Journal of Cell Science, Vol 99, Issue 1 107-114, Copyright © 1991 by Company of Biologists

JOURNAL ARTICLES

Ultrastructure of chromatin. II. Three-dimensional reconstruction of isolated fibers

CL Woodcock, BF McEwen and J Frank
Department of Zoology, University of Massachusetts, Amherst 01003.

Electron-microscope tomography has been used to reconstruct isolated, negatively stained chromatin fibers from Necturus maculosus erythrocytes. Tilt series micrographs from +70 degrees to -70 degrees at 5 degrees intervals were obtained, allowing a reconstruction resolution of 3.3 nm for fibers lying parallel to the tilt axis. The fibers were found to be flattened in the plane of the carbon support, and also stained differentially according to the distance from the carbon. A number of methods of presenting the three-dimensional information were explored. Especially useful was an automatic peak search method for locating putative nucleosome positions coupled with the production of a computer-generated model. Other valuable techniques included the generation of projection stereograms and construction of solid models. A peripheral location of nucleosomes in the chromatin fiber was indicated, and helical arrangements of nucleosomes were observed over short regions. However, no long-range ordering of nucleosomes was apparent. The extent to which this lack of order may be the result of events occurring during the preparation of chromatin for electron microscopy is discussed.

This article has been cited by other articles:

				J. R. McIntosh Electron Microscopy of Cells: A New Beginning for a New Century J. Cell Biol., June 11, 2001; 153(6): F25 - F32. [Abstract] [Full Text] [PDF]

				K. v. Holde and J. Zlatanova Chromatin Higher Order Structure: Chasing a Mirage? J. Biol. Chem., April 14, 1995; 270(15): 8373 - 8376. [Full Text] [PDF]

				P. Giannasca, R. Horowitz, and C. Woodcock Transitions between in situ and isolated chromatin J. Cell Sci., January 6, 1993; 105(2): 551 - 561. [Abstract] [PDF]