Control of Gene Expression

Steps of gene expressionSteps of gene expression

TranscriptioTranscription – n – DNA is DNA is read to make read to make a mRNA in a mRNA in the nucleus of the nucleus of our cellsour cells

Translation Translation – – Reading the Reading the mRNA to mRNA to make a make a protein in the protein in the cytoplasmcytoplasm

Mainly controlled at the level of transcription

Prokaryotic and eukaryotic gene organization

Prokaryotic transcriptional

regulatory regions

(promoters and operators) lie close to the

transcription start siteFunctionally

related genes are frequently

located near each other These “operons”

are transcribed into a single mRNA with

internal translation

initiation sites

Prokaryotic Gene Prokaryotic Gene ExpressionExpression

PromoterCistron1Cistron2CistronNTerminator

Transcription RNA Polymerase

mRNA 5’ 3’

TranslationRibosome, tRNAs,Protein Factors

1 2 N

Polypeptides

NC

NC N

C

1 2 3

Expression mainly by controlling transcription

OperonsOperons A cluster of related genes often coding for enzymes in a metabolic pathway, which are under the control of a single

promoter regulatory region Genes that work together are located together

A promoter plus a set of adjacent genes whose gene products function together.

They are controlled as a unit They usually contain 2 –6 genes (up to 20 genes)

These genes are transcribed as a polycistronic transcript. It is relatively common in prokaryotes

It is rare in eukaryotes

Operon SystemOperon System

Structural genes : DNA that code for a specific polypeptide (protein)

Promoter : DNA segment that recognizes RNA polymerase

Operator : Element that serves as a binding site for an inhibitor protein (modulator) that controls transcription

Repressor : Protein which binds to a specific DNA sequences to determine the transcription of a particular geneRegulatory gene : Gene encode for repressor protein

Regulatory elements of transcription

Regulatory gene:Organization of operon

Operons• The Tryptophan Operon (Repressible and

attenuation) Repressor does not bind to operator unless it interacts with co repressorBiosynthetic pathways

• The Lactose Operon (Induction and catabolite

repression)Repressor is bound to operator unless molecule to be metabolized is present (inducer)Catabolic pathways

A repressible operon

Inducible Operon

Lactose Operon• It codes for the enzymes responsible for lactose catabolism• Within the operon, there are three genes that code for

proteins (structural protein) and an upstream control region including promoter and a regulatory site called the operator

• Laying outside the operon is the repressor gene, which codes for a protein (lac repressor) that binds to the operator site and is responsible for the suppression of the operon by blocking the binding of RNA polymerase

• Transcribed mRNA may contain information for more than one protein (a polycistronic mRNA)

• The synthesis of these mRNA is regulated in accordance with the needs of the cells at any time thus enable the cell to adapt quickly to changing environmental conditions

The lactose (lac) The lactose (lac) operonoperon

• Contains several elementsContains several elements– laclacZ gene = Z gene = ββ-galactosidase-galactosidase– laclacY gene = galactosidase permeaseY gene = galactosidase permease– laclacA gene = thiogalactoside transacetylaseA gene = thiogalactoside transacetylase– laclacI gene = I gene = lac lac repressorrepressor

– PPii = promoter for the = promoter for the laclacI geneI gene– P = promoter for P = promoter for laclac-operon-operon– QQ11 = main operator = main operator– QQ22 and Q and Q33 = secondary operator sites (pseudo- = secondary operator sites (pseudo-

operatorsoperators))

Pi P Z Y A I Q3 Q1 Q2

Regulation of the lac operonRegulation of the lac operon

Pi P Z Y A I Q3 Q1 Q2

Inducer molecules→ Allolactose: - natural inducer, degradable IPTG (Isopropylthiogalactoside)- synthetic inducer, not metabolized

lacI repressor

Pi P Z Y A I Q3 Q1 Q2

LacZ LacY LacA

The lac operon: model for gene expression

Includes three protein synthesis coding region--sometimes called "genes" as well as region of chromosome that controls transcription of genes Genes for proteins involved in the catabolism or breakdown of lactose When lactose is absent, no transcription of gene since no need for these proteinsWhen lactose is present, transcription of genes takes place so proteins are available to catalyze breakdown of lactose

Eukaryotic geneEukaryotic gene

Eukaryotic gene ExpressionEukaryotic gene Expression

1.Transcripts begin and end beyond the coding region

2.The primary transcript is processed by:5’ capping3’ formation / polyA

splicing

3.Mature transcripts are transported to the cytoplasm for translation

Control of Gene Expression

Regulation of gene expression Gene expression is regulated—not all genes Gene expression is regulated—not all genes

are constantly active and having their protein are constantly active and having their protein producedproduced

The regulation or feedback on gene The regulation or feedback on gene expression is how the cell’s metabolism is expression is how the cell’s metabolism is controlled. controlled.

This regulation can happen in different ways:This regulation can happen in different ways:1. Transcriptional control (in nucleus):1. Transcriptional control (in nucleus):

e.g. chromatin density and transcription factorse.g. chromatin density and transcription factors

2. Posttranscriptional control (nucleus)2. Posttranscriptional control (nucleus)e.g. mRNA processinge.g. mRNA processing

3. Translational control (cytoplasm)3. Translational control (cytoplasm)e.g. Differential ability of mRNA to bind ribosomese.g. Differential ability of mRNA to bind ribosomes

4. Posttranslational control (cytoplasm)4. Posttranslational control (cytoplasm)e.g. changes to the protein to make it functionale.g. changes to the protein to make it functional

– 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

Enhancers

Otherproteins

DNA

Transcriptionfactors

Activatorproteins

RNA polymerase

Promoter Gene

Bendingof DNA

Transcription

Transcription control• Transcription factors• Proximal activators• Distal control elements

(enhancers)– DNA binding domain– Activation domains

bind to other proteins– These are cell-specific– A few common

structures, but found in different combinations in different cells

Eukaryotic gene expression

Gene regulation of the transcription

Chr. I

Chr. II

Chr. III

Condition 1

“turned on”

“turned off”

Condition 2

“turned off”

“turned on”

1 2 3 4 5 6 7 8 9

10 11 12 13 14 15 16 17 18

19 20 21 22 23 24 25 26

constitutively expressed gene

induced gene

repressedgene

inducible/ repressible genes

Gene regulationGene regulation

constitutively expressed gene

1 2 3 4 5 6 7 8 9

10 11 12 13 14 15 16 17 18

19 20 21 22 23 24 25 26

Condition 3 Condition 4 upregulated gene expression

down regulated gene expression

Post-Transcriptional Modification in EukaryotesPost-Transcriptional Modification in Eukaryotes

Primary transcriptPrimary transcript formed firstformed first Then processed (3 steps) to form mature mRNA Then processed (3 steps) to form mature mRNA Then transported to cytoplasmThen transported to cytoplasm

Step 1: 7- methyl-guanosine “5’-cap” added to 5’ endStep 2: introns spliced out; exons link up

Step 3: Poly-A tail added to 3’ end

mature mRNA5’-cap- exons -3’ PolyA tail

Alternative picture: co-transcriptional pre-mRNA processing

Cap Functions

1. Protection of the mRNA from degradation (Protection from 5 exoribonucleases)

2. Enhances translation in the cytoplasm (Enhancement of the mRNA’s translatability)

3. Enhances transport from the nucleus4. Proper splicing of the pre-mRNA (Enhances

splicing of the first intron (for some pre-mRNAs))

The attachment of 7Me-GTP to the 5’ end of a nascent mRNA with a 5’ to 5’

phospho-ester linkage

Intron Splicing

•Exons : coding regions•Introns : noncoding regions

Step by step removal of pre-mRNA and joining of remaining exons

Removing intron from pre-mRNA

PolyadenylationThe process of adding poly(A) to RNA

Synthesis of the poly (A) tail involves cleavage of its 3’end and then the addition of about 40-200 adenine residues to form a poly (A) tail

Function - Poly(A) enhances both the lifetime and translatability of mRNA

End ProductEnd Product

The end products of protein synthesis is a The end products of protein synthesis is a primary structure of a proteinprimary structure of a protein..

A sequence of A sequence of amino acid amino acid bonded together bonded together by by peptide bondspeptide bonds..

aa1

aa2 aa3 aa4aa5

aa200

aa199

PolyribosomePolyribosome

incominglarge

subunit

incomingsmall subunit polypeptide

mRNA1 2 3 4 5 6 7

Groups of ribosomes reading same mRNA simultaneously producing many proteins

(polypeptides).

TYPES OF PROTEINSTYPES OF PROTEINS

Enzymes (Helicase)Enzymes (Helicase)Carrier (Haemoglobine)Carrier (Haemoglobine)

Immunoglobulin (Antibodies)Immunoglobulin (Antibodies)Hormones (Steroids)Hormones (Steroids)Structural (Muscle)Structural (Muscle)

Ionic (K+,Na+)Ionic (K+,Na+)

Coupled transcription and translation in bacteria

VALINE

HISTIDINE

LEUCINE

PROLINE THREONINE

GLUTAMATE

VALINE

original base triplet in a DNA strand

As DNA is replicated, proofreadingenzymes detect the mistake and

make a substitution for it:

a base substitution within the triplet (red)

One DNA molecule carries the original, unmutated sequence

The other DNAmolecule carries a gene mutation

POSSIBLE OUTCOMES:

OR

A summary of transcription and translation in a eukaryotic cellA summary of transcription and translation in a eukaryotic cell