The nuclear matrix revealed by eluting chromatin from a cross-linked nucleus.
The nucleus is an intricately structured integration of many functional domains whose complex spatial organization is maintained by a nonchromatin scaffolding, the nuclear matrix. We report here a method for preparing the nuclear matrix with improved preservation of ultrastructure. After the removal of soluble proteins, the structures of the nucleus were extensively cross-linked with formaldehyde. Surprisingly, the chromatin could be efficiently removed by DNase I digestion leaving a well preserved nuclear matrix. The nuclear matrix uncovered by this procedure consisted of highly structured fibers, connected to the nuclear lamina and built on an underlying network of branched 10-nm core filaments. The relative ease with which chromatin and the nuclear matrix could be separated despite extensive prior cross-linking suggests that there are few attachment points between the two structures other than the connections at the bases of chromatin loops. This is an important clue for understanding chromatin organization in the nucleus.
Nickerson
J A
JA
Department of Cell Biology, University of Massachusetts Medical School, Worcester, MA 01655, USA. Nickerson@wccf.mit.edu
Krockmalnic
G
G
Wan
K M
KM
Penman
S
S
D002478:Cells, Cultured; D002843:Chromatin / Q000648:ultrastructure*; D003432:Cross-Linking Reagents; D004247:DNA / Q000187:drug effects; D003850:Deoxyribonuclease I / Q000494:pharmacology; D005557:Formaldehyde; D016591:Histocytological Preparation Techniques; D006657:Histones / Q000302:isolation & purification; D015530:Nuclear Matrix / Q000648:ultrastructure*; D012313:RNA / Q000648:ultrastructure; D012261:Ribonucleoproteins / Q000648:ultrastructure; D013194:Staining and Labeling
D002843:Chromatin; D003432:Cross-Linking Reagents; D006657:Histones; D012261:Ribonucleoproteins; D005557:Formaldehyde; D012313:RNA; D004247:DNA; D003850:Deoxyribonuclease I