THE EFFECT OF NUCLEOSOME PHASING SEQUENCES AND DNA TOPOLOGY ON NUCLEOSOME SPACING

The distances between the nucleosomes in eukaryotic chromatin that def ine the nucleosome repeat length are not universally constant, but var y between different cell types and activity states. We have previously established in a cell-free system that nucleosome spacing is essentia lly governed by electrostatic principles, most likely through charge n eutralisation of linker DNA by cations either free in solution or on f lexible histone domains. On the basis of the tight correlation between the parameters that affect nucleosome spacing and those that influenc e the folding of the nucleosomal fiber into higher order structures, w e suggested that there is an intimate relationship between nucleosome spacing and chromatin folding. Here we describe DNA topology as a new parameter that influences nucleosome spacing in a predictable way. The effects of topology and cation concentrations integrate to define the final repeat length. The phenomenon of ''nucleosome phasing'' describ es nucleosomal arrays that are generated through positioning of nucleo somes by the underlying DNA sequence. To determine the relative contri bution of DNA sequence and the parameters intrinsic to physiological c hromatin for nucleosomal positions, we created situations where these two principles were in conflict. We found that nucleosome repeats dire cted by a strong positioning sequence are dominated by the cation-indu ced spacing as well as by the effects of topology. We conclude that th e DNA sequence effects nucleosome spacing only by ''fine tuning'' of n ucleosome positions within the framework of a repeat pattern that is e stablished by other principles. (C) 1996 Academic Press Limited