Dynamic Chromatin Conformation - a Direct Imaging Approach to Structure-Function Relationships

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MCB9728932 Abstract There is now overwhelming evidence that the folding and compaction of the linear array of nucleosomes that comprises the eukaryotic genome plays a crucial role in transcriptional regulation. Examples range from the precise positioning of nucleosomes on promoter and enhancer control sequences to the large-scale compaction and hence transcriptional shutdown of blocks of genes. In order to define the rules and mechanisms of chromatin folding, it is important to understand the dynamic three dimensional (3D) conformation arrays of nucleosomes. A technique based on cryo-electron microscopy has been developed that allows the 3D hydrated conformation of individual oligonucleosomes to be observed. Preliminary results have demonstrated that nucleosomes array in a 3D zigzag structure that can undergo further compaction depending on the length of linker DNA. Further studies will determine (1) whether the DNA linker sequence as well as its length effects the folding pattern, (2) how salt concentration influences folding, (3) what the effect of histone H1 is and how it is positioned relative to the nucleosomes, (4) how a novel chromatin binding protein termed MENT is involved in heterochromatin condensation, and (5) how transcribing RNA polymerase interacts with nucleosomes that contain DNA segments with transcriptional elements and arrest sequences. The studies are highly significant since very little is known about nucleosome packing and higher orders of chromatin structure even though these features are believed to be extremely important in regulation of chromatin function.