1. Eukaryotic Genomes: Organization, Regulation, and Evolution
Chapter 19
Eukaryotic Genomes: Organization, Regulation, and Evolution

2. Overview: How Eukaryotic Genomes Work and Evolve
Two features of eukaryotic genomes are a major information-processing challenge:
First, the typical eukaryotic genome is much larger than that of a prokaryotic cell
Estimated 25,000 genes in the genome
Second, cell specialization limits the expression of many genes to specific cells

3. The DNA-protein complex, called chromatin, is ordered into higher structural levels than the DNA-protein complex in prokaryotes
Part of the chromatin is packed into the main axis, while those parts that are being actively transcribed are spread out in loops
Both prokaryotes and eukaryotes must alter their patterns of gene expression in response to changes in environmental conditions

5. Concept 19.1: Chromatin structure is based on successive levels of DNA packing
Eukaryotic DNA is precisely combined with a large amount of protein, and the resulting chromatin undergoes changes in the course of the cell cycle
Chromatin—mass within the nucleus
Before mitosis, chromatin coils and folds up to form chromosomes
Eukaryotic chromosomes contain an enormous amount of DNA relative to their condensed length
Average double helix is 1.5 x 108 nucleotide pairs

6. Nucleosomes, or “Beads on a String”
Proteins called histones are responsible for the first level of DNA packing in chromatin
Mass is equal to the mass of DNA
High proportion of positively charged amino acids
Binds tightly to negatively charged DNA (gets negativity from phosphorus groups)
The association of DNA and histones seems to remain intact throughout the cell cycle
Animation: DNA Packing

7. In electron micrographs, unfolded chromatin has the appearance of beads on a string
Each “bead” is a nucleosome, the basic unit of DNA packing
Consists of DNA wound around a protein core composed of 2 molecules each of four types of histone
“String” is called linker DNA

8. How can DNA be transcribed when it is wrapped around histones in a nucleosome?
Researchers have learned that changes in the shapes and positions of nucleosomes can allow RNA-synthesizing polymerases to move along the DNA

9. LE 19-2a 2 nm DNA double helix His- Histone tones tails 10 nm
Histone H1
10 nm
Linker DNA
(“string”)
Nucleosome
(“bead”)
Nucleosomes (10-nm fiber)

10. Higher Levels of DNA Packing
The next level of packing forms the 30-nm chromatin fiber
Due to interactions between the histone tails of one nucleosome and the linker DNA and nucleosomes to either side

11. LE 19-2b
30 nm
Nucleosome
30-nm fiber

12. In turn, the 30-nm fiber forms looped domains, making up a 300-nm fiber

13. LE 19-2c Protein scaffold Loops 300 nm Scaffold
Looped domains (300-nm fiber)

14. In a mitotic chromosome, the looped domains coil and fold, forming the metaphase chromosome

15. LE 19-2d
700 nm
1,400 nm
Metaphase chromosome

16. Interphase chromatin is usually much less condensed than that of mitotic chromosomes
Much of the interphase chromatin is present as a 10-nm fiber, and some is 30-nm fiber, which in some regions is folded into looped domains
Although an interphase chromosome lacks a scaffold, its looped domains seem to be attached to the nuclear lamina, on the inside of the nuclear envelope
May help organize regions of active transport

17. Interphase chromosomes have highly condensed areas, called heterochromatin, and less compacted areas, called euchromatin
Because of its compaction, heterochromatin DNA is largely inaccessible to transcription enzymes and is not transcribed
Looser packing of euchromatin makes its DNA accessible to enzymes and so is not usually transcribed

18. Concept 19.2: Gene expression can be regulated at any stage, but the key step is transcription
All organisms must regulate which genes are expressed at any given time
A multicellular organism’s cells undergo cell differentiation, specialization in form and function
Human body composed of 200 different cell types

19. Differential Gene Expression
Typical human cell expresses about 20% of all its genes at any given time
Differences between cell types result from differential gene expression, the expression of different genes by cells within the same genome
In each type of differentiated cell, a unique subset of genes is expressed
Many key stages of gene expression can be regulated in eukaryotic cells
Most commonly regulated at transcription

20. Chemical modification Degradation of protein
LE 19-3
Signal
NUCLEUS
Chromatin
DNA
Gene available
for transcription
Gene
Transcription
RNA
Exon
Primary transcript
Intro
RNA processing
Tail
Cap
mRNA in nucleus
Transport to cytoplasm
CYTOPLASM
mRNA in cytoplasm
Degradation
of mRNA
Translation
Polypeptide
Cleavage
Chemical modification
Transport to cellular
destination
Active protein
Degradation of protein
Degraded protein

21. Regulation of Chromatin Structure
Structural organization of chromatin not only packs a cell’s DNA into a compact form that fits inside the nucleus, but also is important in helping regulate gene expression
Genes within highly packed heterochromatin are usually not expressed

22. Chemical modifications to histones and DNA of chromatin influence both chromatin structure and gene expression
N terminus end of each histone molecule protrudes outward from the nucleosome
Tails are accessible to various modifying enzymes

23. Histone Modification
In histone acetylation, acetyl groups are attached to positively charged lysines in histone tails
Deacetylation: removal of acetyl groups
When histone tails are acetylated, their positive charges are neutralized and no longer bind to neighboring nucleosomes
This process seems to loosen chromatin structure, thereby promoting the initiation of transcription

24. LE 19-4 Histone tails DNA double helix Amino acids available
for chemical
modification
Histone tails protrude outward from a nucleosome
Unacetylated histones
Acetylated histones
Acetylation of histone tails promotes loose chromatin
structure that permits transcription

25. DNA Methylation
DNA methylation, the addition of methyl groups to certain bases in DNA, is associated with reduced transcription in some species
DNA that is inactive is generally highly methylated compared with DNA that is actively transcribed
In some species, DNA methylation causes long- term inactivation of genes in cellular differentiation
Deficient DNA methylation leads to abnormal embryonic development

26. In genomic imprinting, methylation turns off either the maternal or paternal alleles of certain genes at the start of development

27. Epigenetic Inheritance
Although the chromatin modifications just discussed do not alter DNA sequence, they may be passed to future generations of cells
The inheritance of traits transmitted by mechanisms not directly involving the nucleotide sequence is called epigenetic inheritance
Enzymes that modify chromatin structure appear to be important parts of the cell’s machinery for regulating transcription

28. Regulation of Transcription Initiation
Chromatin-modifying enzymes provide initial control of gene expression by making a region of DNA either more or less able to bind the transcription machinery
Once a gene is optimally modified for expression, the initiation of transcription is the most important and universally used state at which gene expression is regulated

29. Organization of a Typical Eukaryotic Gene
Associated with most eukaryotic genes are control elements, segments of noncoding DNA that help regulate transcription by binding certain proteins
Control elements and the proteins they bind are critical to the precise regulation of gene expression in different cell types

30. (distal control elements)
Enhancer
(distal control elements)
Proximal
control elements
Poly-A signal
sequence
Termination
region
Exon
Intron
Exon
Intron
Exon
DNA
Upstream
Downstream
Promoter
Transcription
Poly-A signal
Primary RNA
transcript
(pre-mRNA)
Exon
Intron
Exon
Intron
Exon
Cleaved 3¢ end
of primary
transcript 5¢
RNA processing:
Cap and tail added;
introns excised and
exons spliced together
Intron RNA
Coding segment
mRNA 3¢
Start
codon
Stop
codon
5¢ Cap
5¢ UTR
(untranslated
region)
3¢ UTR
(untranslated
region)
Poly-A
tail

31. The Roles of Transcription Factors
To initiate transcription, eukaryotic RNA polymerase requires the assistance of proteins called transcription factors
General transcription factors are essential for the transcription of all protein-coding genes
In eukaryotes, high levels of transcription of particular genes depend on control elements interacting with specific transcription factors

32. Enhancers and Specific Transcription Factors
Proximal control elements are located close to the promoter
Distal control elements, groups of which are called enhancers, may be far away from a gene or even in an intron
A gene may have multiple enhancers—each active at a different time or in a different cell type or location in the organism
Animation: Initiation of Transcription

33. An activator is a protein that binds to an enhancer and stimulates transcription of a gene
Some transcription factors function as repressors, inhibiting expression of a particular gene
Some activators and repressors act indirectly by influencing chromatin structure
Gene present in region of chromatin with high levels of histone acetylation is able to bind the transcription machinery

34. Repressors cause inhibition of gene expression in several ways:
Block binding of activators
Bind directly to their own control elements in an enhancer
Turn of transcription in presence of activators

35. Hundreds of transcription factors have been discovered in eukaryotes
Two common structural elements:
DNA-binding domain
Part of 3D protein shape that binds to DNA
Activation domains
Bind other regulatory proteins

36. LE 19-6 Distal control element Activators Promoter Gene DNA Enhancer
TATA
box
General
transcription
factors
DNA-bending
protein
Group of
mediator proteins
RNA
polymerase II
RNA
polymerase II
Transcription
Initiation complex
RNA synthesis

37. Combinatorial Control of Gene Activation
A particular combination of control elements can activate transcription only when the appropriate activator proteins are present
Only about a dozen different nucleotide sequences found in control elements
Each enhancer is made of about ten control elements, each of which can bind only one or two specific transcription factors

38. LE 19-7 Liver cell nucleus Lens cell nucleus Available activators
Enhancer
Promoter
Control
elements
Albumin
gene
Albumin
gene not
expressed
Crystallin
gene
Albumin
gene
expressed
Crystallin gene
not expressed
Crystallin gene
expressed
Liver cell
Lens cell

39. Coordinately Controlled Genes
How does the prokaryotic cell deal with genes of related function that need to be turned off or on at the same time?
Controlled genes often clustered into an operon, which is regulated by a single promoter and transcribed into a single mRNA molecule
Genes are expressed together, and encoded proteins are produced at the same time
Only found in prokaryotic cells

40. How does the eukaryotic cell deal with genes of related function that need to be turned off or on at the same time?
Research has found that some co-expressed genes are clustered near one another on the same chromosome
Each gene has its own promoter and is individually transcribed

41. Systems for coordinating gene regulation probably arose early in evolutionary history and evolved by the duplication and distribution of control elements within the genome

42. Mechanisms of Post-Transcriptional Regulation
Transcription alone does not account for gene expression
More and more examples are being found of regulatory mechanisms that operate at various stages after transcription
Such mechanisms allow a cell to fine-tune gene expression rapidly in response to environmental changes

43. Animation: RNA Processing
RNA processing provide several opportunities for regulating gene expression that are not possible in prokaryotes
In alternative RNA splicing, different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns
Regulatory proteins specific to a cell type control intron-exon choices by binding to a sequence within the primary transcript
Animation: RNA Processing

44. LE 19-8
Exons
DNA
Primary
RNA
transcript
RNA splicing or
mRNA

45. mRNA Degradation
The life span of mRNA molecules in the cytoplasm is a key to determining the protein synthesis
The mRNA life span is determined in part by sequences in the leader and trailer regions
Prokaryotic cells: mRNA degraded within a few minutes of their synthesis
Short life span allows prokaryotes to quickly change proteins being made in response to environmental changes

46. Eukaryotic cells: degraded within hours, days, or weeks
mRNA breakdown begins with shortening of poly-A tail
Then, enzymes remove 5’ cap
After, nuclease enzymes rapidly chew up the mRNA
Sequences that affect mRNA life span are found in UTR at the 3’ end

47. Animation: Blocking Translation Animation: mRNA Degradation
RNA interference by single-stranded microRNAs (miRNAs) can lead to degradation of an mRNA or block its translation
miRNA hydrogen bonds with itself to form double-stranded hairpin structure
Enzyme then cuts it into two strands:
One strand degraded
Other strand binds with protein complex to degrade mRNA or block translation of that mRNA strand
Animation: Blocking Translation
Animation: mRNA Degradation

48. The phenomenon of inhibition of gene expression by RNA molecules is called RNA interference (RNAi)
Due to small interfering RNAs (siRNAs)
Believed that the RNAi pathway originated as a natural defense against infection by RNA viruses

49. LE 19-9 Protein complex Degradation of mRNA Dicer OR miRNA Target mRNA
Hydrogen
bond
Blockage of translation

50. Initiation of Translation
Translation presents another opportunity for regulating gene expression
The initiation of translation of selected mRNAs can be blocked by regulatory proteins that bind to sequences or structures of the mRNA
Found in UTR at the 5’ end
Alternatively, translation of all mRNAs in a cell may be regulated simultaneously
Involves activation or inactivation of protein factors

51. Protein Processing and Degradation
Final opportunity for controlling gene expression occurs after translation
Eukaryotic polypeptides have to be processed to yield functional protein molecules
Proteins have to undergo chemical modifications to make them functional
Regulatory proteins activated or inactivated
Proteins on surface of animal cells require sugar
Cell-surface proteins need to be moved
Animation: Protein Processing
Animation: Protein Degradation

52. Length of time each protein functions in the cell is strictly regulated by means of selective degradation
To mark a protein for destruction, the cell attaches molecules of ubiquitin, a small protein
Proteasomes are giant protein complexes that recognize the ubiquitin-tagged protein molecules and degrade them

53. LE 19-10 Proteasome and ubiquitin to be recycled Ubiquitin Proteasome
Protein to
be degraded
Ubiquitinated
protein
Protein
fragments
(peptides)
Protein entering a
proteasome