DNA methylation-dependent patterns of histone post-translational modification
EMSL Project ID
30404
Abstract
In eukaryotes, chromosomal DNA does not exist as a naked molecule, but is complexed with proteins to form chromatin. As a first step in chromatin formation, 167 bp of DNA is wrapped around a histone octamer made up of two molecules each of the core histone proteins H2A, H2B, H3 and H4, resulting in a bead-like structure known as a nucleosome core particle. The histones can be post-translationally modified by acetylation, methylation, phosphorylation or ubiquitination, thereby positively or negatively modulating the expression of associated genes, depending on the context of the modifications. Likewise, the DNA can be modified by extensive cytosine methylation, which typically correlates with gene silencing in mammals and plants. Genetic and biochemical evidence suggests that there is crosstalk between repressive histone modifications and cytosine methylation, however the full spectrum of histone modifications that go hand-in-hand with different patterns of cytosine methylation have not been defined. We propose to use state-of-the-art mass spectrometry techniques available at EMSL to analyze histones isolated from Arabidopsis thaliana mutants in which specific DNA methyltransferases or key histone modifying enzymes are nonfunctional. The goal of these studies is to define the suites of histone post-translational modifications that correlate with different patterns of cytosine methylation. These patterns include replication-associated maintenance methylation occurring at CG and CHG motifs, accomplished by the cytosine methyltransferases MET1 and CMT3, respectively, as well as de novo methylation of cytosines in any sequence context, which is accomplished by DRM2. Available evidence indicates that DRM2 methylation is directed by 24 nt small interfering RNAs (siRNAs) that guide the methylation to homologous sequences. Therefore mutants that disrupt 24 nt siRNA production and RNA-directed DNA methylation will also be analyzed to determine their effects on histone post-translational modifications. Through these studies, we expect to document the patterns of histone post-translational modifications that result from cross-talk with the DNA modifying machinery.Project Details
Project type
Large-Scale EMSL Research
Start Date
2008-08-20
End Date
2011-09-30
Status
Closed
Released Data Link
Team
Principal Investigator