RNA-directed DNA methylation in plant development
Abstract
In higher plants, organ development occurs mostly post-embryonically. Organ initiation requires tightly controlled cell division patterning and cell fate reprogramming. Auxin, a key plant hormone, plays a crucial role in these processes, ultimately through significantly altering the gene expression landscape. On the other hand, gene expression can also be defined by manipulating locus accessibility, both on protein- (i.e. nucleosomes, histones and their modifications) as well as on DNA level: methylation status of cytosines in coding genes, mainly in their promoter regions, severely affects their expression rate and regulation. The interplay between auxin signaling and DNA methylation is poorly understood and we are planning to address this question in our research. We will characterize “freak show” (fks), a novel missense mutant of NRPE5, a subunit of RNA Polymerase V (Pol V) involved in RNA-directed DNA Methylation (RdDM). We isolated fks in a forward genetic screen aimed at identifying auxin-dependent determinants of lateral root development. fks is absolutely unique among all known Pol V mutants as it shows a series of very strong phenotypic traits in its inbred progenies such as organogenesis defects, dwarfism, sterility, homeotic organ transformations or delayed flowering time. Several of these features had already been described as potentially caused by epialleles, i.e. alterations of DNA methylation at target loci. However, to our knowledge, this is the first example that a mutant in de novo DNA methylation displays such a diverse palette of epigenetic features. Characterization of fks will help us, besides gaining new knowledge about auxin action, to understand how Pol V functions and how RdDM can be directed or regulated on a general level, with insight into developmental and physiological processes regulated by this machinery. We will use next generation sequencing (NGS) for determining the transcriptome and methylome of fks. To couple auxin-dependent gene expression and DNA methylation, we will combine auxin treatment with the NGS analysis. Besides, we will identify epialleles causing aberrant lateral root development and other selected traits with physical mapping, when and where feasible. We will perform double mutant analyses to investigate genetically whether the fks mutation specifically affects any of the known RdDM sub-pathways and whether it interferes with DNA methylation maintenance too. We will obtain triple mutants of NRPE5 and its two homologs of unknown function to further elaborate their role in RdDM and development. Additionally, we will address the function of NRPE5 and its homologs biochemically: protein interaction assays with other RNA polymerase subunits will tell us about possible subunit composition and –affinity of Pol V.
keywords epigenetics resistance
Publikationen
Project staff
Joseph Strauss
Univ.Prof. Mag.rer.nat. Dr.rer.nat. Joseph Strauss
joseph.strauss@boku.ac.at
Tel: +43 1 47654-94420
BOKU Project Leader
01.04.2018 - 30.06.2021