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3. How Genomes Function
In order for the cell to utilize the biological information contained within its genome, groups of genes,
each gene representing a single unit of information, have to be expressed in a coordinated manner. This
coordinated gene expression determines the make-up of the transcriptome, which in turn specifies the
nature of the proteome and defines the activities that the cell is able to carry out.
In Part 3 of Genomes we examine the events that result in the transfer of biological information from
genome to proteome. Our knowledge of these events was initially gained through studies of individual
genes, often as ‘naked' DNA in test-tube experiments. These experiments provided an interpretation of
gene expression that in recent years has been embellished by more sophisticated studies that have taken
greater account of the fact that, in reality, it is the genome that is expressed, not individual genes, and
that this expression occurs in living cells rather than in a test tube.
We begin our investigation of genome expression in Chapter 8, by examining the substantial and
important impact that the nuclear environment has on the utilization of the biological information
contained in the genomes of eukaryotes, the accessibility of that information being dependent on the
way in which the DNA is packaged into chromatin and being responsive to processes that can silence or
inactivate part or all of a chromosome.
Chapter 9 describes the events involved in initiation of transcription, and emphasizes the critical role the
DNA-binding proteins play during the early stages of genome expression. The synthesis of transcripts and
their subsequent processing into functional RNAs is dealt with in Chapter 10, and Chapter 11 covers the
equivalent events that lead to synthesis of the proteome.
As you read Chapters 8–11 you will discover that control over the composition of the transcriptome and
of the proteome can be exerted at various stages during the overall chain of events that make up
genome expression. These regulatory threads will be drawn together in Chapter 12, where we examine
how genome activity changes in response to extracellular signals during differentiation and development.
8. Accessing the Genome
9. Assembly of the Transcription Initiation Complex 1
0. Synthesis and Processing of RNA
11. Synthesis and Processing of the Proteome
12. Regulation of Genome Activity
8. Accessing the Genome
Learning outcomes
When you have read Chapter 8, you should be able to:
1. Explain how chromatin structure influences genome expression
2. Describe the internal architecture of the eukaryotic nucleus
3. Distinguish between the terms ‘constitutive heterochromatin', ‘facultative
heterochromatin' and ‘euchromatin'
4. Discuss the key features of functional domains, insulators, and locus control
regions, and describe the experimental evidence supporting our current
knowledge of these structures
5. Describe the various types of chemical modification that can be made to
histone proteins, and link this information to the concept of the ‘histone code'
6. State why nucleosome positioning is important in gene expression and give
details of a protein complex involved in nucleosome remodeling
7. Explain how DNA methylation is carried out and describe the importance of
methylation in silencing the genome
8. Give details of the involvement of DNA methylation in genomic imprinting and
X inactivation
8.1. Inside the Nucleus
8.2. Chromatin Modifications and Genome Expression
8.1. Inside the Nucleus
Figure 8.1. The internal architecture of the eukaryotic nucleus. (A) Transmission electron
micrograph showing the nuclear matrix of a cultured human HeLa cell. Cells were treated
with a non-ionic detergent to remove membranes, digested with a deoxyribonuclease to
degrade most of the DNA, and extracted with ammonium sulfate to remove histones and
other chromatin-associated proteins. From Molecular Cell Biology, by H Lodish, A Berk, SL
Zipursky, P Matsudaira, D Baltimore and J Darnell. ©1986, 1990, 1995, 2000 by WH
Freeman and Company. Used with permission. (B) and (C) Images of living nuclei
containing fluorescently labeled proteins (see Technical Note 8.1). In (B), the nucleolus is
shown in blue and Cajal bodies in yellow. The purple areas in (C) indicate the positions of
proteins involved in RNA splicing. B and C from Misteli, Science, 291, 843–847. Copyright
2000 American Association for the Advancement of Science.
Figure 8.2. A scheme for organization of DNA in the nucleus. The nuclear matrix is a fibrous
protein-based structure whose precise composition and arrangement in the nucleus has
not been described. Euchromatin, predominantly in the form of the 30 nm chromatin fiber
(see Figure 2.6 ) is thought to be attached to the matrix by AT-rich sequences called matrixassociated or scaffold attachment regions (MARs or SARs)
Figure 8.3. A functional domain in a DNase I sensitive region.
Figure 8.4. Insulator sequences in the fruit-fly genome. The diagram shows the
region of the Drosophila genome containing the two hsp70 genes. The insulator
sequences scs and scs′ are either side of the gene pair. The arrows below the two
genes indicate that they lie on different strands of the double helix and so are
transcribed in opposite directions.
Figure 8.5. The positional effect. (A) A cloned gene
that is inserted into a region of highly packaged
chromatin will be inactive, but one inserted into
open chromatin will be expressed. (B) The results of
cloning experiments without (red) and with (blue)
insulator sequences. When insulators are absent,
the expression level of the cloned gene is variable,
depending on whether it is inserted into packaged
or open chromatin. When flanked by insulators, the
expression level is consistently high because the
insulators establish a functional domain at the
insertion site.
Figure 8.6. Insulators maintain the independence of a functional domain. (A) When placed
between a gene and its upstream regulatory modules, an insulator sequence prevents the
regulatory signals from reaching the gene. (B) In their normal positions, insulators prevent
cross-talk between functional domains, so the regulatory modules of one gene do not
influence expression of a gene in a different domain. For more details about regulatory
modules, see Box 9.6
Figure 8.7. DNase I hypersensitive sites indicate the position of the locus control region
for the human β-globin gene cluster. A series of hypersensitive sites are located in the
20 kb of DNA upstream of the start of the β-globin gene cluster. These sites mark the
position of the locus control region. Additional hypersensitive sites are seen
immediately upstream of each gene, at the position where RNA polymerase attaches to
the DNA. These hypersensitive sites are specific to different developmental stages,
being seen only during the phase of development when the adjacent gene is active. The
60 kb region shown here represents the entire β-globin functional domain. See Figure
2.14 for more information on the developmental regulation of expression of the βglobin gene cluster.
Technical Note 8.1. Fluorescence recovery after photobleaching (FRAP)
8.2. Chromatin Modifications and Genome Expression
Figure 8.8. Two ways in which chromatin structure can influence gene expression. A
region of unpackaged chromatin in which the genes are accessible is flanked by two
more compact segments. Within the unpackaged region, the positioning of the
nucleosomes influences gene expression. On the left, the nucleosomes have regular
spacing, as displayed by the typical ‘beads-on-a-string' structure. On the right, the
nucleosome positioning has changed and a short stretch of DNA, approximately 300 bp,
is exposed. See Figures 2.5 and 2.6 for more details on nucleosomes.
Figure 8.9. Two views of the nucleosome core octamer. The view on the left is downwards
from the top of the barrel-shaped octamer; the view on the right is from the side. The two
strands of the DNA double helix wrapped around the octamer are shown in brown and
green. The octamer comprises a central tetramer of two histone H3 (blue) and two histone
H4 (bright green) subunits plus a pair of H2A (yellow)–H2B (red) dimers, one above and
one below the central tetramer. Note the N-terminal tails of the histone proteins
protruding from the core octamer. Reprinted with permission from Luger et al., Nature,
389, 251–260. Copyright 1997 Macmillan Magazines Limited
Figure 8.10. Nucleosome remodeling, sliding and transfer
Figure 8.11. Maintenance methylation and de novo methylation
Figure 8.12. A model for the link between DNA methylation and genome expression.
Methylation of the CpG island upstream of a gene provides recognition signals for
the methyl-CpG-binding protein (MeCP) components of a histone deacetylase
complex (HDAC). The HDAC modifies the chromatin in the region of the CpG island
and hence inactivates the gene. Note that the relative positions and sizes of the
CpG island and the gene are not drawn to scale.
Figure 8.13. A pair of imprinted genes on human chromosome 11. Igf2 is imprinted
on the chromosome inherited from the mother, and H19 is imprinted on the paternal
chromosome. The drawing is not to scale: the two genes are approximately 90 kb
apart.
Research Briefing 8.1. Discovery of the mammalian