Acetylation and methylation sites in histone H4 from Physarum polycephalum

Abstract

Histone H4 has been isolated and purified from plasmodia of Physarum polycephalum. The four major fragments produced by hydrolysis of H4 by acetic acid were separated and the complete amino acid sequence of two of them was determined. By analogy with calf H4, these peptides are at the C-terminus and give the sequence from residue 68 to the C-terminus (residue 102). In this 35 residue sequence there are two minor differences from calf H4:(i) residue 77 is arginine in Physarum H 4 and lysine in calf H4; (ii) lysine-79 is partially methylated in Physarum. Arginine occurs at position 77 in pea H 4 but the occurrence of methylated lysine at position 79 has not been reported for other species. In the N-terminal region, amino acid compositions of acetic acid, tryptic and chymotryptic peptides indicate that Physarum H 4 and calf H4 have identical sequences from the N-terminus to residue 47. There may be minor differences in the region from residue 46 to residue 67. The sites of acetylation were determined by Edman degradation of acetate-labelled peptide 4 -17 of Physarum H4. Acetylation was observed at positions 5,8, 12, and 16. The only other labelled peptide was the N-terminal peptide, which is not susceptible to Edman degradation and is thus probably a-N-acetylated as in most other organisms. The results confirm the conservation of H4 sequence and place Physarum H 4 in an intermediate position between lower eukaryote H4, such as yeast or Tetrahymena H4, and higher eukaryote H4, such as mammalian H 4 or pea H4.

Post-translational modifications of histones H3 and H4 associated with the histone methyltransferases Suv39h1 and G9a

“…This ten-fold decrease is mostly due to the poor signal, but this ion gives a robust and complete y-series and a poor b-series in the collision fragmentation result. This modification has never been described in mammals but was suggested in Physarum [ 28 ]. We also detected an acetylated form of this amino acid at a level of 6% in the background cell lines.…”

A Class II Histone Deacetylase Acts on Newly Synthesized Histones in Tetrahymena

“…First, two H3/H4 heterodimers are deposited onto the DNA, followed by two H2A/H2B dimers (14). Prior to their deposition, free histones H3 and H4 are acetylated on lysine (Lys) residues by cytoplasmic histone acetyltransferases (HATs) in patterns distinct from transcription acetylation patterns (2,21,42,53). The deposition acetylation promotes histone assembly into nucleosomes through interactions with histone chaperones, such as chromatin assembly factor 1 (CAF-1), Hif1, and Asf1, that incorporate new H3 and H4 specifically onto newly replicated DNA (1,14,45,52).…”

A Chinese hamster ovary cell histone deacetylase that is associated with a unique class of mononucleosomes

“…Peptide H4[ 1-23] contains only four lysines (residues 5,8, 12, and 16), all of which may be aoetylated and deacetylated in H4 in vivo (DeLange et al, 1969;Dixon et al, 1975), and we were able to specifically label them in vitro with acetic anhydride to a very high specific activity. Histones that are labeled with acetate in vivo give substrates with a relatively low specific activity, resulting in assays that are 100-1000 times less sensitive than those that utilize the chemically acetylated H4[ 1-23] as a substrate (Waterborg & Matthews, 1982). …”

“…The histone deacetylase assay is as described by Waterborg and Matthews (1982) with the following modifications. Twenty microliters of enzyme was added to 20 pL of 25 mM Tris-HC1 buffer, pH 8.0, containing 0.5 pL of tritiated, acetylated peptide H4[ 1-23].…”