Kinetics of accumulation and depletion of soluble newly synthesized histone in the reciprocal regulation of histone and DNA synthesis.

Procedures are presented which permit the identification and analysis of cellular histone that is not bound to chromatin. This histone, called soluble histone, could be distinguished from that bound to chromatin by the state of H4 modification and the lack of H2A ubiquitination. Changes in the levels of newly synthesized soluble histone were analyzed with respect to the balance between histone and DNA synthesis in hamster ovary cells. Pulse-chase protocols suggested that the chase of newly synthesized histone from the soluble fraction into chromatin may have two kinetic components with half-depletion times of about 1 and 40 min. When protein synthesis was inhibited, the pulse-chase kinetics of newly synthesized histone from the solubl fraction into chromatin were not significantly altered from those of the control. However, in contrast to the control, when protein synthesis was inhibited, DNA synthesis was also inhibited with kinetics similar to those of the chase of newly synthesized histone from the soluble fraction. There was a rapid decrease in the rate of DNA synthesis with a half-deceleration time of 1 min down to about 30% of the control rate, followed by a slower decrease with an approximate half-deceleration time of 40 min. When DNA synthesis was inhibited, newly synthesized histone accumulated in the soluble fraction, but H2A and H2B continued to complex with chromatin at a significant rate. Soluble histone in G1 cells showed the same differential partitioning of H4/H3 and H2A/H2B between the soluble and chromatin-bound fractions as was found in cycling cells with inhibited DNA synthesis. These results support a unified model of reciprocal regulatory mechanisms between histone and DNA synthesis in the assembly of chromatin.

Histones and their modifications.

Histones constitute the protein core around which DNA is coiled to form the basic structural unit of the chromosome known as the nucleosome. Because of the large amount of new histone needed during chromosome replication, the synthesis of histone and DNA is regulated in a complex manner. During RNA transcription and DNA replication, the basic nucleosomal structure as well as interactions between nucleosomes must be greatly altered to allow access to the appropriate enzymes and factors. The presence of extensive and varied post-translational modifications to the otherwise highly conserved histone primary sequences provides obvious opportunities for such structural alterations, but despite concentrated and sustained effort, causal connections between histone modifications and nucleosomal functions are not yet elucidated.

Occurrence of the low-mobility H1 histones subfraction in embryonic, differentiated, and neoplastic tissues of the Syrian hamster.

Electrophoretically slow H1 histone subfractions with mobilities identical to that of the subfraction found in the Kirkman-Robbins hamster hepatoma chromatin have been shown to be present in 12-day hamster embryos and in a sarcoma-type hamster tumor induced by SV40. No subfractions of such mobility were found in hamster liver, regenerating liver, thymus, spleen, and a fast-growing transplantable amelanotic hamster melanoma. A suggestion is made that some defective mechanisms of differentiation may affect the regulation of expression of the genes coding for the H1 histone subfractions. The same mechanisms may possibly but not necessarily be connected with the molecular events leading to neoplastic growth.