Chapter 18:

Regulation of Gene Expression

AP Biology

Overview: Conducting the Genetic Orchestra

• Prokaryotes and eukaryotes alter gene expression in response to changes in

environmental conditions

– Multicellular eukaryotes must also develop and maintain multiple cell types

• Though multicellular eukaryotes have different types of cells, all of these

cells contain the same genome

– A significant challenge in the gene regulation of these organisms is

controlling the expression of different subsets of genes to create

different cell types

• Gene expression is often regulated at the stage of transcription, but control at other

levels of gene expression is also important

– RNA molecules play many roles in regulating gene expression in eukaryotes

Concept 18.1: Bacteria often respond to

environmental change by regulating transcription

Fig. 18-2

Regulationof geneexpression

trpE gene

trpD gene

trpC gene

trpB gene

trpA gene

(b) Regulation of enzymeproduction

(a) Regulation of enzymeactivity

Enzyme 1

Enzyme 2

Enzyme 3

Tryptophan

Precursor

Feedback

inhibition

Regulation of Enzyme Activity and Production

• Natural selection has favored bacteria that produce only the products needed by that

cell

– By doing so, these bacteria can conserve resources and energy for other

important tasks

• Metabolic control occurs on 2 levels:

– First, cells can adjust the activity of enzymes that are already present by

feedback inhibition

• In this type of inhibition, the

activity of an enzyme is inhibited

by a product in an anabolic

pathway

– Second, cells can adjust the

production level of certain enzymes

by regulating the expression of the

genes encoding these enzymes

• Gene expression in bacteria is

controlled by the operon model

Operons: The Basic Concept

• A cluster of functionally related genes can be under

coordinated control by a single on-off “switch”

– The regulatory “switch” is a segment of DNA called an

operator usually positioned within the promoter

• The operator controls access of RNA polymerase to the

genes

• An operon is the entire stretch of DNA that includes the

operator, the promoter, and the genes that they control

Repressors

• The operon can be switched off by a protein repressor

– The repressor prevents gene transcription by binding to the

operator and blocking RNA polymerase (no transcription)

– A repressor protein is specific for the operator of a particular

operon

– The repressor is the product of a separate regulatory gene

• Regulatory genes are expressed continuously, although at a

low rate, so that a few repressor molecules are always

present within the cell

Repressors (Continued)

• The binding of repressors to operators is reversible

• Operators can be in one of 2 states at any given time:

• One with repressor bound (“off” mode)

• One without the repressor bound (“on” mode)

• The relative duration of each state depends on the number of active

repressor molecules present

• The repressor can be in an active or inactive form, depending on the presence

of other molecules

• In its inactive form, the repressor has little affinity for its operator

• In its active form, a specific substrate binds to the repressor at an allosteric

site, triggering a change in conformation

• These types of substrates are examples of molecules called

corepressors that cooperates with a repressor protein to switch an

operon off

• Ex) E. coli can synthesize the amino acid tryptophan

Fig. 18-3a

Polypeptide subunits that make upenzymes for tryptophan synthesis

(a) Tryptophan absent, repressor inactive, operon on

DNA

mRNA 5

Protein Inactiverepressor

RNApolymerase

Regulatorygene

Promoter Promoter

trp operon

Genes of operon

Operator

Stop codonStart codon

mRNA

trpA

5

3

trpR trpE trpD trpC trpB

ABCDE

Operon “On”

• By default the trp operon is on and the genes for tryptophan

synthesis are transcribed

• Occurs when tryptophan is absent

• Repressor is inactive

Fig. 18-3b-2

(b) Tryptophan present, repressor active, operon off

Tryptophan(corepressor)

No RNA made

Activerepressor

mRNA

Protein

DNA

Operon “Off”

• When tryptophan is present, it binds to the trp repressor protein,

which turns the operon off

• The repressor is active only in the presence of its corepressor

tryptophan

• Thus the trp operon is turned off (repressed) if tryptophan levels

are high

Repressible and Inducible Operons: Two Types of Negative Gene Regulation

• There are 2 types of negative gene regulation:

– 1) A repressible operon is one that is usually on

• Binding of a repressor to the operator shuts off transcription

– Ex) trp operon

– 2) An inducible operon is one that is usually off

• A molecule called an inducer inactivates the repressor and turns

on transcription

– Ex) lac operon

The lac Operon: An Inducible Operon

• The lac operon is an inducible operon found in E.coli cells

– This operon contains genes that code for enzymes used in the hydrolysis and

metabolism of lactose

• Lactose metabolism begins with the hydrolysis of lactose into its

component monosaccharides – glucose and galactose

– This reaction is catalyzed by the enzyme β- galactosidase

– The gene for β-galactosidase is one of the 3 genes that code for

enzymes that function in lactose utilization

• The entire transcription unit is under the command of a single operator and

promoter

• A regulatory gene located outside the operon called lacI codes for an

allosteric repressor protein that can switch the operon “off” by binding to

the operator

Fig. 18-4b

(b) Lactose present, repressor inactive, operon on

mRNA

Protein

DNA

mRNA 5

Inactiverepressor

Allolactose(inducer)

5

3

RNApolymerase

Permease Transacetylase

lac operon

-Galactosidase

lacYlacZ lacAlacI

The lac Operon: Allolactose as an Inducer

• A molecule called an inducer inactivates the repressor to turn the lac operon on

– For the lac operon, the inducer is an isomer of lactose called allolactose

• Allolactose is formed in small amounts from lactose that enters the cell

– Allolactose binds to the lac repressor and alters its shape, preventing the

repressor from binding to the operator

• Without a bound repressor, the lac operon is transcribed into mRNA, and

the proteins needed for lactose utilization are produced

Fig. 18-4a

(a) Lactose absent, repressor active, operon off

DNA

ProteinActiverepressor

RNApolymerase

Regulatory

genePromoter

Operator

mRNA5

3

NoRNAmade

lacI lacZ

The lac Operon: Lactose Absent

• By itself, the lac repressor is active and switches the lac operon off

– Occurs due to the absence of lactose (and hence allolactose)

Inducible vs. Repressible Enzymes

• The enzymes of the lactose pathway are referred to as inducible enzymes because

their synthesis is induced by a chemical signal (allolactose)

– Inducible enzymes usually function in catabolic pathways

• The enzymes for tryptophan synthesis are referred to as repressible enzymes

because their synthesis is repressed by high levels of the end product

– Repressible enzymes usually function in anabolic pathways

• Regulation of both the trp and lac operons involves negative control of genes

because operons are switched off by the active form of the repressor

– Gene regulation is said to be positive only when a regulatory protein interacts

directly with the genome to switch transcription on

Positive Gene Regulation

• An example of positive gene regulation also involves the lac operon

– When glucose and lactose are both present, E.coli preferentially use glucose,

since the enzymes for glycolysis are always present

• E.coli use lactose as an energy source only when glucose is in short

supply

– When glucose is scarce, a small organic molecule called cyclic AMP (cAMP)

accumulates

• In this case, the lac operon is subject to positive control through a

stimulatory protein called catabolite activator protein (CAP), an activator

of transcription

• CAP is activated by binding

with cAMP, which allows it to

attach to a specific site at the

upstream end of the lac promoter

• This attachment increases the

affinity of RNA polymerase for the

promoter, thus accelerating

transcription

• When glucose levels in the cell increase, cAMP concentration decreases

– Without cAMP, CAP detaches from the lac operon

– Because CAP is inactive, the affinity of RNA polymerase for the promoter of the

lac operon is lowered

– Transcription of the lac operon will

thus proceed only at a low level, even

in the presence of lactose

• Therefore, the lac operon is under dual control:

– Negative control by the lac repressor

(like on-off switch)

• The state of the lac repressor (with or without bound allolactose)

determines whether transcription of the lac operon’s genes will occur at all

– Positive control by CAP (like volume control)

• The state of CAP (with or without bound cAMP) controls the rate of

transcription if the operon is repressor-free

• CAP also helps regulate other operons that encode enzymes used in catabolic

pathways

Dual Control of the lac Operon

Concept 18.2: Eukaryotic gene expression can be

regulated at any stage

Gene Expression and Cell Specialization

• All organisms must regulate which genes are expressed at any

given time

– In multicellular organisms gene expression is essential for

cell specialization

• To perform its role, each cell type must maintain a

specific program of gene expression in which certain

genes are expressed and others are not

Differential Gene Expression

• Almost all the cells in an organism are genetically identical

– Differences between cell types result from differential gene

expression, the expression of different genes by cells with the same

genome

• A typical human cell expresses only ~20% of its genes at any given

time

– Errors in gene expression can lead to diseases including cancer

• Gene expression in eukaryotic cells is regulated at many stages

– Each stage is a potential control point at which gene expression can be

turned on or off, accelerated, or slowed down

Regulation of Gene Expression at Transcription

• In all organisms, a common control point for gene expression is at

transcription

– Regulation at this stage is often in response to signals

(hormones, signaling molecules) coming from outside the cell

– For this reason, the term “gene expression” is often equated with

transcription for both bacteria and eukaryotes

• The greater complexity of eukaryotes, however, also

provides opportunities for regulating gene expression at

many additional stages

Fig. 18-6

DNA

Signal

Gene

NUCLEUS

Chromatin modification

Chromatin

Gene available

for transcription

Exon

Intron

Tail

RNA

Cap

RNA processing

Primary transcript

mRNA in nucleus

Transport to cytoplasm

mRNA in cytoplasm

Translation

CYTOPLASM

Degradation

of mRNA

Protein processing

Polypeptide

Active protein

Cellular function

Transport to cellular

destination

Degradation

of protein

Transcription

• In this diagram, the colored boxes indicate

processes most often regulated

– Each color indicates the type of

molecule affected (blue=DNA,

orange=RNA, purple=protein)

– The nuclear envelope separating

transcription and translation in

eukaryotic cells offers opportunities

for post-transcriptional control in the

form of RNA processing

– In addition, eukaryotes have a greater

variety of control mechanisms

operating before transcription and

after translation

Regulation of Chromatin Structure

• The structural organization of chromatin not only packs a cell’s DNA

into a compact form that fits inside the nucleus, but it also helps

regulate gene expression in several ways

– The location of a gene’s promoter can affect whether a gene will

be transcribed

– In addition, genes within highly packed heterochromatin are

usually not expressed

– Chemical modifications to histones and DNA of chromatin also

influence both chromatin structure and gene expression

Fig. 18-7

Histonetails

DNA

double helix

(a) Histone tails protrude outward from anucleosome

Acetylated histones

Aminoacidsavailablefor chemicalmodification

(b) Acetylation of histone tails promotes loosechromatin structure that permits transcription

Unacetylated histones

Histone Modifications

• There is mounting evidence that chemical modifications to histones play a

direct role in regulation of gene transcription

– The N-terminus of each histone molecule protrudes outward from the

nucleosome

– These histone tails are

accessible to various

modifying enzymes that

catalyze the addition or

removal of specific

chemical groups

Fig. 18-7

Histonetails

DNA

double helix

(a) Histone tails protrude outward from anucleosome

Acetylated histones

Aminoacidsavailablefor chemicalmodification

(b) Acetylation of histone tails promotes loosechromatin structure that permits transcription

Unacetylated histones

Histone Acetylation

• In histone acetylation, acetyl groups (-COCH3) are attached to positively

charged lysines in histone tails

– When lysines are acetylated, their positive charges are neutralized

• As a result, histone tails no longer bind to neighboring

nucleosomes

– This process loosens chromatin structure and allows transcription

proteins easier access to

genes, thereby promoting

the initiation of

transcription

Other Histone Modifications

• Several other chemical groups can be reversibly attached to amino acids in

histone tails, including methyl and phosphate groups

– The addition of methyl groups (methylation) can condense chromatin

– The addition of phosphate groups (phosphorylation) next to a methylated

amino acid can loosen chromatin

• The discovery that these and other modifications to histone tails can affect

chromatin structure and gene expression has led to the histone code hypothesis

– This hypothesis proposes that specific combinations of modifications,

rather than the overall level of histone acetylation, help determine

chromatin configuration

• Chromatin configuration, in turn, has a direct influence on transcription

DNA Methylation

• Some enzymes can methylate certain bases of DNA itself

– DNA methylation, the addition of methyl groups to certain bases in DNA, is

associated with reduced transcription in some species

• Ex) The inactivated mammalian X chromosome is generally more

methylated than DNA that is actively transcribed

– DNA methylation can also cause long-term inactivation of genes in cellular

differentiation

• Methylation patterns are passed on to successive generations of cells, so

that cells keep a chemical record of what occurred during embryonic

development

• A methylation pattern maintained in this way accounts for genomic

imprinting in mammals

– In genomic imprinting, methylation regulates expression of either the

maternal or paternal alleles of certain genes at the start of

development

Epigenetic Inheritance

• Although chromatin modifications 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

• Epigenetic variations might help explain why one

identical twin acquires a genetically based disease

(schizophrenia), but the other does not, despite their

identical genomes

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 the chromatin of a gene is optimally modified for expression, the

initiation of transcription is the next major step at which gene

expression is regulated

• Involves proteins that bind to DNA and either facilitate or inhibit

binding of RNA polymerase (transcription factors)

• Before looking at how eukaryotic cells control transcription,

however, it is helpful to review the structure of a typical eukaryotic

gene

Fig. 18-8-3

Enhancer

(distal control elements)Proximal

control elements

Poly-A signalsequence

Terminationregion

DownstreamPromoter

UpstreamDNA

ExonExon ExonIntron Intron

Exon Exon ExonIntronIntronCleaved 3 endof primarytranscript

Primary RNAtranscript

Poly-Asignal

Transcription

5

RNA processing

Intron RNA

Coding segment

mRNA

5 Cap 5 UTRStart

codonStop

codon 3 UTR Poly-A

tail

3

• In a typical eukaryotic gene, a cluster of proteins called a transcription initiation

complex assembles on the promoter sequence at the “upstream” end of a gene

– One of these proteins (RNA polymerase II) then proceeds to transcribe the

gene, producing a primary RNA transcript

• RNA processing follows, including enzymatic addition of a 5’ cap and a

poly-A tail, as well as splicing out of introns

– 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

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

• Most of these transcription factors do not bind DNA directly, but bind to

proteins (including each other) and RNA polymerase II

• These protein-protein interactions are crucial to the initiation of eukaryotic

transcription

• The interactions of general transcription factors and RNA polymerase II

with a promoter, however, usually only lead to a low rate of transcription

– In eukaryotes, high levels of transcription of particular genes depend on control

elements interacting with another set of proteins called specific transcription

factors

Proximal vs. Distal Control Elements

• Some of these specific transcription factors are called proximal control

elements because they are located close to the promoter

• More distant groups of specific transcription factors called enhancers may

be located 1000s of nucleotides upstream or downstream of a gene, or even

within an intron

– These enhancers are referred to as distal control elements

• A given gene may have multiple enhancers, each active at a

different time or in a different cell type or location within an

organism

• Each enhancer, however, is only associated with one specific gene

Activators and Mediator Proteins

• In eukaryotes, the rate of gene expression can be strongly controlled by the binding

of special proteins to the control elements of enhancers

– An activator is a protein that binds to an enhancer and stimulates transcription

of a gene

• Protein-mediated bending of DNA is thought to bring bound activators in

contact with another group of proteins called mediator proteins

– These mediator proteins will, in turn, interact with proteins at the

promoter

• These multiple protein-protein interactions help assemble and position the

initiation complex on the promoter

Animation: Initiation of Transcription

Fig. 18-9-3

Enhancer TATAbox

PromoterActivators

DNAGene

Distal controlelement

Group ofmediator proteins

DNA-bending

protein

Generaltranscriptionfactors

RNApolymerase II

RNApolymerase II

Transcriptioninitiation complex RNA synthesis

• Step 1: Activator proteins bind to distal control elements grouped as an enhancer in

the DNA

– This particular enhancer has 3 binding sites

• Step 2: A DNA-bending protein brings the bound activator closer to the promoter

– General transcription

factors, mediator

proteins, and RNA

polymerase are

nearby

• Step 3: The activator bind

to certain mediator proteins

and general transcription

factors, helping them form

an active transcription

initiation complex on

the promoter

• Some transcription factors function as repressors, inhibiting expression of a

particular gene

– Some repressors bind directly to control element DNA, like enhancers

• This may block activator binding or turn off transcription even when

activators are bound

– Other repressors block the binding of activators to proteins that allow activators

to bind to DNA

• Some activators and repressors act indirectly by influencing chromatin structure to

promote or silence transcription

– Activators may recruit proteins that acetylate histones near the promoters of

specific genes, thereby promoting transcription

– Some repressors recruit proteins that deacetylate histones, leading to reduced

transcription

Coordinately Controlled Genes in Eukaryotes

• In bacteria, coordinately controlled genes are often clustered in an operon that is

regulated by a single promoter and transcribed in a single mRNA molecule

– In eukaryotic cells, some co-expressed genes are also clustered near one

another of the same chromosomes, but each has its own promoter and control

elements

– More commonly, however, these genes are scattered over different

chromosomes, but each has the same combination of control elements

• Copies of the activators recognize these specific control elements and

promote simultaneous transcription of the genes, no matter where they are

in a genome

– This coordinated control often occurs in response to chemical signals

(ex: hormones) from outside the cell

• These signals bind to receptor proteins, forming complexes that

serves as transcription activators

– Every gene whose transcription is stimulated by a particular chemical

signal has a control element recognized by the same complex,

regardless of its chromosomal location

Mechanisms of Post-Transcriptional Regulation

• Transcription alone does not account for gene

expression

– Regulatory mechanisms can operate at various

stages after transcription

– Such mechanisms allow a cell to fine-tune gene

expression rapidly in response to environmental

changes

Fig. 18-11

or

RNA splicing

mRNA

PrimaryRNAtranscript

Troponin T gene

Exons

DNA

RNA Processing

• RNA processing in the nucleus and export of mature RNA to the cytoplasm provide

several opportunities for regulating gene expression in eukaryotic cells

– 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 each cell type control intron-exon choices

by binding to regulatory sequences

in the primary transcript

– Ex) The troponin T gene

encodes 2 different

proteins

mRNA Degradation

• The life span of mRNA molecules in the cytoplasm is a key to determining

protein synthesis

– Eukaryotic mRNA is more long lived than prokaryotic mRNA, allowing

them to be translated repeatedly in these cells

– The mRNA life span is determined in part by sequences in the leader

and trailer regions

• Nucleotide sequences that affect how long an mRNA remains

intact are often found in the untranslated region (UTR) at the 3’ end

Animation: mRNA Degradation

Initiation of Translation

• Translation presents another opportunity for regulating gene expression

– Occurs most commonly at the initiation stage

• The initiation of translation of some mRNAs can be blocked by regulatory

proteins that bind to sequences or structures within the 5’ UTR of the

mRNA

– This prevents attachment of ribosomes and hence translation

• Alternatively, translation of all mRNAs

in a cell may be regulated simultaneously

– Usually involves activation or inactivation of one or more protein

factors required to initiate translation

• Ex) Translation initiation factors are simultaneously

activated in an egg following fertilization

Animation: Blocking Translation

Protein Processing and Degradation

• The final opportunity for controlling gene expression occurs after translation

– Eukaryotic polypeptides must often be processed to yield functional protein

molecules

• These various types of protein processing include cleavage and chemical

modifications

– Ex) Regulatory proteins are commonly activated or inactivated by the

reversible addition of phosphate groups

– The length of time each protein functions in a cell is also strictly regulated by

means of selective degradation

• To mark a particular protein for destruction, the cell often attaches

molecules of a small protein called ubiquitin to that protein

• Giant protein complexes called proteasomes recognize these ubiquitin-

tagged proteins and degrade them

Animation: Protein Degradation Animation: Protein Processing

Degradation of a Protein by a Proteasome

• Step 1: Multiple ubiquitin molecules are attached to a protein by enzymes in

the cytosol

• Step 2: The ubiquitin-tagged protein is recognized by a proteasome , which

unfolds the protein and sequesters it within a central cavity

• Step 3: Enzymatic components of the proteasome cut the protein into small

peptides, which can be further degraded by other enzymes in the cytosol

Fig. 18-12

Proteasomeand ubiquitinto be recycledProteasome

Proteinfragments(peptides)Protein entering a

proteasome

Ubiquitinatedprotein

Protein tobe degraded

Ubiquitin

Concept 18.3: Noncoding RNAs play multiple roles

in controlling gene expression

Noncoding RNAs and Regulation of Gene Expression

• Only a small fraction (1.5% in humans) of DNA codes for

proteins, rRNA, and tRNA

– A significant amount of the genome may be transcribed into

noncoding RNAs

• Noncoding RNAs regulate gene expression at two

points:

– mRNA translation

– Chromatin configuration

Effects on mRNAs by MicroRNAs and Small Interfering RNAs

• MicroRNAs (miRNAs) are small single-stranded RNA molecules that can bind to

complementary sequences of mRNA

– miRNAs are formed from longer RNA precursors that fold back on themselves,

forming one or more double-stranded hairpin structures, each held together by

hydrogen bonds

– After each hairpin is cut away from the precursor, it is trimmed by an enzyme

called a Dicer into a short, double-stranded fragment of ~20 nucleotide pairs

– One of the two strands is degraded, while the other strand (the miRNA) forms a

complex with one or more proteins

• The miRNA allows this complex to bind to any mRNA molecule with a

complementary sequence

• The miRNA-protein complex then either degrades the target mRNA or

blocks its translation

– The expression of an estimated 1/3 of all human genes may be

regulated by miRNAs

Fig. 18-13

miRNA-proteincomplex(a) Primary miRNA transcript

Translation blocked

Hydrogenbond

(b) Generation and function of miRNAs

Hairpin miRNA

miRNA

Dicer

3

mRNA degraded

5

• Step 1: An enzyme cuts each hairpin from the primary miRNA transcript

• Step 2: A second enzyme called Dicer trims the loop and the single-stranded ends

from the hairpin (cuts are made at the arrows)

• Step 3: One strand of the double-stranded RNA is degraded

– The other strand (miRNA) than forms a complex with one or more proteins

• Step 4: The miRNA in the complex can bind to any target mRNA that contains at

least 6 bases of complementary

sequence

• Step 5: If miRNA and mRNA

bases are complementary all

along their length, the

mRNA is degraded (left)

– If the match is less

complete, translation

is blocked (right)

Small Interfering RNAs

• Gene expression can also be blocked by RNA molecules called small

interfering RNAs (siRNAs)

– The phenomenon of inhibition of gene expression by RNA molecules is

called RNA interference (RNAi)

• siRNAs and miRNAs are similar but form from different RNA

precursors

– miRNA is formed from a single hairpin in a precursor RNA

– siRNAs are formed from much longer double-stranded RNA

molecules, each which gives rise to many siRNAs

Chromatin Remodeling and Silencing of Transcription by Small RNAs

• Small RNA molecules can also cause remodeling of chromatin structure

– siRNAs play a role in heterochromatin formation and can block large

regions of the chromosome

• An RNA transcript produced from DNA is copied into double-

stranded RNA, which is then processed into several siRNAs

• These siRNAs associate with a complex of proteins, which then

recruit enzymes that modify the chromatin, turning it into the highly

condensed heterochromatin

– Small RNAs may also block transcription of specific genes

Concept 18.4: A program of differential gene

expression leads to the different cell types in a multicellular organism

A program of differential gene expression leads to the different cell types in a multicellular organism

• During embryonic development, a fertilized egg gives rise to

many different cell types

– Cell types are organized successively into tissues, organs,

organ systems, and the whole organism

• Gene expression orchestrates this developmental

program, producing cells of different types that form

these higher-level structures

• The transformation from zygote to adult results from 3 interrelated processes:

– Cell division

• The zygote gives rise to a large number of cells through a succession of

mitotic cell division

– Cell differentiation

• These daughter cells then become specialized in structure and function

– Morphogenesis

• These different types of

cells are organized into

tissues and organs in a

particular 3-dimensional

arrangement that give an

organism its shape

Fig. 18-14

(a) Fertilized eggs of a frog (b) Newly hatched tadpole

A Genetic Program for Embryonic Development

• Differential gene expression results from genes being

regulated differently in each cell type

– Materials placed into an egg by the mother set up

a sequential program of gene regulation that is

carried out as cells divide

– This program makes the cells become different

from each other in a coordinated fashion

Fig. 18-15a

(a) Cytoplasmic determinants in the egg

Two differentcytoplasmicdeterminants

Unfertilized egg cell

Sperm

Fertilization

Zygote

Mitoticcell division

Two-celledembryo

Nucleus

Cytoplasmic Determinants and Inductive Signals

• Two sources of information “tell” a cell which genes to express at any given time

during embryonic development

– An egg’s cytoplasm contains RNA, proteins, and other substances that are

distributed unevenly in the unfertilized egg

• These substances include

cytoplasmic determinants,

maternal substances in the

egg that influence early development

– As the zygote divides by mitosis,

cells contain different cytoplasmic

determinants, which lead to

different gene expression

Fig. 18-15b

(b) Induction by nearby cells

Signalmolecule(inducer)

Signaltransductionpathway

Early embryo(32 cells)

NUCLEUS

Signalreceptor

Induction

• The other important source of developmental information is the environment around

the cell, especially signals from nearby embryonic cells

– In the process called induction, signal molecules from embryonic cells cause

transcriptional changes in nearby target cells

• Gene expression is therefore altered in these cells

• Thus, interactions between cells

induce differentiation of

specialized cell types

Animation: Cell Signaling

Sequential Regulation of Gene Expression During Cellular Differentiation

• The term determination refers to the events that lead to the observable

differentiation of a cell

– Once a cell has undergone determination, it is irreversibly committed to its final

fate

• If a committed cell is experimentally placed in another location in the

embryo, it will still differentiate into the cell type that is its normal fate

• Determination precedes differentiation

– Observable cellular differentiation is marked by the expression of genes for

tissue-specific proteins

• These proteins are found only in a specific cell type and give the cell its

characteristic structure and function

Differentiation of Skeletal Muscle Cells

• We can look at the differentiation of skeletal muscle cells as an example:

– Muscle cells develop from embryonic precursor cells that have the potential to

develop into a number of cell types, including cartilage and fat cells

– Once determination occurs, these cells are called myoblasts

• Myoblasts produce muscle-specific proteins and eventually differentiate to

form skeletal muscle cells

– MyoD is one of several “master regulatory genes” that produce proteins that

commit the cell to becoming skeletal muscle

• This gene encodes MyoD protein, a transcription factor that binds to

enhancers of various target genes and stimulates their expression

• Then, secondary transcription factors activate the genes for proteins such

as myosin and actin that confer the unique properties of skeletal muscle

cells

Fig. 18-16-3

Embryonicprecursor cell

Nucleus

OFF

DNA

Master regulatory gene myoD Other muscle-specific genes

OFF

OFFmRNA

MyoD protein(transcriptionfactor)

Myoblast(determined)

mRNA mRNA mRNA mRNA

Myosin, othermuscle proteins,and cell cycle–blocking proteinsPart of a muscle fiber

(fully differentiated cell)

MyoD Anothertranscriptionfactor

• Step 1: Determination - Signals from other cells lead to activation of the master

regulatory gene myoD, allowing the cell to make MyoD protein, which acts as an

activator

– The cell is now called a myoblast and is irreversibly committed to becoming a

skeletal muscle cell

• Step 2: Differentiation - MyoD protein stimulates the myoD gene further and

activates genes encoding for other muscle-specific transcription factors

– These transcription factors

activate genes for muscle

proteins like myosin and actin

– MyoD also turns on

genes that block the

cell cycle, thus

stopping cell division

– The nondividing

myoblasts fuse to

become mature

multinucleate muscle

cells, also called

muscle fibers

Pattern Formation: Setting Up the Body Plan

• For differentiated cells and tissues to function effectively in the organism as a whole,

the organism’s body plan (its 3-D arrangement) must be established and

superimposed on the differentiation process

– Pattern formation is the development of a spatial organization of tissues and

organs

• In animals, pattern formation begins with the establishment of the major

axes

– The three major axes of a bilaterally symmetrical animal include head

and tail, right and left sides, and back and front

• The molecular cues that control pattern formation are collectively known as

positional information

– These cytoplasmic determinants and inductive signals tell a cell its

location relative to the body axes and to neighboring cells

Pattern Formation in Drosophila

• Pattern formation has been extensively studied in

the fruit fly Drosophila melanogaster

– Combining anatomical, genetic, and

biochemical approaches, researchers have

discovered developmental principles common

to many other species, including humans

The Life Cycle of Drosophila

• In Drosophila, cytoplasmic determinants in the unfertilized egg provide

positional information for the placement of anterior-posterior and dorsal-

ventral axes even before fertilization

– This egg develops in the female’s ovary, surrounded by ovarian cells

called nurse cells and follicle cells

• These support cells supply the egg with nutrients, mRNAs, and

other substances needed for development and make the egg shell

• After fertilization, the embryo develops into a segmented larva with

three larval stages

Fig. 18-17bFollicle cell

Nucleus

Eggcell

Nurse cell

Egg celldeveloping withinovarian follicle

Unfertilized egg

Fertilized egg

Depletednurse cells

Eggshell

FertilizationLaying of egg

Bodysegments

Embryonicdevelopment

Hatching

0.1 mm

Segmentedembryo

Larval stage

(b) Development from egg to larva

1

2

3

4

5

• 1) The yellow egg is surrounded by other cells that form a structure called the follicle

within one of the mother’s ovaries

• 2)Nurse cells shrink as they supply nutrients and mRNAs to the developing egg,

which grows larger

– Eventually, the mature egg fills

the egg shell that is secreted by

the follicle cells

• 3)The egg is fertilized within the

mother and then laid

• 4-5) Embryonic development forms

a larva that goes through 3 stages

– The 3rd stage forms a cocoon (not shown), within

which the larva metamorphoses into the

adult shown

Fig. 18-18

Antenna

MutantWild type

Eye

Leg

Genetic Analysis of Early Development: Scientific Inquiry

• Edward B. Lewis, Christiane Nüsslein-Volhard, and Eric Wieschaus won a Nobel

1995 Prize for decoding pattern formation in Drosophila

– These scientists studied mutant flies with developmental defects that led to

extra wings or legs in the wrong places

• They located these mutations on the fly’s genetic map, thus connecting

developmental abnormalities to specific genes

– Their research supplied the first concrete evidence that genes somehow direct

developmental processes

• These genes that

control pattern

formation in the late

embryo, larva, and

adult are called

homeotic genes

• Thirty years later, Nüsslein-Volhard and Wieschaus set out to identify all the

genes that affect segment formation in Drosophila

– They created mutants, conducted breeding experiments, and looked for

corresponding genes

• Breeding experiments were complicated by embryonic lethals,

embryos with lethal mutations

– They found 120 genes essential for normal segmentation

• The researchers were able to group these segmentation genes by

general function, to map them, and to clone many of them for

further study in the lab

Axis Establishment

• Maternal effect genes encode for cytoplasmic determinants that initially establish

the axes of the body of Drosophila

– When these genes are mutant in the mother, any offspring display the mutant

phenotype regardless of the offspring’s own genotype

– These maternal effect genes are also called egg-polarity genes because they

control orientation of the egg and consequently, that of the fly

• One group of these genes sets up the anterior-posterior axis of the

embryo, while a second group establishes the dorsal-ventral axis

– Mutations in these maternal effect genes are generally embryonic lethals

Animation: Development of Head-Tail Axis in Fruit Flies

Fig. 18-19a

T1 T2T3

A1 A2 A3 A4 A5 A6A7

A8

A8

A7 A6 A7

Tail

TailTail

Head

Wild-type larva

Mutant larva (bicoid)

EXPERIMENT

A8

The Bicoid Gene

• One maternal effect gene, the bicoid gene, affects the front half of the body

– An embryo whose mother has a mutant bicoid gene lacks the front half of its

body and has duplicate posterior structures at both ends

• This phenotype suggested that the product of the mother’s bicoid gene is

essential for setting up the anterior end of the fly and therefore might be

concentrated at the future anterior end of the embryo

• This hypothesis is an example of the

morphogen gradient hypothesis, in

which gradients of substances called

morphogens establish an embryo’s axes

and other features of its form

Fig. 18-19b

Fertilization,

translation

of bicoid

mRNA Bicoid protein in earlyembryo

Anterior end

Bicoid mRNA in matureunfertilized egg

100 µm

RESULTS

• Experiment: many embryos and larvae with defects in their body patterns were

obtained

– Some of these defects were due to mutations in the mother’s genes, including

the bicoid (“two-tailed”) gene, which resulted in larvae with two tails and no

head

– The researchers hypothesized that bicoid normally codes for a morphogen

specifying the head (anterior) end of the embryo

• To test this hypothesis, they used molecular techniques to determine

where the mRNA and protein encoded by this gene were found in the

fertilized egg and early embryo

• Results: bicoid mRNA (dark blue) was confined to the anterior end of the

unfertilized egg

– Later in development, Bicoid protein was seen to be concentrated in cells at the

anterior end of the embryo

• Conclusion: the results support the hypothesis that Bicoid protein is a morphogen

specifying formation of head-

specific structures

Importance of Bicoid Research • The bicoid research is important for three reasons:

– It identified a specific protein required for some early steps in pattern formation

• This helped us understand how different regions of the egg can give rise to

cells that go down different developmental pathways

– It increased understanding of the mother’s role in embryo development

– It demonstrated the key developmental principle that a gradient of molecules

can determine polarity and position in the embryo

• In Drosophila, gradients of specific proteins determine the posterior and

anterior ends, as well as the dorsal-ventral axis

• Positional information later establishes a specific number of correctly

oriented segments and triggers the formation of each segment’s

characteristic structures

– The pattern of the adult is abnormal when the genes operating in this

final step are abnormal

Concept 18.5: Cancer results from genetic changes

that affect cell cycle control

Cancer and Gene Regulation

• The gene regulation systems that go wrong during cancer are the very same

systems involved in embryonic development

– Cancer can be caused by mutations to genes that regulate cell growth

and division

• The agents of these changes can be random spontaneous

mutation, or they may be caused by environmental influences,

including chemical carcinogens and X-rays

• Tumor viruses can also cause cancer in animals, including

humans

– Ex) Human papillomaviruses (HPV) are associated with

cervical cancer

Oncogenes and Proto-Oncogenes

• Oncogenes are cancer-causing genes

– Proto-oncogenes are the corresponding normal cellular

genes that are responsible for normal cell growth and

division

• An oncogene usually arises from a genetic change that

leads to an increase either in the amount of the gene’s

protein product or in the activity of each protein

molecule

– Conversion of a proto-oncogene to an oncogene can

lead to abnormal stimulation of the cell cycle

Conversion of Proto-Oncogenes to Oncogenes

• Genetic changes that convert proto-oncogenes to oncogenes fall into 3 main

categories:

– Movement of DNA (translocation) within the genome: if it ends up near an

active promoter, transcription may increase

– Amplification of a proto-oncogene: increases the number of copies of the proto-

oncogene in the cell

– Point mutation in a control element or in the proto-oncogene itself, causing an

increase in gene expression

Fig. 18-20

Normal growth-stimulatingprotein in excess

Newpromoter

DNA

Proto-oncogene

Gene amplification:Translocation ortransposition:

Normal growth-stimulatingprotein in excess

Normal growth-stimulatingprotein in excess

Hyperactive ordegradation-resistant protein

Point mutation:

Oncogene Oncogene

within a control element within the gene

Tumor-Suppressor Genes

• Cells also contain genes known as tumor-suppressor genes whose normal

products inhibit cell division

– The proteins they encode help prevent uncontrolled cell growth

– Mutations that decrease protein products of tumor-suppressor genes may

contribute to cancer onset

• The protein products of tumor-suppressor genes have various functions:

– Repair damaged DNA, preventing the cell from accumulating cancer-causing

mutations

– Control adhesion of cells to one another or to the extracellular matrix, which is

crucial in normal tissues and often absent in cancers

– Inhibit the cell cycle in the cell-signaling pathway

Interference with Normal Cell-Signaling Pathways

• The proteins encoded by many proto-oncogenes and tumor-

suppressor genes are components of cell-signaling pathways

– The products of 2 key genes, ras proto-oncogene and the

p53 tumor-suppressor gene, can be examined in order to

elucidate what goes wrong with the functioning of these

proteins in cancer cells

• Mutations in the ras gene can lead to production of a

hyperactive Ras protein and increased cell division

The Ras Protein

• The Ras (named for rat sarcoma) protein is a G protein that relays a signal

from a growth factor receptor on the plasma membrane to a cascade of

protein kinases

– The cellular response at the end of the pathway is the synthesis of a

protein that stimulates the cell cycle

• Normally, this pathway will not operate unless triggered by the

appropriate growth factor

• Certain mutations in the ras gene can lead to production of a

hyperactive Ras protein that triggers the kinase cascade even in

the absence of growth factor

– Results in increased cell division

Fig. 18-21a

Receptor

Growthfactor

G protein GTP

Ras

GTP

Ras

Protein kinases(phosphorylationcascade)

Transcriptionfactor (activator)

DNA

HyperactiveRas protein(product ofoncogene)issuessignalson its own

MUTATION

NUCLEUS

Gene expression

Protein thatstimulatesthe cell cycle

(a) Cell cycle–stimulating pathway

11

3

4

5

2

• The normal cell cycle-stimulating pathway is triggered by a growth factor (1) that

binds to its receptor (2) in the plasma membrane

• The signal is relayed to a G protein (3) called Ras

– Ras is active when GTP is bound to it

• Ras passes the signal to a series of protein kinases (4)

• The last kinase (5) activates a

transcription activator that turns on

one or more genes for proteins that

stimulate the cell cycle

– Results in excessive

cell division that may

cause cancer

The p53 Gene • If the DNA of a cell is damaged, another signaling pathway blocks the cell cycle until

the damage has been repaired

– Thus, the genes for components of this pathway act as tumor-suppressor

genes

• One is these genes, called the p53 gene, encodes a specific transcription

factor that promotes the synthesis of cell cycle-inhibiting proteins

– Activates a gene called p21 whose product halts the cell cycle by

binding to cyclin-dependent kinases, thus allowing time for DNA repair

– Also turns on genes directly involved in DNA repair

– When DNA damage is irreparable, p53 activates “suicide genes”

whose proteins cause cell death by apoptosis

• A mutation that knocks out the p53 gene can lead to excessive cell growth

and cancer

• In the normal cell cycle-inhibiting pathway, DNA damage (1) is an

intracellular signal that is passed via protein kinases (2) and leads to

activation of p53 (3)

– Activated p53 promotes transcription of the gene for a protein that

inhibits the cell cycle

• This suppression ensures that damaged DNA is not replicated Fig. 18-21b

MUTATION

Protein kinases

DNA

DNA damagein genome

Defective or

missing

transcription

factor, such

as p53, cannot

activate

transcription

Protein that

inhibits

the cell cycle

Activeformof p53

UVlight

(b) Cell cycle–inhibiting pathway

2

3

1

Fig. 18-21

Receptor

Growth

factor

G protein

GTP

Ras

GTP

Ras

Protein kinases

(phosphorylation

cascade)

Transcription

factor (activator)

DNA

Hyperactive

Ras protein

(product of

oncogene)

issues

signals

on its own

MUTATION

NUCLEUS

Gene expression

Protein that

stimulates

the cell cycle

(a) Cell cycle–stimulating pathway

MUTATION

Protein kinases

DNA

DNA damage

in genome

Defective or

missing

transcription

factor, such

as p53, cannot

activate

transcription

Protein that

inhibits

the cell cycle

Active

form

of p53

UV

light

(b) Cell cycle–inhibiting pathway

(c) Effects of mutations

EFFECTS OF MUTATIONS

Cell cycle not

inhibited

Protein absent

Increased cell

division

Protein

overexpressed

Cell cycle

overstimulated

1

2

3

4

5

2

1

3

• Mutations causing deficiencies in

any pathway component can

contribute to the development

of cancer

– Increased cell division that

may lead to cancer can

result if the cell cycle is

over-stimulated via the

cell cycle-stimulating

pathway (a)

– A similar effect can be seen

if the mutation affects the

cell cycle-inhibiting

pathway (b)

The Multistep Model of Cancer Development

• Multiple mutations are generally needed for full-fledged cancer

– The longer we live, the more mutations we accumulate

• This may help explain why the incidence of cancer

increases greatly with age

– At the DNA level, a cancerous cell is usually characterized

by at least one active oncogene and the mutation of

several tumor-suppressor genes

• The model of a multistep path to cancer is well-supported by studies of colorectal

cancer

– Like most cancers, colorectal cancer develops gradually

• The 1st sign is often a polyp, made up of cells that look normal but divide

unusually frequently

• The tumor grows and may eventually become malignant, spreading to

other tissues

– The development of this malignant tumor is caused by a gradual accumulation

of mutations that convert proto-oncogenes to oncogenes and knock out tumor-

suppressor genes

• A ras oncogene and a mutated p53 tumor-suppressor gene are often

involved

Fig. 18-22

EFFECTS OF MUTATIONS

Malignant tumor

(carcinoma)

Colon

Colon wall

Loss of tumor-

suppressor gene

APC (or other)

Activation of

ras oncogene

Loss of

tumor-suppressor

gene DCC

Loss of

tumor-suppressor

gene p53

Additional

mutations

Larger benign

growth (adenoma)

Small benign

growth (polyp)

Normal colon

epithelial cells

5

42

3

1

Inherited Predisposition and Other Factors Contributing to Cancer

• Individuals can inherit oncogenes or mutant alleles of

tumor-suppressor genes

– Inherited mutations in the tumor-suppressor gene

adenomatous polyposis coli are common in

individuals with colorectal cancer

– Mutations in the BRCA1 or BRCA2 gene are found

in at least half of inherited breast cancers

You should now be able to:

1. Explain the concept of an operon and the

function of the operator, repressor, and

corepressor

2. Explain the adaptive advantage of grouping

bacterial genes into an operon

3. Explain how repressible and inducible operons

differ and how those differences reflect

differences in the pathways they control

4. Explain how DNA methylation and histone

acetylation affect chromatin structure and the

regulation of transcription

5. Define control elements and explain how they

influence transcription

6. Explain the role of promoters, enhancers,

activators, and repressors in transcription

control

7. Explain how eukaryotic genes can be

coordinately expressed

8. Describe the roles played by small RNAs on

gene expression

9. Explain why determination precedes

differentiation

10. Describe two sources of information that

instruct a cell to express genes at the

appropriate time

11. Explain how maternal effect genes affect polarity and development in Drosophila embryos

12. Explain how mutations in tumor-suppressor genes can contribute to cancer

13. Describe the effects of mutations to the p53 and ras genes