The Core Histones from the Slime Mold Physarum polycephalum

Abstract

The histones of Physarum polycephalum have been isolated and fractionated using a combination of gel exclusion and ion-exchange chromatography or differential precipitation. The four core histones were unambiguously identified by gel electrophoresis, amino acid composition and N-terminal analysis.The molecular weight of Physarum histones H3, H2B and H4 are very close or identical to the corresponding histones from chicken erythrocytes. Physarum H2A is significantly larger than chicken erythrocyte H2A and its N-terminal residue is alanine. From the differences in amino acid compositions and in the peptide maps between Physarum and calf H4, one can expected some changes in the amino acid sequence of Physarum H4.The slime mold Physarum polycephalum is widely used as a model system for cell cycle studies [I]. In most instances, information gained in studying Physarum can be extrapolated to more complex eukaryotic organisms. However, to our knowledge, the four 'core histones' of Physarum have never been described unambiguously. As pointed out by Mohberg and Rusch [15], Physarum histones are difficult to isolate. The nuclei are extremely resistant to breakage and frequently contain an acidic polysaccharide which forms complexes with the histones.Physarum histones were named by Mohberg and Rusch [15] in reverse order of their electrophoretic mobilities in an acidlurea system: P 1, 2a, 2b, 3, 4, 5 and 6 [16-201. The molecular weights of these fractions were determined by electrophoresis on polyacrylamide gels containing sodium dodecyl sulfate [17,18]. Gel filtration chromatography was used with limited success and, on the basis of amino acid analysis and peptide mapping [18,19], the principal histone fractions P1, P4a, P4b, P5 and P6 were assumed to be homologous to the histones H1, H3, H2B, H2A and H4 of calf thymus, respectively. More recently it was suggested that histone P3 of Physarum and histone H2B from calf thymus have regions of homology [20].We describe here a method for purifying the four core histones of Physarum polycephalum. These proteins were characterized by polyacrylamide gel electrophoresis, amino acid analysis, end-group determination and peptide mapping. We compare the data to those obtained with other known histones. Finally we conclude that histone P3 from Physarum polycephalum is homologous to histone H2A. MATERIALS AND METHODSCulture of Physarum polycephalum Plasmodia as described by Daniel and Rusch [21].Macroplasmodia were grown in petri dishes for two days Nuclei were suspended in 1 M CaCI2 (60 ml for ten macroplasmodia) and 1 mM phenylmethylsulfonyl fluoride was added. Nuclei were lysed for 5 min at 80°C, cooled in ice and centrifuged at 4 "C for 30 min at 40000 x g. HCI was added to the supernatant to a final concentration of 0.25 M.The mixture was stirred at 4°C for 1 h, then centrifuged 30 min at 40000 x g.Histones were precipitated from the supernatant by 25 trichloroacetic acid at 4 "C overnight, centrifuged at 40000 x g, washed with acidified acetone, then with acetone, and finally dri...

High mobility group like chromosomal proteins from amebas of the acellular slime mold Physarum polycephalum

“…It is only recently that histones from the acellular slime mold Physarum polycephalum were isolated and characterized. Using either plasmodia (Chahal et al, 1980;Mende et al, 1983;Champagne et al, 1982) or amoebas (Cote et al, 1982),…”