant-specific polymerases IV and V. In this project, knock-out lines of polymerase IV subunits overexpressing wild type SUVR4 were investigated to find out if SUVR4 is connected to the function of polymerase IV. The knock-out of polymerase IV subunit genes led to less H3K9 trimethylation on specific transposon chromatin, suggesting that SUVR4 is dependent on the function of polymerase IV to be able to trimethylate H3K9. The investigation of SUVR1 and SUVR2 indicated that they too can bind ubiquitin, arguing that they may have a function similar to that of SUVR4. However, they possess a domain (MET) absent in SUVR4, suggesting a possible difference in chromatin substrate preference. In previous, unpublished experiments, overexpression of SUVR1 and SUVR2 have been shown to lead to early flowering, indicating a function in gene regulation related to genes involved in regulating flowering time.