NEW DNA-SEQUENCE RULES FOR HIGH-AFFINITY BINDING TO HISTONE OCTAMER AND SEQUENCE-DIRECTED NUCLEOSOME POSITIONING

DNA sequences that position nucleosomes are of increasing interest bec ause of their relationship to gene regulation in vivo and because of t heir utility in studies of nucleosome structure and function in vitro. However, at present our understanding of the rules for DNA sequence-d irected nucleosome positioning is fragmentary, and existing positionin g sequences have many Limitations. We tarried out a SELEX experiment s tarting with a large pool of chemically synthetic random DNA molecules to identify those individuals having the highest affinity for histone octamer. A set of highest-affinity molecules were selected, cloned, a nd sequenced, their affinities (free energies) for histone octamer in nucleosome reconstitution measured, and their ability to position nucl eosomes in vitro assessed by native gel electrophoresis. The selected sequences have higher affinity than previously known natural or non-na tural sequences, and have a correspondingly strong nucleosome position ing ability. A variety of analyses including Fourier transform, real-s pace correlation, and direct counting computations were carried out to assess non-random features in the selected sequences. The results rev eal sequence rules that were already identified in earlier studies of natural nucleosomal DNA, together with a large set of new rules having even stronger statistical significance. Possible physical origins of the selected molecules' high affinities are discussed. The sequences i solated in this study should prove valuable for studies of chromatin s tructure and function in vitro and, potentially, for studies in vivo. (C) 1998 Academic Press Limited.