B. Alberts, A. Johnson, J. Lewis и др. - Molecular Biology of The Cell (6th edition) (1120996), страница 80
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The results suggest that three majortypes of repressive chromatin predominate in this organism, along with two majortypes of chromatin on actively transcribed genes, and that each type is associatedwith a different complex of non-histone proteins. Thus, classical heterochromatincontains more than six such proteins, including heterochromatin protein 1 (HP1),interbandsbands(A)2 µm(B)1 µmFigure 4–51 Micrographs of polytenechromosomes from Drosophila salivaryglands. (A) Light micrograph of a portion ofa chromosome.
The DNA has been stainedwith a fluorescent dye, but a reverse imageis presented here that renders the DNAblack rather than white; the bands areclearly seen to be regions of increasedDNA concentration. This chromosomehas been processed by a high-pressuretreatment so as to show its distinct patternof bands and interbands more clearly.(B) An electron micrograph of a smallsection of a Drosophila polytenechromosome seen in thin section. Bandsof very different thickness can be readilydistinguished, separated by interbands,which contain less condensed chromatin.(A, adapted from D.V. Novikov, I.
Kireevand A.S. Belmont, Nat. Methods 4:483–485, 2007. With permission fromMacmillan Publishers Ltd; B, courtesyof Veikko Sorsa.)THE GLOBAL STRUCTURE OF CHROMOSOMES211Figure 4–52 RNA synthesis in polytene chromosome puffs.An autoradiograph of a single puff in a polytene chromosome from thesalivary glands of the freshwater midge Chironomus tentans. As outlinedin Chapter 1 and described in detail in Chapter 6, the first step in geneexpression is the synthesis of an RNA molecule using the DNA as a template.The decondensed portion of the chromosome is undergoing RNA synthesisand has become labeled with 3H-uridine, an RNA precursor molecule that isincorporated into growing RNA chains. (Courtesy of José Bonner.)RNA synthesiswhereas the so-called Polycomb form of heterochromatin contains a similarnumber of proteins of a different set (PcG proteins). In addition to the five majorchromatin types, other more minor forms of chromatin appear to be present, eachof which may be differently regulated and have distinct roles in the cell.The set of proteins bound as part of the chromatin at a given locus variesdepending on the cell type and its stage of development.
These variations makethe accessibility of specific genes different in different tissues, helping to generatethe cell diversification that accompanies embryonic development (described inChapter 21).10 µmChromatin Loops Decondense When the Genes Within Them AreExpressedMBoC6 m4.62/4.51When an insect progresses from one developmental stage to another, distinctive chromosome puffs arise and old puffs recede in its polytene chromosomesas new genes become expressed and old ones are turned off (Figure 4–52).
Frominspection of each puff when it is relatively small and the banding pattern is stilldiscernible, it seems that most puffs arise from the decondensation of a singlechromosome band.The individual chromatin fibers that make up a puff can be visualized withan electron microscope. In favorable cases, loops are seen, much like thoseobserved in amphibian lampbrush chromosomes. When genes in the loop arenot expressed, the loop assumes a thickened structure, possibly that of a folded30-nm fiber, but when gene expression is occurring, the loop becomes moreextended.
In electron micrographs, the chromatin located on either side of thedecondensed loop appears considerably more compact, suggesting that a loopconstitutes a distinct functional domain of chromatin structure.Observations in human cells also suggest that highly folded loops of chromatinexpand to occupy an increased volume when a gene within them is expressed. Forexample, quiescent chromosome regions from 0.4 to 2 million nucleotide pairs inlength appear as compact dots in an interphase nucleus when visualized by fluorescence microscopy.
However, the same DNA is seen to occupy a larger territorywhen its genes are expressed, with elongated, punctate structures replacing theoriginal dot.New ways of visualizing individual chromosomes have shown that each of the46 interphase chromosomes in a human cell tends to occupy its own discrete territory within the nucleus: that is, the chromosomes are not extensively entangledwith one another (Figure 4–53). However, pictures such as these present onlyan average view of the DNA in each chromosome.
Experiments that specificallylocalize the heterochromatic regions of a chromosome reveal that they are often10 µm4910143Figure 4–53 Simultaneous visualization of the chromosome territoriesfor all of the human chromosomes in a single interphase nucleus. Here,a mixture of DNA probes for each chromosome has been labeled so as tofluoresce with a different spectra; this allows DNA–DNA hybridization to beused to detect each chromosome, as in Figure 4–10. Three-dimensionalreconstructions were then produced. Below the micrograph, eachchromosome is identified in a schematic of the actual image.
Note thathomologous chromosomes (e.g., the two copies of chromosome 9) are notin general co-located. (From M.R. Speicher and N.P. Carter, Nat. Rev. Genet.6:782–792, 2005. With permission from Macmillan Publishers Ltd.)1119 913 21152218127386X13141226417715 1718215 20212Chapter 4: DNA, Chromosomes, and GenomesFigure 4–54 The distribution of gene-rich regions of the human genomein an interphase nucleus.
Gene-rich regions have been visualized with afluorescent probe that hybridizes to the Alu interspersed repeat, which ispresent in more than a million copies in the human genome (see page 292).For unknown reasons, these sequences cluster in chromosomal regionsrich in genes. In this representation, regions enriched for the Alu sequenceare green, regions depleted for these sequences are red, while the averageregions are yellow. The gene-rich regions are seen to be largely absent inthe DNA near the nuclear envelope.
(From A. Bolzer et al., PLoS Biol.3:826–842, 2005.)5 µmclosely associated with the nuclear lamina, regardless of the chromosome examined. And DNA probes that preferentially stain gene-rich regions of human chromosomes produce a striking picture of the interphase nucleus that presumablyreflects different average positions for active and inactive genes (Figure 4–54).How is most of the chromatin in each interphase chromosome condensedwhen its genes are not being expressed? A powerful extension of the chromosomeconformation capture method described previously (see Figure 4–48), whichexploits a high-throughput DNA sequencing technology called massive parallelsequencing (see Panel 8–1, pp.
478–481), allows the connections between all ofthe different one-megabase (1 Mb) segments of the human genome to be mappedin human interphase chromosomes. The results reveal that most regions of ourchromosomes are folded into a conformation referred to as a fractal globule: aknot-free arrangement that facilitates maximally dense packing while, at the sametime, preserving the ability of the chromatin fiber to unfold and fold (Figure 4–55).Chromatin Can Move to Specific Sites Within the Nucleus to AlterGene ExpressionA variety of different types of experiments has led to the conclusion that theposition of a gene in the interior of the nucleus changes when it becomes highlyexpressed. Thus, a region that becomes very actively transcribed is sometimesfound to extend out of its chromosome territory, as if in an extended loop (Figure4–56).
We will see in Chapter 6 that the initiation of transcription—the first step ingene expression—requires the assembly of over 100 proteins, and it makes sensethat this would be facilitated in regions of the nucleus enriched in these proteins.More generally, it is clear that the nucleus is very heterogeneous, with functionally different regions to which portions of chromosomes can move as they aresubjected to different biochemical processes—such as when their gene expression changes. It is this issue that we discuss next.chromosome~5 megabasesCHROMOSOME FOLDINGIN NUCLEUSnucleuschromosome territorycross sectionMBoC6 m4.64/4.53Figure 4–55 A fractal globule model forinterphase chromatin.
An extension ofthe 3C method in Figure 4–48, called Hi-C,was used to measure the extent to whicheach of the three thousand 1 Mb segmentsin the human genome was located adjacentto any other of these segments. Theresults support the type of model shown.In the enlarged fractal globule illustrated,a region of 5 million base pairs is seen tofold in a way that keeps regions that areneighbors along the one-dimensional DNAhelix as neighbors in three dimensions;this gives rise to monochromatic blocks inthis representation that are obvious bothon the surface and in cross section. Thefractal globule is a knot-free conformationof the DNA that permits dense packing, yetretains an ability to easily fold and unfoldany genomic locus.
(Adapted fromE. Lieberman-Aiden et al., Science326:289–293, 2009. With permissionfrom AAAS.)THE GLOBAL STRUCTURE OF CHROMOSOMES213(A)5 µmnuclear envelopehomologous chromosomesdetected by hybridizationtechniquesFigure 4–56 An effect of high levels ofgene expression on the intranuclearlocation of chromatin. (A) Fluorescencemicrographs of human nuclei showinghow the position of a gene changes whenit becomes highly transcribed. The regionof the chromosome adjacent to the gene(red) is seen to leave its chromosomalterritory (green) only when it is highlyactive.
(B) Schematic representation ofa large loop of chromatin that expandswhen the gene is on, and contracts whenthe gene is off. Other genes that are lessactively expressed can be shown by thesame methods to remain inside theirchromosomal territory when transcribed.(From J.R. Chubb and W.A. Bickmore, Cell112:403–406, 2003. With permission fromElsevier.)specially marked gene(B)GENE OFFGENE ONNetworks of Macromolecules Form a Set of Distinct BiochemicalEnvironments inside the NucleusIn Chapter 6, we shall describe the function of a variety of subcompartments thatare present within the nucleus. The largest and most obvious of these is the nucleolus, a structure well known to microscopists even in the nineteenth century (seeFigure 4–9). The nucleolus is the cell’s site of ribosome subunit formation, as wellas the place where many other specializedreactions occur (see Figure 6–42): itMBoC6 m4.65/4.54consists of a network of RNAs and proteins concentrated around ribosomal RNAgenes that are being actively transcribed.
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