The internal structure and packing of eukaryotic chromosome fibers under physiological conditions is to be investigated by a combination of physical, chemical, and biological methods. The primary objective is to use electron microscopy to determine the internal arrangement of nucleosomes in transcriptionally inactive chromosome fibers. We propose to use x-ray scattering to find conditions under which fibers can be successfully preserved in the dehydrated state for microscopy. In separate experiments we will circumvent the dehydration problem entirely by imaging the fibers in the frozen hydrated state. X-ray experiments will be performed, examining fibers in solution and in nuclei to fill in gaps in our microscopic data and to compare structures in different types of cells. In order to characterize and model the well known salt dependent condensation phenomena of chromatin, a polyelectrolyte model will be tested by structural and thermodynamic studies. When the structure and structural transitions of inactive chromatin fibers are better understood, many experiments to understand the relationships between structures and function can be proposed. Differences between the structure of fibers taken from different cells and from different points in the cell cycle can be studied. In vitro studies of the salt dependent phase transitions of chromatin with modified histone or non-histone proteins might detect alterations in the relative stabilities of the """"""""heterochromatic"""""""" state that would be relevant to understanding the structural basis for constitutive and facultative repression of transcriptional activity.
Showing the most recent 10 out of 14 publications
