How Genes are Controlled Muse 2009. CONTROL OF GENE EXPRESSION

How Genes are Controlled

Muse 2009

CONTROL OF GENE EXPRESSION

– Gene expression is the overall process of information flow from genes to proteins

– Mainly controlled at the level of transcription

– A gene that is “turned on” is being transcribed to produce mRNA that is translated to make its corresponding protein

– Organisms respond to environmental changes by controlling gene expression

11.1 Proteins interacting with DNA turn prokaryotic genes on or off in response to

environmental changes

11.1 Proteins interacting with DNA turn prokaryotic genes on or off in response

to environmental changes– An operon is a group of genes under

coordinated control in bacteria

– The lactose (lac) operon includes

– Three adjacent genes for lactose-utilization enzymes

– Promoter sequence where RNA polymerase binds

– Operator sequence is where a repressor can bind and block RNA polymerase action


to environmental changes

Regulation of the lac operon– Regulatory gene codes for a repressor protein

– In the absence of lactose, the repressor binds to the operator and prevents RNA polymerase action

– Lactose inactivates the repressor, so the operator is unblocked


to environmental changes– Types of operon control

– Inducible operon (lac operon)– Active repressor binds to the operator

– Inducer (lactose) binds to and inactivates the repressor

– Repressible operon (trp operon)– Repressor is initially inactive

– Corepressor (tryptophan) binds to the repressor and makes it active

– For many operons, activators enhance RNA polymerase binding to the promoter (ie NAC)

DNA

DNA

RNA polymerasecannot attach to promoter

Lactose-utilization genesPromoter OperatorRegulatorygene

OPERON

Protein

mRNA

Inactiverepressor

Lactose Enzymes for lactose utilization

RNA polymerasebound to promoter

Operon turned on (lactose inactivates repressor)

mRNA

Activerepressor

Operon turned off (lactose absent)

Protein

DNA

RNA polymerasecannot attach to promoter

Lactose-utilization genesPromoter OperatorRegulatorygene

OPERON

mRNA

Activerepressor

Operon turned off (lactose absent)

Protein

DNA

Protein

Inactiverepressor

Lactose Enzymes for lactose utilization

RNA polymerasebound to promoter

Operon turned on (lactose inactivates repressor)

mRNA

DNA

Inactiverepressor

Activerepressor

Inactiverepressor

Activerepressor

Lactose

Promoter

Tryptophan

Operator Gene

lac operon trp operon

11.2 Differentiation results from the expression of different combination

genes

– Differentiation involves cell specialization, in both structure and function

– Differentiation is controlled by turning specific sets of genes on or off

11.3 DNA packing in eukaryotic chromosomes helps regulate gene

expression– Eukaryotic chromosomes undergo multiple levels of folding and

coiling, called DNA packing – Nucleosomes are formed when DNA is wrapped around

histone proteins– “Beads on a string” appearance– Each bead includes DNA plus 8 histone molecules– String is the linker DNA that connects nucleosomes

– Tight helical fiber is a coiling of the nucleosome string– Supercoil is a coiling of the tight helical fiber– Metaphase chromosome represents the highest level of

packing– DNA packing can prevent transcription

11.3 DNA packing in eukaryotic chromosomes helps regulate

gene expression

Animation: DNA Packing

DNA double helix(2-nm diameter)

“Beads ona string”

Linker

Histones

Metaphasechromosome

Tight helical fiber(30-nm diameter)

Nucleosome(10-nm diameter)

Supercoil(300-nm diameter)

700 nm

11.4 In female mammals, one X chromosome is inactive in each somatic cell

– X-chromosome inactivation

– In female mammals, one of the two X chromosomes is highly compacted and transcriptionally inactive

– Random inactivation of either the maternal or paternal chromosome

– Occurs early in embryonic development and all cellular descendants have the same inactivated chromosome

– Inactivated X chromosome is called a Barr body

– Tortoiseshell fur coloration is due to inactivation of X chromosomes in heterozygous female cats

Two cell populationsin adult

X chromosomes

Early embryo

Allele forblack fur

Inactive X

Black furAllele fororange fur

Orange fur

Cell divisionand random

X chromosomeinactivation Active X

Inactive X

Active X

11.5 Complex assemblies of proteins control

eukaryotic transcription

– Eukaryotic genes – Each gene has its own promoter and terminator

– Are usually switched off and require activators to be turned on

– Are controlled by interactions between numerous regulatory proteins and control sequences

11.5 Complex assemblies of proteins control

eukaryotic transcription

– Regulatory proteins that bind to control sequences

– Transcription factors promote RNA polymerase binding to the promoter

– Activator proteins bind to DNA enhancers and interact with other transcription factors

– Silencers are repressors that inhibit transcription

– Control sequences

– Promoter

– Enhancer

– Related genes located on different chromosomes can be controlled by similar enhancer sequences

11.5 Complex assemblies of proteins control eukaryotic

transcription

Animation: Initiation of Transcription

Regions on the DNA called enhancers control genes that may be quitedistant.Enhancer binding proteins may have multiple controls built into them

Enhancers

Otherproteins

DNA

Transcriptionfactors

Activatorproteins

RNA polymerase

Promoter

Gene

Bendingof DNA

Transcription

11.6 Eukaryotic RNA may be spliced in more than one way

– Alternative RNA splicing – Production of different mRNAs from the same

transcript

– Results in production of more than one polypeptide from the same gene

– Can involve removal of an exon with the introns on either side

Animation: RNA Processing

1

or

Exons

DNA

RNA splicing

RNAtranscript

mRNA

2 3 4 5

1 2 3 4 5

1 2 4 51 2 3 5

11.7 Small RNAs play multiple roles in

controlling gene expression

– RNA interference (RNAi)

– Prevents expression of a gene by interfering with translation of its RNA product

– Involves binding of small, complementary RNAs to mRNA molecules

– Leads to degradation of mRNA or inhibition of translation

– MicroRNA

– Single-stranded chain about 20 nucleotides long

– Binds to protein complex

– MicroRNA + protein complex binds to complementary mRNA to interfere with protein production

11.7 Small RNAs play multiple roles in controlling gene expression

Animation: Blocking Translation

Animation: mRNA Degradation

Anti-sense RNA and snRNAi

miRNA 1

Translation blockedORmRNA degraded

Target mRNA

Protein

miRNA-proteincomplex

2

3 4

11.8 Translation and later stages of gene expression are also subject

to regulation

– Control of gene expression also occurs with – Breakdown of mRNA

– Initiation of translation

– Protein activation

– Protein breakdown

Folding ofpolypeptide andformation ofS—S linkages

Initial polypeptide(inactive)

Folded polypeptide(inactive)

Active formof insulin

Cleavage

Disulfide bonds

Quartenary structure

11.9 Review: Multiple mechanisms regulate

gene expression in eukaryotes

– Many possible control points exist; a given gene may be subject to only a few of these

– Chromosome changes (1)– DNA unpacking

– Control of transcription (2)– Regulatory proteins and control sequences

– Control of RNA processing– Addition of 5’ cap and 3’ poly-A tail (3)

– Splicing (4)

– Flow through nuclear envelope (5)



– Many possible control points exist; a given gene may be subject to only a few of these

– Breakdown of mRNA (6)

– Control of translation (7)

– Control after translation – Cleavage/modification/activation of proteins (8)

– Breakdown of protein (9)

– Applying Your KnowledgeFor each of the following, determine whether an increase or decrease in the amount of gene product is expected

– The mRNA fails to receive a poly-A tail during processing in the nucleus

– The mRNA becomes more stable and lasts twice as long in the cell cytoplasm

– The region of the chromatin containing the gene becomes tightly compacted

– An enzyme required to cleave and activate the protein product is missing



– Applying Your KnowledgeFor each of the following, determine whether an increase or decrease in the amount of gene product is expected

– The mRNA fails to receive a poly-A tail during processing in the nucleus ---------

– The mRNA becomes more stable and lasts twice as long in the cell cytoplasm ++++++

– The region of the chromatin containing the gene becomes tightly compacted --------

– An enzyme required to cleave and activate the protein product is missing +++++ feedback



11.9 Review: Multiple mechanisms regulate gene expression in

eukaryotes

Animation: Protein Processing

Animation: Protein Degradation

NUCLEUS

DNA unpackingOther changes to DNA

Addition of cap and tail

Chromosome

Gene

RNA transcript

GeneTranscription

Intron

Exon

Splicing

CapmRNA in nucleus

Tail

Flow throughnuclear envelope

Broken-downmRNA

CYTOPLASM

Breakdown of mRNA

Translation

mRNA in cytoplasm

Broken-downprotein

Cleavage / modification /activation

Breakdown of protein

Polypeptide

Active protein

NUCLEUS

DNA unpackingOther changes to DNA

Addition of cap and tail

Chromosome

Gene

RNA transcript

GeneTranscription

Intron

Exon

Splicing

CapmRNA in nucleus

Tail

Flow throughnuclear envelope

Broken-downmRNA

CYTOPLASM

Breakdown of mRNA

Translation

mRNA in cytoplasm

Broken-downprotein

Cleavage / modification /activation

Breakdown of protein

Polypeptide

Active protein

11.10 Cascades of gene expression direct the development

of an animal

– Role of gene expression in fruit fly development

– Orientation from head to tail

– Maternal mRNAs present in the egg are translated and influence formation of head to tail axis

– Segmentation of the body

– Protein products from one set of genes activate other sets of genes to divide the body into segments

– Production of adult features

– Homeotic genes are master control genes that determine the anatomy of the body, specifying structures that will develop in each segment

Animation: Development of Head-Tail Axis in Fruit Flies

11.10 Cascades of gene expression direct the development of an

animal

Animation: Cell Signaling

Head of a normal fruit fly

Antenna

Eye

Head of a developmental mutant

Leg

Egg cellwithin ovarianfollicle

Follicle cells

“Head”mRNA

Proteinsignal

Egg cell

Gene expression1

Cascades ofgene expression2

Embryo Bodysegments

Adult fly

Gene expression

3

4

DNA microarrays test for the transcription of many genes at

once– DNA microarray

– Contains DNA sequences arranged on a grid

– Used to test for transcription– mRNA from a specific cell type is isolated

– Fluorescent cDNA is produced from the mRNA

– cDNA is applied to the microarray

– Unbound cDNA is washed off

– Complementary cDNA is detected by fluorescence

cDNA

Nonfluorescent spotFluorescent

spot

Actual size(6,400 genes)

Each well contains DNAfrom a particular gene

DNA microarray

mRNAisolated

DNA of anexpressed gene

DNA of anunexpressed gene

Reverse transcriptaseand fluorescent DNAnucleotides

1

cDNA madefrom mRNA

2

cDNA appliedto wells

3

UnboundcDNA rinsedaway

4

Global transcription analysis

11.12 Signal transduction pathways convert messages received at the cell surface to responses within

the cell – Signal transduction pathway is a series of molecular changes that converts a signal at the cell’s surface to a response within the cell

– Signal molecule is released by a signaling cell

– Signal molecule binds to a receptor on the surface of a target cell

11.12 Signal transduction pathways convert messages received at the cell surface to responses within

the cell – Relay proteins are activated in a series of reactions

– A transcription factor is activated and enters the nucleus

– Specific genes are transcribed to initiate a cellular response

Animation: Signal Transduction Pathways

Animation: Overview of Cell Signaling

Signaling cell

DNA

Nucleus

Transcriptionfactor(activated)

Signaling molecule Plasma

membraneReceptorprotein

Relayproteins

TranscriptionmRNA

Newprotein

Translation

Target cell

2

1

3

4

5

6

CLONING OF PLANTS AND ANIMALS

Hello Dolly!

11.14 Plant cloning shows that differentiated cells may retain all of their genetic

potential

– Most differentiated cells retain a full set of genes, even though only a subset may be expressed

– Evidence is available from – Plant cloning

– A root cell can divide to form an adult plant

– Animal limb regeneration

– Remaining cells divide to form replacement structures

– Involved dedifferentiation followed by redifferentiation into specialized cells

Root ofcarrot plant

Root cells culturedin nutrient medium

Cell divisionin culture

Single cell

Plantlet Adult plant

11.15 Nuclear transplantation can be used to clone animals

– Nuclear transplantation– Replacing the nucleus of an egg cell or zygote

with a nucleus from an adult somatic cell

– Early embryo (blastocyst) can be used in – Reproductive cloning

– Implant embryo in surrogate mother for development

– New animal is genetically identical to nuclear donor

– Therapeutic cloning

– Remove embryonic stem cells and grow in culture for medical treatments

– Induce stem cells to differentiate

Removenucleusfrom eggcell

Implant blastocyst insurrogate mother

Add somatic cellfrom adult donor

Donorcell

Remove embryonicstem cells fromblastocyst andgrow in culture

Reproductivecloning

Nucleus fromdonor cell

Grow in cultureto produce anearly embryo(blastocyst)

Therapeuticcloning

Clone ofdonor is born

Induce stemcells to formspecialized cells

Removenucleusfrom eggcell

Add somatic cellfrom adult donor

Donorcell Nucleus from

donor cell

Grow in cultureto produce anearly embryo(blastocyst)

Implant blastocyst insurrogate mother

Remove embryonicstem cells fromblastocyst andgrow in culture

Reproductivecloning

Therapeuticcloning

Clone ofdonor is born

Induce stemcells to formspecialized cells

Reproductive cloning has valuable applications, but human reproductive

cloning raises ethical issues– Cloned animals can show differences from their

parent due to a variety of influences during development

– Reproductive cloning is used to produce animals with desirable traits

– Agricultural products

– Therapeutic agents

– Restoring endangered animals

– Human reproductive cloning raises ethical concerns

CAT1 CC

11.17 CONNECTION: Therapeutic cloning can produce stem cells with great

medical potential– Stem cells can be induced to give rise to

differentiated cells – Embryonic stem cells can differentiate into a

variety of types

– Adult stem cells can give rise to many but not all types of cells

– Therapeutic cloning can supply cells to treat human diseases

– Research continues into ways to use and produce stem cells

Blood cells

Different types ofdifferentiated cells

Nerve cells

Adult stemcells in bone

marrow

Different cultureconditions

Culturedembryonicstem cells

Heart muscle cells

THE GENETIC BASIS OF CANCER

11.18 Cancer results from mutations in genes that control cell division

– Mutations in two types of genes can cause cancer

– Oncogenes– Proto-oncogenes normally promote cell division

– Mutations to oncogenes enhance activity

– Tumor-suppressor genes– Normally inhibit cell division

– Mutations inactivate the genes and allow uncontrolled division to occur


– Oncogenes

– Promote cancer when present in a single copy

– Can be viral genes inserted into host chromosomes (src, ras)

– Can be mutated versions of proto-oncogenes, normal genes that promote cell division and differentiation

– Converting a proto-oncogene to an oncogene can occur by– Mutation causing increased protein activity

– Increased number of gene copies causing more protein to be produced

– Change in location putting the gene under control of new promoter for increased transcription


– Tumor-suppressor genes– Promote cancer when both copies are mutated

Mutation withinthe gene

Hyperactivegrowth-stimulatingprotein in normalamount

Proto-oncogene DNA

Multiple copiesof the gene

Gene moved tonew DNA locus,

under new controls

Oncogene New promoter

Normal growth-stimulatingprotein in excess

Normal growth-stimulatingprotein in excess

Mutated tumor-suppressor geneTumor-suppressor gene

Defective,nonfunctioningprotein

Normalgrowth-inhibitingprotein

Cell divisionunder control

Cell division notunder control

11.19 Multiple genetic changes underlie the

development of cancer

– Four or more somatic mutations are usually required to produce a cancer cell

– One possible scenario for colorectal cancer includes

– Activation of an oncogene increases cell division

– Inactivation of tumor suppressor gene causes formation of a benign tumor

– Additional mutations lead to a malignant tumor

1

Colon wall

Cellularchanges:

DNAchanges:

Oncogeneactivated

Increasedcell division

Tumor-suppressorgene inactivated

Growth of polyp

Second tumor-suppressor geneinactivated

Growth of malignanttumor (carcinoma)

2 3

Chromosomes 1mutation

Normalcell

4mutations

3mutations

2mutations

Malignantcell

11.20 Faulty proteins can interfere with normal signal transduction

pathways

– Path producing a product that stimulates cell division

• Product of ras proto-oncogene relays a signal when growth hormone binds to receptor

• Product of ras oncogene relays the signal in the absence of hormone binding, leading to uncontrolled growth

11.20 Faulty proteins can interfere with

normal signal transduction pathways

– Path producing a product that inhibits cell division– Product of p53 tumor-suppressor gene is a

transcription factor

– p53 transcription factor normally activates genes for factors that stop cell division

– In the absence of functional p53, cell division continues because the inhibitory protein is not produced

Growth factor

Protein thatStimulatescell division

Translation

Nucleus

DNA

Target cell

Normal productof ras gene

Receptor

Relayproteins


Hyperactiverelay protein(product ofras oncogene)issues signalson its own

Transcription

Growth-inhibiting factor

Protein thatinhibitscell division

Translation

Normal productof p53 gene

Receptor

Relayproteins


Nonfunctional transcriptionfactor (product of faulty p53tumor-suppressor gene) cannot trigger transcription

Transcription

Protein absent(cell divisionnot inhibited)

Lifestyle choices can reduce the risk of cancer

– Carcinogens are cancer-causing agents that damage DNA and promote cell division

– X-rays and ultraviolet radiation

– Tobacco

– Healthy lifestyle choices– Avoiding carcinogens

– Avoiding fat and including foods with fiber and antioxidants

– Regular medical checkups

Wrap up and Review

• Bacteria arrange multiple genes into operons and control promoters with repressors and activators.

• Eukaryotes have multiple ways to control genes.

Regulatorygene

Promoter

DNA

Gene 1Operator

A typical operon

Gene 2Gene 3

Encodes repressorthat in active formattaches to operator

RNApolymerasebinding site

Switches operonon or off


Early embryoresulting fromnuclear trans-plantation

Surrogatemother

Cloneof donor

Whole Organism Cloning


Early embryoresulting fromnuclear trans-plantation

Embryonicstem cellsin culture

Specializedcells

Therapeutic cloning

Generegulation

(a)prokaryoticgenes oftengrouped into

in eukaryotesmay involve when

abnormalmay lead to

can produce

is a normal gene thatcan be mutated to an

oncogene

can cause

(c)

(g)(f)

multiple mRNAsper genetranscription

femalemammals

(e)

(d)

(b)

operons

areswitchedon/off by

in activeform binds to

controlled byprotein called

occurs inare proteinsthat promote

§ Explain how prokaryotic gene control occurs in the operon

§ Describe the control points in expression of a eukaryotic gene

§ Describe DNA packing and explain how it is related to gene expression

§ Explain how alternative RNA splicing and microRNAs affect gene expression

§ Compare and contrast the control mechanisms for prokaryotic and eukaryotic genes

You should now be able to

§ Distinguish between terms in the following groups: promoter—operator; oncogene—tumor suppressor gene; reproductive cloning— therapeutic cloning

§ Define the following terms: Barr body, carcinogen, DNA microarray, homeotic gene; stem cell; X-chromosome inactivation

§ Describe the process of signal transduction, explain how it relates to yeast mating, and explain how it is disrupted in cancer development


§ Explain how cascades of gene expression affect development

§ Compare and contrast techniques of plant and animal cloning

§ Describe the types of mutations that can lead to cancer

§ Identify lifestyle choices that can reduce cancer risk


Documents

How Genes are Controlled Muse 2009. CONTROL OF GENE EXPRESSION