Nuclear scaffold

The detachment of chromatin from the nuclear scaffold, leading to chromatin conden.sation. 

It has been assumed that the nucleus contains an immobile structure , in which chromatin fibers are partially attached and fixed. These structures are called nuclear matrix or nuclear scaffold . When cells are successively treated with detergent, high-salt solution and DNase I, the nuclear scaffold can be observed as a fibrous network in the cell nucleus (Fey et al, 1986 Nickerson, 2001 Yoshimura et al, 2003) (Fig. 2b). The biochemical analyses of the nuclear scaffold have identified a 174kDa protein as a major component (Fisher et al, 1982). This protein is now known as topo II (Berrios et al, 1985). 

Amati B, Pick L, Laroche T, Gasser SM (1990) Nuclear scaffold attachment stimulates, but is not essential for ARS activity in Saccharomyces cerevisiae Analysis of the Drosophila ftz SAR. EMBO J 9(12) 4007-4016... 

Mirkovitch J, Mirault ME, Laemmli UK (1984) Organization of the higher-order chromatin loop Specific DNA attachment sites on nuclear scaffold. Cell 39(l) 223-232 Mitchell RS, Beitzel BF, Schroder AR, Shinn P, Chen H, Berry CC, Ecker JR, Bushman FD (2004) Retroviral DNA integration ASLV, HIV, and MLV show distinct target site preferences. PLoS Biol 2(8) E234... 

Two histone molecules each of types H2A (blue), H2B (green), H3 (yellow), and H4 (red) form an octameric complex, around which 146 bp of DNA are wound in 1.8 turns. These particles, with a diameter of 7 nm, are referred to as nucleosomes. Another histone (HI) binds to DNA segments that are not directly in contact with the histone octamers ( linker DNA). It covers about 20 bp and supports the formation of spirally wound superstructures with diameters of 30 nm, known as solenoids. When chromatin condenses into chromosomes, the solenoids form loops about 200 nm long, which already contain about 80 000 bp. The loops are bound to a protein framework (the nuclear scaffolding), which in turn organizes some 20 loops to form minibands. A large number of stacked minibands finally produces a chromosome. In the chromosome, the DNA is so densely packed that the smallest human chromosome already contains more than 50 million bp. 

Nucleosomes are organized into 30 nm fibers, and the fibers are extensively folded to provide the 10,000-fold compaction required to fit a typical eukaryotic chromosome into a cell nucleus. The higher-order folding involves attachment to a nuclear scaffold that contains histone HI, topoisomerase II, and SMC proteins. 

Nucleosomes can be packed more tightly to form a polynucleosome (also called a nucle-ofilament), which is organized into loops that are anchored by a nuclear scaffold containing several proteins. Additional levels of organization create a chromosome. 

Fig. 11.17 Topoisomerase 1 Allows Strain Relief Ahead of the Replication Fork. Prokaryotic chromosomes are Mobius strips. Topoisomerase 11 creates gates by nicking both strands of the dsDNA, allowing helices to cross one another (Fig 11.18). Tangles and linked loops can therefore be separated. It also forms part of the eukaryotic nuclear scaffold, for similar reasons.

The discussion of supercoiling also applies to the linear DNA molecules found in the nuclei of eukaryotic cells. Such molecules are constrained by their attachment to nuclear scaffolds, which are structural components of chromosomes. 

In one proposal for the structure of 200-nm filaments, the 30-nm fiber is looped and attached to a nuclear scaffold composed of protein. 

There is good evidence for independent chromosomal domains of about 50- to 100-thousand base pairs (50100 kb) of DNA. In such a domain, the ends of the domain are anchored so that the domain may be torsionally constrained, even though the chromosome is linear. One domain can contain any or all of the elements of chromatin structure already mentioned. The current hypothesis is that the ends of the domains are anchored to a large, stable protein structure, called the nuclear matrix (in interphase) or the nuclear scaffold (in metaphase). In some cases, one domain may represent one independently controlled transcriptional unit. The possibility of DNA becoming supercoiled within a torsionally constrained domain is discussed in Chapter 2. 

Proteins help to compact DNA this is important because the DNA in a chromosome could not fit inside its cell if it were not compacted. Histones are positively charged proteins that neutralize negative DNA strands when they wrap around and form complexes with the DNA. This wrapped structure, called beads on a string, represents the first level of compaction. The beads are condensed to form fibers, fibers fold into loops, loops combine with nuclear scaffold proteins to form rosettes, and rosettes condense to form coils. Finally, a chromatid with ten or more coils is formed. Nonhistone proteins within chromosomes are also important. These proteins have varied functions, including assisting in the unwinding of DNA and in the repairing of DNA. 

Mirkovitch, J., Mirault, M.-E., and Laemmli, U. K. (1984). Organization of the higher-order chromatin loop specific DNA attachment sites on the nuclear scaffold. Cell (Cambridge, Mass.) 39,223-232. 

Adolph. K. W. (1980). Organization of chromosomes in HeLa cells Isolation of histone-depleted nuclei and nuclear scaffolds. J. Cell Set. 42, 291-304. 

The bands observed at 69 and 64 kD may be due to ADP-ribosylation of lamin A and lamin B, major constituents of the nuclear matrix, as was previously suggested by Song and Adolph [26]. They reported that nuclear scaffolds isolated by mild micrococcal nuclease digestion from in vitro labeled HeLa cell nuclei contained labeled proteins at 116 kD and in the 65 to 70 kD range. 

---