Biophysical Society Thematic Meeting| Les Houches 2019

Multiscale Modeling of Chromatin: Bridging Experiment with Theory

Tuesday Speaker Abstracts

IMAGING LOCAL AND GLOBAL CHROMATIN STRUCTURE AS A 3D CONTINUUM WITHIN THE NUCLEUS: PROGRESS AND FUTURE STRATEGIES Mark Ellisman 1 ; Guillaume Castillon 1 ; Thomas Deerinck 1 ; Hiroyuki Hakozaki 1 ; Mason Mackey 1 ; Horng Ou 2 ; Steven Peltier 1 ; Sebastien Phan 1 ; Ranjan Ramachandra 1 ; Jan Soroczynski 2 ; Jingwen Yin 2 ; Clodagh O’Shea 2 ; 1 University of California San Diego, Department of Neurosciences, National Center for Microscopy and Imaging Research (NCMIR), La Jolla, California, United States 2 The Salk Institute , Biological Studies, La Jolla, California, United States Recent 3D EM analyses have advanced knowledge of how local nucleosome organization and global 3D organization of DNA in the nucleus relates to the functional activity of our genome in different cell states. To provide insight and understanding on this front, we are actively developing new probes and methods for correlated, multimodal microscopy to enable the visualization of chromatin in situ across scales - from the global organization of chromatin polymers within megabase 3D domains to the level of individual nucleosomes. We have generated and the first 3D EM datasets and multiscale reference maps of chromatin ultrastructure in whole cell nuclei and made these data openly available. This work was enabled by our development of ChromEMT 1 , which combines ChromEM with new advances in multi-tilt EM tomography (EMT) 2 . Progress on this ongoing project will be described with some emphasis given to future plans, focusing on new opportunities to further improve resolution, field-of-view, sample preservation and multimodal data integration. In particular, plans for the refinement and application of a new CryoChem technique 3 , which allows for genetically labeled and cryofixed samples to be characterized with 3D CLEM, exploiting cryofixation for high quality ultrastructural preservation while retaining the use of diaminobenzidine labeling using genetic EM tags – offering a roadmap to advance our EM (and cryoEM) imaging strategies into tissue. We also preview novel strategies for labeling specific genomic loci or intranuclear bodies that can be imaged with LM, mapped in 3D with x-ray microCT, and then localized and imaged at EM resolution with STEM tomography (and eventually cryo-STEM tomography).


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