Purification and characterization of the histones associated with the macronucleus of Tetrahymena

Abstract

Histone fractions were isolated from the macronucleus of T. pyriformis. Five classes of macronuclear histone were purified, using a combination of gel exclusion and ion-exchange chromatography and were examined with respect to their solubility, electrophoretic, chromatographic and chemical properties. Tetrahymena histone fraction, H4, is very similar to vertebrate H4, but exhibits a larger number of acetylated subfractions. The other Tetrahymena histones vary more extensively from their calf thymus counterparts. Tetrahymena H3 resembles calf thymus H3 in its solubility properties and is the only macronuclear histone containing cysteine. It differs from vertebrate H3 in composition and has a faster electrophoretic mobility on both urea-acrylamide and sodium dodecyl sulfate-acrylamide gel electrophoresis. Tetrahymena H3 also displays a level of acetylation higher than that reported for its vertebrate homologue. Approximately 45% of macronuclear H2B which resembles calf thymus H2B in composition and solubility is present in a (mono)acetylated form, not detected in vertebrate somatic H2B. H1, though similar to its calf thymus homologue in solubility, modification (by phosphorylation), and other properties, differs considerably in its content of basic, acidic, and hydrophobic amino acids. Tetrahymena does not contain a histone strictly homologous to H2A. Although macronuclear histone X resembles H2A in chromatographic and some solubility properties, it displays compositional and other solubility properties more like H2B than H2A. Fraction X is polymorphic in sodium dodecyl sulfate-acrylamide gels, migrating as 2 distinct molecular forms. While it is possible that 1 form is H2A-like and the other more H2B-like, the observation that both forms of X behave identically in solubility fractionation schemes makes this unlikely. Fraction X is both phosphorylated and acetylated which, in addition to 2 molecular forms, results in a characteristic heterogenous pattern on urea-acrylamide gels. Characterization of the histone complement of this lower eukaryote shoud contribute to the understanding of the evolution and biological role of these basic proteins. This description represents the most extensive analysis to date of the histones associated with an amitotic, genetically active nucleus. It serves as a reference to which the histones of the morphologically distinct, mitotically dividing, and genetically inactive micronucleus of this organism can be compared.