Structure of chromatin and the linking number of DNA.

Abstract

Recent observations suggest that the basic supranucleosomal structure of chromatin is a zigzag helical ribbon with a repeat unit made of two nucleosomes connected by a relaxed spacer DNA. A remarkable feature of one particular ribbon is that it solves the apparent paradox between the number of DNA turns per nucleosome and the total linking number of a nucleosome-containing closed circular DNA molecule. We show here that the repeat unit of the proposed structure, which contains two nucleosomes with -13/4 DNA turns per nucleosome and one spacer crossover per repeat, contributes -2 to the linking number of closed circular DNA. Space-filling models show that the cylindrical 250-A chromatin fiber can be generated by twisting the ribbon.It is now well-established that the unit structure of chromatin, the nucleosome (1-3), is a flat cylindrical particle (110 X 110 x 57 A) with DNA wrapped around a histone octamer in a lefthanded toroidal supercoil of approximately 80 base pairs (bp) per turn (4). There are 146 bp of DNA in the nucleosome core describing -13/4 superhelical turns (5) and 20-95 bp of spacer DNA between neighboring nucleosome cores (6).The nature of the supranucleosomal structure of chromatin is less clear. Electron microscopic studies of eukaryotic nuclei have revealed a "thin" 100-A chromatin fiber and a "thick" 250-A fiber (7-9). Many past models have assumed that neighboring nucleosomes are stacked side by side to generate a 100-A nucleofilament, which is further coiled into a 250-A "solenoid" (10, 11). The unstacked 100-A fiber ("beads on a string") can be readily observed in nuclei and chromatin preparations spread under isotonic conditions (1, 2, 12). A stacked 100-A nucleofilament also has been detected, but only at high salt concentrations (11). Although the 100-A fiber is present in histone Hi-depleted chromatin, the 250-A fiber is never observed under these conditions (11, 12). Thus, histone HI must play a role in the further compaction of the linear chain of beads (12, 13). Reconstitution studies with the four intranucleosomal histones and small circular DNAs in the presence of nicking-closing enzyme have revealed that the DNA coiling around a nucleosome changes the DNA linking number by -1 (14). The histone Hl-induced compaction of the "beads on a string" into a 250-A-diameter supranucleosomal structure (15) does not cause further changes in the linking number (16). Because the nicking-closing enzyme relaxes the spacer DNA (16), the observed change in the linking number must be due to the particular DNA structure in the nucleosome.The linking number L of a closed circular DNA (ccDNA) molecule is the number of topological revolutions made by one strand about the other [counted after the molecule is constrained to lie in a plane (17)]. L is a topological invariant (i.e., an integer which is unchanged by all deformations that leave the strands intact). Moreover, it is related to the writhing number W and the twist number T by the equation (18-21)W is a real number that measures the shape...

The regulatory role of DNA supercoiling in nucleoprotein complex assembly and genetic activity

“…Whether the helical form of the 30-nm fiber is a 1-start helix or solenoid (Finch and Klug 1976;Thoma et al 1979;Felsenfeld and McGhee 1986) with a single stack of nucleosomes, a 2-start helix with two continuous stacks (Williams et al 1986;Bordas et al 1986a, b;Woodcock et al 1984;Worcel et al 1981) analogous to DNA, or a multi-start helix (Daban and Bermúdez 1998), or whether fibers with different NRLs differ in topology (Wong et al 2007) has been extensively debated. There is now compelling experimental evidence that the '30-nm' fiber can adopt a 2-start structure.…”

The Nuclear Actin-related Protein ofSaccharomyces cerevisiae, Act3p/Arp4, Interacts with Core Histones

“…The effect of an act3 mutation on plasmid DNA linking number was checked, because previous studies have correlated alterations in chromatin structure with changes in linking number of DNA (Worcel et al, 1981). In these experiments, the linking number of circular DNA molecules is measured by gel electrophoresis in the presence of the intercalating dye chloroquine.…”

The higher-order structure of chromatin: evidence for a helical ribbon arrangement.

“…The results suggest that the initial folding step in nucleosome packing involves the formation of a zig-zag ribbon as has been proposed by others (Thoma F., T. Koller, and A. Klug, 1979, 1. Cell Biol., 83 :403-427 ;Worcel A., S. Strogartz, and D. Riley, 1981, Proc . Nad.…”

“…Studies of the arrangements of nucleosomes in these fibers are facilitated by the reversible salt-induced transition between the relaxed beaded chain ofnucleosomes and the 30-nm fiber (9,35,36). Electron micrographs of chromatin undergoing this transition often show a fairly regular zig-zag pattern of chromatosome-linker DNA-chromatosome (36,46), but as compaction proceeds, the individual nucleosomes are no longer resolved and their arrangement in the 30-nm fiber becomes less accessible to direct observation . However, a substantial body of data concerning the biophysical and biochemical properties of both compact and relaxed chromatin has been accumulated, which places many constraints on their possible architecture (reviewed in references 12 and 17).…”

“…A second model, based principally on morphological evidence, suggests that clusters or "superbeads" of nucleosomes in some undefined arrangement are strung together to make up the 30-nm fiber (23,27,32,33,47,48). A third proposal suggests that the underlying architecture is a flat zig-zag ribbon of nucleosomes which is twisted to generate the 30-nm fiber (34,46).…”