Gene Regulation

1
Gene Regulation

• Chapter 14

2
Learning Objective 1

• Why do bacterial and eukaryotic cells have
  different mechanisms of gene regulation?

3
Prokaryotes

• Bacterial cells
• grow rapidly
• have a short life span
• Transcriptional-level control
• usually regulates gene expression

4
Eukaryotic Cells

• Have long life span
• respond to many different stimuli
• One gene
• may be regulated in different ways
• Transcriptional-level control
• and control at other levels of gene expression

5
KEY CONCEPTS

• Cells can synthesize thousands of proteins
• but not all proteins are required in all cells
• Cells regulate which parts of the genome will be
  expressed, and when

6
Learning Objective 2

• What is an operon?
• What are the functions of the operator and
  promoter regions?

7
Operon

• A gene complex
• structural genes with related functions
• controlled by closely linked DNA sequences
• Regulated genes in bacteria
• are organized into operons

8
Promoter Region

• Each operon has a promoter region
• upstream from protein-coding regions
• where RNA polymerase binds to DNA before
  transcription

9
Operator (1)

• Regulatory switch for transcriptional-level
  control of operon
• Repressor protein
• binds to operator sequence
• prevents transcription

10
Operator (2)

• RNA polymerase
• bound to promoter
• is blocked from transcribing structural genes
• If repressor is not bound to operator
• transcription proceeds

11
Learning Objective 3

• What is the difference between inducible,
  repressible, and constitutive genes?

12
Inducible Genes (1)

• An inducible operon
• such as lac operon
• is normally turned off
• Repressor protein
• is synthesized in active form
• binds to operator

13
Inducible Genes (2)

• If lactose is present
• is converted to allolactose (inducer)
• binds to repressor protein
• changes repressors shape
• Altered repressor
• cannot bind to operator
• operon is transcribed

14
The lac Operon
15

lac operon
Repressor gene
Promoter
Operator
lac Z
lac Y
lac A
DNA
Repressor protein
Transcription
mRNA
Translation
Ribosome
Fig. 14-2a, p. 307
16


lac operon
Repressor gene
Promoter
Operator
lac Z
lac Y
lac A
RNA polymerase
Transcription
mRNA
mRNA
Translation
Transacetylase
Inducer (allolactose)
Lactose permease
ß-galactosidase
Repressor protein (inactive)
Enzymes for lactose metabolism
Fig. 14-2b, p. 307
17
Repressible Genes (1)

• A repressible operon (trp operon)
• is normally turned on
• Repressor protein
• is synthesized in inactive form
• cannot bind to operator
• A metabolite (metabolic end product)
• acts as corepressor

18
Repressible Genes (2)

• With high intracellular corepressor levels
• corepressor molecule binds to repressor
• changes repressors shape
• Altered repressor
• binds to operator
• turns off transcription of operon

19
The trp Operon
20

trp operon
Repressor gene
Operator
trp E
trp D
trp C
trp B
trp A
Promoter
DNA
RNA polymerase
Transcription
mRNA
mRNA
Translation
Repressor protein (inactive)
Enzymes of the tryptophan biosynthetic pathway
Tryptophan
(a) Intracellular tryptophan levels low.
Fig. 14-4a, p. 310
21

trp operon
Repressor gene
Promoter
Operator
trp E
trp D
trp C
trp B
trp A
DNA
Active repressor corepressor complex
mRNA
Inactive repressor protein
Tryptophan (corepressor)
(b) Intracellular tryptophan levels high.
Fig. 14-4b, p. 310
22
Constitutive Genes (1)

• Are neither inducible nor repressible
• active at all times
• Regulatory proteins
• produced constitutively
• catabolite activator protein (CAP)
• repressor proteins

23
Constitutive Genes (2)

• Regulatory proteins
• recognize and bind to specific base sequences in
  DNA
• Activity of constitutive genes
• controlled by binding RNA polymerase to promoter
  regions

24
Learning Objective 4

• What is the difference between positive and
  negative control?
• How do both types of control operate in
  regulating the lac operon?

25
Negative Control

• Repressible and inducible operons are under
  negative control
• When repressor protein binds to operator
• transcription of operon is turned off

26
Positive Control (1)

• Some inducible operons are under positive control
  
• Activator protein binds to DNA
• stimulates transcription of gene

27
Positive Control (2)

• CAP activates lac operon
• binds to promoter region
• stimulates transcription by tightly binding RNA
  polymerase
• To bind to lac operon
• CAP requires cyclic AMP (cAMP)
• cAMP levels increase
• as glucose levels decrease

28
Positive Control
29

Promoter
RNA polymerase binding site
CAP- binding site
Repressor gene
Operator
lac Z
lac Y
lac A
DNA
mRNA
RNA polymerase binds poorly
CAP (inactive)
Allolactose
Repressor protein (inactive)
(a) Lactose high, glucose high, cAMP low.
Fig. 14-5a, p. 311
30

Promoter
CAP- binding site
RNA polymerase binding site
Repressor gene
Operator
lac Z
lac Y
lac A
DNA
RNA polymerase binds efficiently
Transcription
mRNA
mRNA
CAP
Translation
Galactoside transacetylase
cAMP
Lactose permease
ß -galactosidase
Allolactose
Enzymes for lactose metabolism
Repressor protein (inactive)
(b) Lactose high, glucose low, cAMP high.
Fig. 14-5b, p. 311
31
Binding CAP
32

DNA
cAMP
CAP dimer
Fig. 14-6, p. 312
33
Learning Objective 5

• What are the types of posttranscriptional control
  in bacteria?

34
Posttranscriptional Controls in Bacteria

• Translational control
• regulates translation rate of particular mRNA
• Posttranslational controls
• include feedback inhibition of key enzymes in
  metabolic pathways

35
KEY CONCEPTS

• Prokaryotes regulate gene expression in response
  to environmental stimuli

36
KEY CONCEPTS

• Gene regulation in prokaryotes occurs primarily
  at the transcription level

37
Learning Objective 6

• Discuss the structure of a typical eukaryotic
  gene and the DNA sequences involved in regulating
  that gene

38
Eukaryotic Genes

• Are not normally organized into operons
• Regulation occurs at levels of
• Transcription
• mRNA processing
• Translation
• Modifications of protein product

39
Transcription

• Requires
• Transcription initiation site
• where transcription begins
• Promoter
• to which RNA polymerase binds
• In multicellular eukaryotes
• RNA polymerase binds to promoter (TATA box)

40
Transcription
41

TATA box
Transcription initiation site
T T
TATA A
UPE
A A
pre-mRNA
(a) Eukaryotic promoter elements.
Fig. 14-9a, p. 316
42

TATA box
Transcription initiation site
T T
TATA A
UPE
A A
pre-mRNA
(b) A weak eukaryotic promoter.
Fig. 14-9b, p. 316
43

Transcription initiation site
TATA box
T T
TATA A
A A
UPE
UPE
UPE
UPE
pre-mRNA
(c) A strong eukaryotic promoter.
Fig. 14-9c, p. 316
44

TATA box
Transcription initiation site
T T
TATA A
Enhancer
UPE
UPE
A A
pre-mRNA
(d) A strong eukaryotic promoter plus an enhancer.
Fig. 14-9d, p. 316
45
Regulated Eukaryotic Gene

• Promoter
• RNA polymerase-binding site
• short DNA sequences (upstream promoter elements
  (UPEs) or proximal control elements)
• UPEs
• number and types within promoter region determine
  efficiency of promoter

46
Enhancers (1)

• Located far away from promoter
• control some eukaryotic genes
• Help form active transcription initiation complex

47
Enhancers (2)

• Specific regulatory proteins
• bind to enhancer elements
• activate transcription by interacting with
  proteins bound to promoters

48
Enhancers
49

Enhancer
Target proteins
RNA polymerase
TATA box
DNA
(a) Little or no transcription.
Fig. 14-11a, p. 317
50

Enhancer
Activator (transcription factor)
TATA box
DNA
(b) High rate of transcription.
Fig. 14-11b, p. 317
51
Learning Objective 7

• In what ways may eukaryotic DNA-binding proteins
  bind to DNA?

52
Transcription Factors

• DNA-binding protein regulators control eukaryotic
  genes
• some transcriptional activators
• some transcriptional repressors

53
Transcription Factors

• Each has DNA-binding domain
• 3 types of regulatory proteins
• Helix-turn-helix
• Zinc fingers
• Leucine zippers

54
Helix-Turn-Helix

• Inserts one helix into DNA

55

Turn
a -helix
DNA
(a) Helix-turn-helix.
Fig. 14-10a, p. 317
56
Zinc Fingers

• Loops of amino acids
• held together by zinc ions
• each loop has a-helix that fits into DNA

57

COO
Finger 2
Finger 3
Zinc ion
Finger 1
NH3
DNA
(b) Zinc fingers.
Fig. 14-10b, p. 317
58
Leucine Zipper Proteins

• Associate as dimers that insert into DNA

59

Leucine zipper region
DNA
(c) Leucine zipper.
Fig. 14-10c, p. 317
60
Learning Objective 8

• How may a change in chromosome structure affect
  the activity of a gene?

61
Gene Activity (1)

• Changes in chromosome structure
• inactivates genes
• Heterochromatin
• densely packed regions of chromosomes
• contain inactive genes

62
Gene Activity (2)

• Active genes
• associated with loosely packed chromatin
  structure (euchromatin)
• Cells change chromatin structure
• from heterochromatin to euchromatin
• by chemically modifying histones (proteins
  associated with DNA to form nucleosomes)

63
Chromatin Structure
64

Heterochromatin genes silent
Chromatin decondensation
Nucleosome
Histones
DNA
Transcribed region
Euchromatin genes active
Fig. 14-7, p. 314
65
Gene Activity (3)

• Histone tail
• string of amino acids that extends from the
  DNA-wrapped nucleosome
• Methyl groups, acetyl groups, sugars, and
  proteins
• may chemically attach to the histone tail
• may expose or hide genes (turn on or off)

66
Gene Activity (4)

• Epigenetic inheritance
• changes how a gene is expressed
• important mechanism of gene regulation
• DNA methylation
• perpetuates gene inactivation
• patterns repeat in successive cell generations
• mechanism for epigenetic inheritance

67
Gene Amplification

• Some genes
• products are required in large amounts
• have multiple copies in the chromosome
• Gene amplification
• some cells selectively amplify genes by DNA
  replication

68
Gene Amplification
69

Drosophila chorion gene
Gene amplification by repeated DNA replication of
chorion gene region
Chorion gene in ovarian cell
Fig. 14-8, p. 315
70
Learning Objective 9

• How may a gene in a multicellular organism
  produce different products in different types of
  cells?

71
Differential mRNA Processing

• Single gene produces different forms of protein
  in different tissues
• depending on how pre-mRNA is spliced
• Gene contains a segment that can be either intron
  or exon
• as intron, sequence is removed
• as exon, sequence is retained

72
Differential mRNA Processing
73

Potential splice sites
Exon or intron
Exon
Intron
Exon
pre-mRNA
Differential mRNA processing
Exon
Exon
Exon
Exon
Exon
Functional mRNA in tissue A
Functional mRNA in tissue B
Fig. 14-12, p. 318
74
Learning Objective 10

• What types of regulatory controls operate in
  eukaryotes after mature mRNA is formed?

75
mRNA Stability

• Certain regulatory mechanisms increase RNA
  stability
• allowing more protein synthesis before mRNA
  degradation
• Sometimes under hormonal control

76
Posttranslational Control (1)

• In eukaryotic gene expression
• feedback inhibition
• modification of protein structure
• Protein function change
• by kinases adding phosphate groups
• by phosphatases removing phosphates

77
Protein Degradation (1)

• Proteins targeted for destruction
• covalently bonded to ubiquitin
• Protein tagged by ubiquitin
• degraded in a proteasome

78
Protein Degradation (2)

• Proteasome
• large macromolecular structure
• recognizes ubiquitin tags
• Proteases
• protein-degrading enzymes
• associated with proteasomes
• degrade protein into peptide fragments

79
Protein Degradation
80

Target protein
Ubiquitin
Ubiquitin molecules attach to protein tar- geted
for degradation.
1
Ubiquitinylated protein
Protein enters proteasome.
2
Proteasome
3
Ubiquitins are released and available for reuse.
Protein is degraded into peptide fragments.
Peptide fragments
Fig. 14-13, p. 318
81

Stepped Art
Fig. 14-13, p. 318
82
KEY CONCEPTS

• Gene regulation in eukaryotes occurs at the
  levels of transcription, posttranscription,
  translation, and posttranslation

83
Animation Controls of Eukaryotic Gene Expression
CLICKTO PLAY