Reversible in vitro packing of nucleosomal filaments into globular supranucleosomal units in chromatin of whole chick erythrocyte nuclei.
Structural transitions between extended and condensed forms of chromatin have been studied by electron microscopy of whole chicken erythrocyte nuclei incubated in low buffer concentrations (0.5 mM sodium borate; 1 to 5 mM Tris-HCl) with and without additions of monovalent or divalent cations. Both spread preparations and ultrathin sections have been examined. It is shown that the condensed chromatin present in chicken erythrocytes is organized, at the supranucleosomal level, in granular particles (mean diameter 29 +/- 3 nM0 tightly apposed against each other in beaded chains (higher order fibrils). These chains of higher order granules are gradually unravelled in low salt buffers lacking divalent cations into extended nucleosomal filaments. When nuclear chromatin swollen in low salt buffer and extended uniformly into nucleosomal filaments is exposed to buffers containing higher concentrations of monovalent ions (e.g., 100 mM NaCl) or low concentrations of divalent cations (e.g., 0.05-0.5 mM MgCl2) rapid and almost complete recondensation of the entire chromatin of a nucleus into one compact spheroidal aggregate of chains of higher order granules (diameters 25-30 nm) is observed. This transition is reversible and whole nuclear chromatin can be induced, in vitro, to undergo several cycles of such conformational changes. The results show that reconstitution of globular forms of higher order packing of nucleosomes is not restricted to oligonucleosomal units but also takes place, in a rather uniform and regular mode, in long nucleosomal filaments as they are present in whole nuclei. The data are discussed in relation to other studies of chromatin ultrastructure and are interpreted to indicate that the predominant, if not exclusive form of supranucleosomal packing existing in condensed inactive chromatin of living cells is a chromatin fibril made up of serially arranged, close-packed granular particles ("suprabeads" sensu in [64]). In addition, the results suggest that higher order packing of chromatin into such supranucleosomal granules is not simply a function of ionic strength but that the type of ions present strongly influences the state of chromatin packing.
D000818:Animals; D002122:Calcium Chloride / Q000494:pharmacology; D002467:Cell Nucleus / Q000648:ultrastructure; D002645:Chickens / Q000097:blood; D002843:Chromatin / Q000648:ultrastructure*; D004247:DNA; D004912:Erythrocytes / Q000648:ultrastructure*; D008274:Magnesium / Q000494:pharmacology; D008854:Microscopy, Electron; D009707:Nucleosomes / Q000648:ultrastructure*; D011487:Protein Conformation
D002843:Chromatin; D009707:Nucleosomes; D004247:DNA; D008274:Magnesium; D002122:Calcium Chloride