Untersuchung des Zusammenhanges zwischen der Funktion pflanzlicher microRNAs und der DNA-Methylierung des PHABULOSA Locus in Arabidopsis thaliana

本研究旨在阐明拟南芥PHABULOSA（PHB）基因座甲基化的生物学功能。PHB是拟南芥叶器官极性形成的关键转录因子。最近已经表明，PHB信使RNA（mRNA）的水平由microRNA 165/166控制（McConnell等人，2001; Rhoades等人，2002;马洛里等人，2004），并且微小RNA/mRNA相互作用是微小RNA互补位点下游的PHB基因座内编码区甲基化所必需的（Bao et al.，2004年）。尽管已知siRNA指导同源DNA序列的甲基化（Mette等人，2000），DMA的甲基化是迄今为止尚未归因于microRNA的功能。此外，参与microRNA依赖性甲基化过程的机制似乎与负责siRNA介导的甲基化的机制不同，因为序列与miRNA不同源并且位于更远的下游。这指出了microRNA能够顺式指导染色体修饰的新特征。我们目前还不了解这种编码序列甲基化在PHB基因座的生物学作用。然而，在从富含干细胞的组织制备的DNA样品中观察到较低的下游甲基化水平，表明甲基化可能是发育控制的过程。我们的目标是解开生物学作用，并确定参与甲基化过程的蛋白质，希望能更深入地了解microRNA如何控制重要的发育过程。

This proposal aims to unravel the biological function of genie methylation observed at the PHABULOSA (PHB) locus in Arabidopsis thaliana. PHB is a key transcription factor required for the establishment of organ polarity in the Arabidopsis leaf. Recently it has been shown that the levels of PHB messenger RNA (mRNA) are controlled by microRNA165/166 (McConnell et al., 2001; Rhoades et al.f 2002; Mallory et al., 2004) and that microRNA/mRNA-interaction is required for methylation of a coding region within the PHB locus downstream of the microRNA complementary site (Bao et al., 2004). Although siRNAs are known to direct methylation of homologous DMA sequences (Mette et al., 2000), methylation of DMA is a function which has so far not been ascribed to microRNAs. Furthermore the machinery involved in the microRNA dependent methylation process appears to be different from that responsible for siRNA-mediated methylation as the sequences are not homologous to the miRNA and are more distantly located downstream. This points towards a novel feature of microRNAs being able to direct chromosomal modification in cis. We do not currently understand the biological role of this coding sequence methylation at the PHB locus. However, lower downstream methylation levels were observed in DNA samples made from tissues enriched for stem cells indicating that the methylation may be a developmentally controlled process. Our aim is to unravel the biological role and identify proteins involved in the methylation process hoping to gain a deeper understanding of how microRNAs control important developmental processes.