Alan P. Wolffe

A positive role for nucleosome mobility in the transcriptional activity of chromatin templates: restriction by linker histones.

Nucleosome mobility facilitates the transcription of chromatin templates containing only histone octamers. Inclusion of linker histones in chromatin inhibits nucleosome mobility, directs nucleosome positioning and represses transcription. Transcriptional repression by linker histone occurs preferentially on templates associated with histone octamers relative to naked DNA. Mobile nucleosomes and the restriction of mobility by linker histones might be expected to exert a major influence on the accessibility of chromatin to regulatory molecules.

Competition between transcription complex assembly and chromatin assembly on replicating DNA.

We have used a Xenopus egg extract to show that a competition exists between the assembly of transcription complexes and nucleosomes on replicating 5S DNA. This competition results in the establishment of a transcriptionally repressed state for 5S DNA that is dependent on core histones but not on the precise positioning of the cores. The repression is selective, since satellite I DNA is not significantly repressed under these conditions. We demonstrate that the efficiency of chromatin assembly compared with transcription complex assembly is an important variable in determining gene activity.

Chapter 28 Chromatin Assembly

Publisher Summary This chapter provides an overview of chromation assembly. Chromatin represents an enigma for many molecular and developmental biologists. A major current challenge for molecular and developmental biologists is to understand chromatin. Chromatin and nuclear structure change during Xenopus embryogenesis. These changes relate to alterations in the length of the cell cycle, the activation of transcription, and the repression of certain genes. Cell-free systems exist that are derived from Xenopus eggs and oocytes that allow the reconstruction of chromatin and eventually nuclei in vitro . The reconstruction of chromatin in these extracts is highlighted in the chapter. This in vitro reconstruction of chromatin templates is the first step to the analysis of how transcription and replication occur within a chromosome. The solution of this problem leads to important insights into how these processes actually occur within the nucleus in vivo . The first experiments that clearly demonstrated the utility of cell-free preparations derived from Xenopus eggs and oocytes for assembling chromatin came from the discovery of Laskey and others that cloned circular simian virus 40 (SV40) DNA would be assembled into nucleosomes by a high speed supernatant of homogenized Xenopus eggs. The chapter also outlines the preparation of extracts.

Histone and DNA Contributions to Nucleosome Structure

Publisher Summary The nucleosome has an unexpectedly complex anatomy. The histone proteins undergo highly selective interactions between themselves and with DNA. Although the basis for this selectivity has not been resolved, several new features have been recently recognized. Most notable is the extended helical structure of the C-terminal domains of the histones. The C-terminal domains of the core histones do not form monomeric globules, but have extensive protein–protein and protein–DNA contacts. The interfaces between histone heterodimers are not extensive, and although specific, offer the possibility of conformational flexibility. Moreover, the stability of the interaction of the histones with DNA depends on the presence of continuous contact with DNA. This may be disrupted by trans-acting factors or RNA polymerase. Conformational changes in the histone octamer may follow from sequestration of linker histones into the nucleosome or from modification or mutation of the tail domains of the core histones. Since these changes are implicated in the regulation of trans-acting factor access to DNA in chromatin, they are likely to be an important area for future investigation.