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Isolation, identification, and characterization of histones from plasmodia of the true slime mold Physarum
Abstract:
Histones from plasmodia of the true slime mold Physarum polycephalum have been prepared free of slime by an approach to histone isolation that uses extraction of nuclei with 40% guanidine hydrochloride and chromatography of the extract on Bio-Rex 70. This procedure followed by chromatography or electrophoresis has been used to obtain pure fractions of histones from Physarum microplasmodia. Physarum microplasmodia have five major histone fractions, and we show by amino acid analysis, apparent molecular weight on three gel systems containing sodium dodecyl sulfate, mobility on gels containing Triton X-100, and other characterizations that these fractions are analogous to mammalian histones H1, H2A, H2B, H3, and H4. Significant differences between Physarum and mammalian histones are noted, with histone H1 showing by far the greatest variation. Histones H1 and H4 from Physarum microplasmodia have similar, but not identical, products of partial chymotryptic digestion compared with those of calf thymus histones H1 and H4. Labeling experiments, in vivo, showed that histone H1 is the major phosphorylated histone and approximately 15 separate phosphopeptides are present in a tryptic digest of Physarum histone H1. The core histones from Physarum, histones H2A, H2B, H3, and H4, are rapidly acetylated; histone H4 shows five subfractions, analogous to the five subfractions of mammalian histone H4 (containing zero to four acetyllysine residues per molecule); histone H3 has a more complex pattern that we interpret as zero to four acetyllysine residues on each of two sequence variants of histone H3; histones H2A and H2B show less heterogeneity. Overall, the data show that Physarum microplasmodia have a set of histones that is closely analogous to mammalian histones.
Insights
This study isolated and characterized histones from the slime mold Physarum polycephalum, finding they are analogous to mammalian histones. Differences were noted, particularly in histone H1, but core histones showed similar acetylation patterns.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Histones are crucial proteins involved in DNA packaging and regulation.
- Understanding histone diversity across species provides insights into evolutionary conservation and functional divergence.
Purpose of the Study:
- To isolate and characterize histone proteins from the true slime mold Physarum polycephalum.
- To compare Physarum histones with their mammalian counterparts to identify similarities and differences.
Main Methods:
- Histone isolation using guanidine hydrochloride extraction and Bio-Rex 70 chromatography.
- Characterization via amino acid analysis, SDS-PAGE, Triton X-100 gel electrophoresis, and partial chymotryptic digestion.
- In vivo labeling experiments to study histone phosphorylation and acetylation.
Main Results:
- Five major histone fractions analogous to mammalian H1, H2A, H2B, H3, and H4 were identified in Physarum.
- Histone H1 exhibited significant variation compared to mammalian H1.
- Core histones (H2A, H2B, H3, H4) showed rapid acetylation, with H4 displaying subfractions similar to mammals.
- Histone H1 was identified as the major phosphorylated histone.
Conclusions:
- Physarum polycephalum possesses a set of histones that are highly analogous to mammalian histones.
- Despite overall similarity, notable differences exist, especially in histone H1, suggesting evolutionary divergence.
- The conserved acetylation patterns of core histones highlight their fundamental importance in eukaryotic organisms.
