Date of Award
Doctor of Philosophy (PhD)
Eukaryotic genomic DNA is packaged in the form of chromatin, which contains repeating nucleosomal units consisting of roughly two super-helical turns of DNA wrapped around an octamer of core histone proteins composed of four histone species: one histone H3/H4 tetramer and two histone H2A/H2B dimers. Histones are basic globular proteins rich in lysine and arginine residues, with unstructured N-terminal “tail” regions protruding outside the nucleosome structure, and structured “core” domains in the DNA-associated portion. Several core residues, and in particular arginines in H3 and H4, mediate key interactions between the histone octamer and DNA in forming the nucleosomal particle. Histone post-translational modifications (PTMs) lead to downstream effects indirectly by allowing or preventing docking of effector molecules, or directly by changing the intrinsic biophysical properties of local chromatin. To date, little has been done to study PTMs that lie outside of the unstructured tail domains of histones. I describe here the identification by mass spectrometry of a novel methylation site on histone H3, the asymmetric dimethylation of arginine 42 (H3R42me2a). H3R42 is conserved through evolution and is at the DNA entry/ exit position within the nucleosome core, with likely interactions with the DNA backbone. I show that methyltransferases CARM1 and PRMT6 methylate this residue in vitro and in vivo. Using chemically-defined “designer” histones I also show that methylation of H3R42 stimulates transcription in vitro from chromatinized templates. Using peptide pull down experiments combined with enzymatic assays I demonstrate that H3R42me2a prevents the stimulation of the histone deacetylase activity of the N-CoR co-repressive complex by impeding its binding to H3. Thus, H3R42 is a new histone methylation site with stimulating effects on transcription. I propose that methylation of basic histone residues at the DNA interface may be a general mechanism to disrupt histone:DNA interactions, with effects on downstream processes, including transcription.
Casadio, Fabio, "Discovery and Characterization of Methylation of Arginine 42 on Histone H3: A Novel Histone Modification with Positive Transcriptional Effects" (2013). Student Theses and Dissertations. 224.