welcome to mb class 2 molecular biology of the gene, 5/e --- watson et al. (2004) part i: chemistry...

Welcome to

MB Class

Welcome to

MB Class

2

Molecular Biology of the Gene, 5/E --- Watson et al. (2004)

Part I: Chemistry and Genetics

Part II: Maintenance of the Genome

Part III: Expression of the Genome

Part IV: Regulation

Part V: Methods

2005-5-10

The revised central dogma

RNA processingRNA processing

Gene regulationGene regulation

基因组的保持

基因组的表达

The structure of DNA and RNAThe structure of DNA and RNA

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Part IV RegulationCh 16: Transcriptional regulation in Ch 16: Transcriptional regulation in prokaryotesprokaryotes

Ch 17: Transcriptional regulation in Ch 17: Transcriptional regulation in eukaryoteseukaryotes

Ch18: Regulatory RNAsCh18: Regulatory RNAs

Ch 19: Gene regulation in Ch 19: Gene regulation in development and evolutiondevelopment and evolution

Ch 20: Genome Analysis and Ch 20: Genome Analysis and Systems BiologySystems Biology

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Housekeeping genes: expressed constitutively, essential for basic processes involving in cell replication and growth.

Inducible genes: expressed only when they are activated by inducers or cellular factors.

Expression of many genes in cells are regulated

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Surfing the contents of Part IV

--The heart of the frontier biological

disciplines

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Some of the peoples who significantly contribute to

the knowledge of gene regulation

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Chapter 16Gene Regulation

in Prokaryotes

Chapter 16Gene Regulation

in Prokaryotes

•Molecular Biology Course

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TOPIC 1 Principles of Transcriptional Regulation [watch the animation]

TOPIC 2 Regulation of Transcription Initiation: Examples from Bacteria (Lac operon, alternative factors, NtrC,MerR, Gal rep, araBAD operon)

TOPIC 3 The Case of Phage λ: Layers of Regulation

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Topic 1: Principles Topic 1: Principles of Transcription of Transcription

Regulation Regulation

Topic 1: Principles Topic 1: Principles of Transcription of Transcription

Regulation Regulation

CHAPTER 16 Gene Regulation in Prokaryotes

1.1. What are the regulatory proteins?What are the regulatory proteins?2.2. Which steps of gene expression to be Which steps of gene expression to be

targeted?targeted?3.3. How to regulate? (recruitment, allostery, How to regulate? (recruitment, allostery,

blocking, action at a distance, cooperative blocking, action at a distance, cooperative binding)binding)

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1. Gene Expression is Controlled by Regulatory Proteins ( 调控蛋白 )

Gene expression is very often controlled by Extracellular Signals, which are communicated to genes by regulatory proteins:

Positive regulators or activators INCREASE the transcription

Negative regulators or repressors

DECREASE or ELIMINATE the transcription

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2. Most activators and repressors act at the level of transcription initiation

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Why that?Why that?1.1. Transcription initiation is the Transcription initiation is the

most energetically efficient most energetically efficient step to regulate. [A wise step to regulate. [A wise decision at the beginning]decision at the beginning]

2.2. Regulation at this step is Regulation at this step is easier to do well than easier to do well than regulation of the translation regulation of the translation initiation.initiation.

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Regulation also occurs at all Regulation also occurs at all stages after transcription stages after transcription initiation. Why?initiation. Why?

1.1. Allows more inputs and Allows more inputs and multiple checkpoints.multiple checkpoints.

2.2. The regulation at later stages The regulation at later stages allow a quicker response.allow a quicker response.

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Fig 12-3-initiation

Promoter Binding (closed complex)Promoter Binding (closed complex)

Promoter “melting” (open complex)

Promoter escape/Initial transcription

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Fig 12-3-Elongation and termination

Termination

Elongation

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3. Targeting promoter binding:

Many promoters are regulated by activators ( 激活蛋白 ) that help RNAP bind DNA (recruitmentrecruitment) and by repressors ( 阻遏蛋白 ) that blockblock the binding.

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Generally, RNAP binds many promoters weakly. Why?

Activators contain two binding sites to bind a DNA sequence and RNAP simultaneously, can therefore enhance the RNAP affinity with the promoters and increases gene transcription. This is called recruitment regulation ( 招募调控 ).***

On the contrary, Repressors can bind to the operator inside of the promoter region, which prevents RNAP binding and the transcription of the target gene.

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a. Absence of Regulatory Proteins: basal level expression

b. Repressor binding to the operator repressesexpressionc. Activator binding activates expression

Fig 16-1

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4 Targeting transition to the open complex: Allostery regulation ( 异构调控 ) after the RNA Polymerase Binding

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In some cases, RNAP binds the promoters efficiently, but no spontaneous isomerization ( 异构化 ) occurs to lead to the open complex, resulting in no or low transcription.

Some activators can bind to the closed complex, inducing conformational change in either RNAP or DNA promoter, which converts the closed complex to open complex and thus promotes the transcription. This is an example of allostery regulation.

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Fig 16-2

Allostery regulation

Allostery is not only a mechanism of gene activation , it is also often the way that regulators are controlled by their specific signals.

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Repressors Repressors can work in ways:

(1)blocking the promoter binding.

(2)blocking the transition to the open complex.

(3)blocking promoter escape

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5. Action at a Distance and DNA Looping. The regulator proteins can function even binding at a DNA site far away from the promoter region, through protein-protein interaction and DNA looping.

Fig 16-3

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Fig 16-4 DNA-binding protein can facilitate interaction between DNA-binding proteins at a distance

Fig 16-4

Architectural protein

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6. Cooperative binding (recruitment) and allostery have many roles in gene regulation

For example: group of regulators often bind DNA cooperatively (activators and/or repressors interact with each other and with the DNA, helping each other to bind near a gene they regulated) :

(1) produce sensitive switches to rapidly turn on a gene expression. (1+1>2)

(2) integrate signals (some genes are activated when multiple signals are present).

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Watch the animation-regulation of the transcription initiation!

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Topic 2: Regulation Topic 2: Regulation of Transcription of Transcription

Initiation : Initiation : Examples Examples

from Bacteriafrom Bacteria

Topic 2: Regulation Topic 2: Regulation of Transcription of Transcription

Initiation : Initiation : Examples Examples

from Bacteriafrom Bacteria


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OperonOperon:: a unit of prokarytoic gene expression and regulation which typically includes: 1. Structural genes for enzymes in a specific biosynthetic and metabolic pathway whose expression is coordinately controlled. 2. Control elements, such as operator sequence. 3. Regulator gene(s) whose products recognize the control elements. These genes is usually transcribed from a different promoter.

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Control element

Structural genes

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First example: Lac First example: Lac operonoperon

First example: Lac First example: Lac operonoperon

Regulation of Transcription Regulation of Transcription Initiation in Bacteria Initiation in Bacteria

The lactose Operon The lactose Operon (( 乳糖操纵子乳糖操纵子 ))

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Point 1: Composition of Point 1: Composition of the the LacLac operon operon

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The enzymes encoded by lacZ, lacY, lacA are required for the use of lactose as a carbon source. These genes are only transcribed at a high level when lactose is available as the sole carbon source.

Fig 16-5

The LAC operon

1. Lactose operon contains 3 structural genes and 2 control elements.

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lacY encodes a cell membrane protein called lactose permease ( 半乳糖苷渗透酶 ) to transport Lactose across the cell wall

lacZ codes for β-galactosidase ( 半乳糖苷酶 ) for lactose hydrolysis

lacA encodes a thiogalactoside transacetylase ( 硫代半乳糖苷转乙酰酶 )to get rid of the toxic thiogalacosides

The LAC operon

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The lacZ, lacY, lacA genes are transcribed into a single lacZYA mRNA (polycistronic mRNApolycistronic mRNA) under the control of a single promoter Plac .

LacZYA transcription unit contains an operator site Olac

position between bases -5 and +21 at the 3’-end of Plac

Binds with the lac repressor

The LAC operon

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Point 2: Regulatory Point 2: Regulatory proteins and their proteins and their

response to extracellular response to extracellular signalssignals

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2. An activator and a repressor together control the Lac operon expression

The activator: CAP (Catabolite Activator Protein, 代谢产物激活蛋白 ) or CRP (cAMP Receptor Protein,cAMP 受体蛋白 ); responses to the glucose level.The repressor: lac repressor that is encoded by LacI gene; responses to the lactose.

Sugar switch-off mechanismThe LAC operon

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3. The activity of Lac repressor and CAP are controlled allosterically by their signals.

Lactose is converted to allolactose by -galactosidase, therefore lactose can indirectly turn off the repressor. Glucose lowers the cellular cAMP level, therefore, glucose indirectly turn off CAP.

The LAC operon

Allolactose binding: turn of Lac repressor

cAMP binding: turn on CAP

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Fig 16-6

The LAC operon

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ii pp oo zz yy aa

Very low level of lac mRNAVery low level of lac mRNA

Absence of lactoseAbsence of lactose

ActiveActive

i p o z y a

-Galactosidase

PermeaseTransacetylase

Presence of lactose

Inactive

Lack of inducer: the lac repressor block all but a very low level of trans-cription of lacZYA .

When Lactose is present, the low basal level of permease allows its uptake, and -galactosidase catalyzes the conversion of some lactose to allolactose.

Allolactose acts as an inducer, binding to the lac repressor and inactivate it.

Response to lactose

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Response to glucose: 注意该图的 CRP 结合位点有误

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Point 3: The mechanism Point 3: The mechanism of the binding of of the binding of

regulatory proteins to regulatory proteins to their sitestheir sites

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The LAC operon

Repressor binding physically Repressor binding physically prevents RNAP from binding prevents RNAP from binding to the promoterto the promoter, because the site bound by lac repressor is called the lac operator (Olac ), and the Olac overlaps promoter (Plac).

4. CAP and Lac repressor have opposing effects on RNA polymerase binding to the promoter

The LAC operon

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The LAC operon

CAP binds to a site upstream of CAP binds to a site upstream of the promoter, and helps RNA the promoter, and helps RNA polymerase binds to the polymerase binds to the promoter by physically promoter by physically interacting with RNAP. interacting with RNAP. This cooperative binding stabilizes the binding of polymerase to Plac.

The LAC operon

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Fig 16-8

The LAC operon

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The LAC operon

5. CAP interacts with the CTD domain of the a-subunit of RNAP

The LAC operon

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CAP site has the similar structure as the operator, which is 60 bp upstream of the start site of transcription.

CAP interacts with the CTD domain of the -subunit of RNAP and thus promotes the promoter binding by RNAP.

Fig 16-9

CTD: C-terminal domain of the subunit of RNAP

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CAP binds as a dimer

CTD

Fig 16-10. CAP has separate activating and DNA-binding surface

The LAC operon

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6. CAP and Lac repressor bind DNA using a common structural motif: helix-turn-helix motif

The LAC operon

Fig 16-11

One is the recognition helix that can fits into the major groove of the DNA. Another one sits across the major grove and makes contact with the DNA backbone.

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DNA binding by a helix-turn-helix motif

Fig 16-12 Hydrogen Bonds between repressor and the major groove of the operator.

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Lac operon contains three operators: the primary operator and two other operators located 400 bp downstream and 90 bp upstream.

Lac repressor binds as a tetramer ( 四聚体 ), with each operator is contacted by a repressor dimer ( 二聚体 ). respectively.

Fig 16-13

DNA looping

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A regulator (CAP) works together with different repressors at different genes, this is an example of Combinatorial Control.

In fact, CAP acts at more than 100 genes in E.coli, working with an array of partners.

7: Combinatorial Control ( 组合调控 ): CAP controls other genes as well.

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Second example: Second example: Alternative Alternative factor factor

Second example: Second example: Alternative Alternative factor factor


Alternative factors ( 可变因子 ) direct RNA polymerase to alternative promoters.

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factor subunit bound to RNA polymerase for transcription

initiation (Ch 12)

Fig 12-7 and subunits recruit RNA pol core enzyme to the promoter

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Different factors binding to the same RNAP, conferring each of them a new promoter specificity.

70 factors is the most common one in E. coli under the normal growth condition.

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Many bacteria produce alternative sets of σfactors to meet the regulation requirements of transcription under normal and extreme growth condition. Bacteriophage has its own σfactors

E. coli: Heat shock 32

Sporulation in Bacillus subtilis

Bacteriophage σ factors

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Heat shock ( 热休克 ) Around 17 proteins are specifically

expressed in E. coli when the temperature is increased above 37ºC.

These proteins are expressed through transcription by RNA polymerase using an alternative factor 32 coded by rhoH gene. 32 has its own specific promoter consensus sequences.

Alternative Alternative factors factors

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Many bacteriophages synthesizetheir own σfactors to endow thehost RNA polymerase with a different promoter specificity and hence to selectively express their own phage genes .

Bacteriophages

Alternative Alternative factors factors

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B. subtilis SPO1 phage expresses a cascade of σfactors which allow a defined sequence of expression of different phage genes.

Fig 16-14Alternative Alternative factors factors

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Third example: NtrC Third example: NtrC and MerR use and MerR use

allosteric activationallosteric activation

Third example: NtrC Third example: NtrC and MerR use and MerR use

allosteric activationallosteric activation


Transcriptional activators NtrC and MerRwork by allostery rather than by recruitment.

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Review The majority of activators work by

recruitment, such as CAP. These activators simply bring an active form of RNA polymerase to the promoter.

The beautiful exceptions: allosteric activation by NtrC and MerR.

In allosteric activation RNAP initially binds the promoter in an inactive complex, and the activator triggers an allosteric change in that complex to activate transcription.

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1. NtrC has ATPase activity and works at DNA sites far away from the gene.

NtrC and MerR and allosteric NtrC and MerR and allosteric activationactivation

NtrC controls expression of genes involved in nitrogen metabolism ( 氮代谢 ), such as the glnA gene.

NtrC has separate activating and DNA-binding domains, and binds DNA only when the nitrogen levels are low.

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Low nitrogen levels ( 低水平氮 )NtrC phosphorylation and conformational change NtrC (?) binds DNA sites at ~-150 bp position as a dimer (?)NtrC interacts 54 in RNAP bound to the glnA promoter NtrC ATPase activity provides energy needed to induce a conformation change in RNAP transcription STARTs

Fig 16-15 activation by NtrC

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2. MerR activates transcription by twisting promoter DNA

NtrC and MerR and allosteric NtrC and MerR and allosteric activationactivation

MerR controls a gene called merT, which encodes an enzyme that makes cells resistant to the toxic effects of mercury ( 抗汞酶 )

In the presence of mercury ( 汞 ), MerR binds to a sequence between –10 and –35 regions of the merT promoter and activates merT expression.

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As a 70 promoter, merT contains 19 bp between –10 and –35 elements (the typical length is 15-17 bp), leaving these two elements recognized by 70 neither optimally separated nor aligned.

66Fig 16-15 Structure of a merT-like promoter

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Fig 16-15

When Hg2+ is absent, MerR binds to the promoter and locks it in the unfavorable conformationWhen Hg2+ is present, MerR binds Hg2+ and undergoes conformational change, which twists the promoter to restore it to the structure close to a strong 70

promoter

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Repressors work in many ways-reviewBlocking RNA polymerase binding through binding to a site overlapping the promoter. Lac repressorBlocking the transition from the closed to open complex. Repressors bind to sites beside a promoter, interact with polymerase bound at that promoter and inhibit initiation. E.coli Gal repressorBlocking the promoter escape. P4 protein interaction with PA2c (bacteriophage 29 )

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Fourth example: Fourth example: araaraBADBAD operon operon

Fourth example: Fourth example: araaraBADBAD operon operon


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1. AraC and control of the araBAD operon by anti-activation

The araBAD The araBAD operonoperon

The promoter of the araBAD operon from E. coli is activated in the presence of arabinose ( 阿拉伯糖 ) and the absence of glucose and directs expression of genes encoding enzymes required for arabinose metabolism. This is very similar to the Lac operon.

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Different from the Lac operon, two activators AraC and CAP work together to activate the araBAD operon expression

Fig 16-18

CAP site

194 bp

DNA looping

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Because the magnitude of induction of the araBAD promoter by arabinose is very large, the promoter is often used in expression vector.

If fusing a gene to the araBAD promoter, the expression of the gene can be easily controlled by addition of arabinose （阿拉伯糖） .

What is an expression vector ? [The answer is in the Methods part.]

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Topic 3: The Case of Topic 3: The Case of Bacteriophage Bacteriophage : :

Layers of RegulationLayers of Regulation

Topic 3: The Case of Topic 3: The Case of Bacteriophage Bacteriophage : :

Layers of RegulationLayers of Regulation


Bacteriophage Bacteriophage is a virus that infects is a virus that infects E. coliE. coli. . Upon infection, the phage can Upon infection, the phage can propagatepropagate in in either of two ways : lytically or lysogenically.either of two ways : lytically or lysogenically.

TheThe phage phage has a 50-kb genome and has a 50-kb genome and ~50 genes. Most of these genes encode ~50 genes. Most of these genes encode protein for replication, packing or lysis. protein for replication, packing or lysis.

How the lytic and lysogenic How the lytic and lysogenic growth is regulated? growth is regulated? ---regulatory ---regulatory proteins and cis-acting control proteins and cis-acting control elements.elements.

1.1. Alternative patterns of Alternative patterns of gene expression control gene expression control lytic and lysogenic lytic and lysogenic growth. growth.

Fig. 16-21: Promoters in the right and Fig. 16-21: Promoters in the right and left left control regions control regions of phage of phage

Fig. 16-22: Transcription in the Fig. 16-22: Transcription in the control regions in lytic and control regions in lytic and lysogenic growthlysogenic growth

2. Regulatory Proteins and 2. Regulatory Proteins and Their Binding SitesTheir Binding Sites

The cI gene encodes The cI gene encodes repressor, repressor, that can both activate and repress that can both activate and repress transcriptiontranscription

As a repressor: As a repressor: similarly as Lac similarly as Lac repressor (?)repressor (?)

As an As an activator: activator: similarly as similarly as CAP (?) CAP (?)

repressorrepressorAs a repressor, it binds to sites that As a repressor, it binds to sites that

overlap the promoter and excludes RNA overlap the promoter and excludes RNA

polymerasepolymerase

As an activator, it works like CAP by As an activator, it works like CAP by

recruitment.recruitment.

Cro (another regulatory protein), stands for control of repressor and other things. It is a single domain protein that binds as a dimer to 17-bp DNA sequences using a HTH motif. It only represses transcription.

repressor and Cro can each bind to any one of six operators, but with dramatically different affinity. repressor binds OR1 most easily while Cro binds OR3 with highest affinity.repressor binds OR1 tenfold better than OR2. Cro binds OR3 tenfold better than OR1 and OR2.

There are 6 operators in the right (3) and There are 6 operators in the right (3) and left (3) control regions of bacteriophage left (3) control regions of bacteriophage

Sequences are not identical

3. 3. repressor binds to operator repressor binds to operator sites cooperatively. sites cooperatively. Two dimmers Two dimmers of repressor bind cooperatively to Oof repressor bind cooperatively to OR1R1 and Oand OR2R2, The binding at O, The binding at OR1R1 helps the helps the binding at Obinding at OR2R2..

MonomerMonomer

DimerDimer

TetramerTetramer

High affinityHigh affinity10-fold 10-fold low low affinityaffinity

Not boundNot bound

Box 16-3 Concentration, affinity, and Box 16-3 Concentration, affinity, and cooperative bindingcooperative binding

Two factors determine whether two interacting molecules find and bind each others: (1) the binding affinity (2) their concentrations.

Cooperativity binding can be Cooperativity binding can be expressed in terms of increased expressed in terms of increased affinity. affinity.

The curve of l The curve of l repressor repressor binding to its binding to its operator operator DNA.DNA.

Binding of Binding of a protein a protein to a single to a single sitesite

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The benefit of cooperative binding The benefit of cooperative binding of regulatory proteins of regulatory proteins is to is to ensure dramatic changes in the ensure dramatic changes in the expression level of a given gene expression level of a given gene even in response to small even in response to small changes in the level of the changes in the level of the control signal.control signal.

Lytic growth

Lysogen

4. Repressor and Cro bind in 4. Repressor and Cro bind in different patterns to control lytic different patterns to control lytic and lysogenic growthand lysogenic growth

1. DNA damage activates RecA in E. coli 2. RecA stimulates repressor to undergo

autocleavage, resulting in the removal the C-terminal domain and the immediate loss of dimerization and binding cooperativity.

3. Repressor dissociates from OR1-OR2 & OR1-OR2, which triggers transcription from PR and PL

4. leading to lytic growth.

5. Lysogenic induction requires 5. Lysogenic induction requires proteolytic cleavage of proteolytic cleavage of repressorrepressor

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For induction to work efficiently, the level For induction to work efficiently, the level of repressor in a lysogen must be tightly of repressor in a lysogen must be tightly regulated. regulated. How?How?

Keep it not too Keep it not too lowlow by by positive positive autoregulation: autoregulation: repressor binding at repressor binding at OOR2R2 activates its own transcription from activates its own transcription from PPRMRM. . Keep it nKeep it not too ot too highhigh by by negative negative autoregulation: when the repressor level autoregulation: when the repressor level goes too high, it will bind to goes too high, it will bind to OOR3 R3 as well, as well, which will prevents which will prevents transcription from transcription from PPRMRM..

When repressor level is high, it occupies both OR1-OR2 and OL1-OL2, and the interaction between two tetramer forms the repressor octomer and bring together OR3 and OL3 for another cooperative binding of the repressor.

That’s why lysogeny can be so stable That’s why lysogeny can be so stable while also ensuring that induction is while also ensuring that induction is very efficient. very efficient.

6. Negative autoregulation of 6. Negative autoregulation of repressor requires long-distance repressor requires long-distance interactions and a large DNA interactions and a large DNA loop: cooperative binding at loop: cooperative binding at OOR3 R3

and and OOL3.L3.

Fig. 16-27Fig. 16-27

Figure 16-28 Interactions between the c-terminal domain of repressors.

cII is transcribed from cII is transcribed from PPRR and cIII is and cIII is transcribed from transcribed from PPLL. .

CII protein is a transcriptional activator that CII protein is a transcriptional activator that binds to PRE and stimulate the binds to PRE and stimulate the transcription of cI gene (transcription of cI gene ( repressor). repressor).

7. Another activator, 7. Another activator, CII, controls CII, controls the decision between lytic and the decision between lytic and lysogenic growth upon infection of lysogenic growth upon infection of a new host-an earlier event.a new host-an earlier event.

Establishment of lysogeny: Establishment of lysogeny: synthesis of the essential synthesis of the essential lysogenic lysogenic repressor repressor is is establishedestablished by transcription by transcription from one promoter and then from one promoter and then maintainedmaintained by by transcription from another one.transcription from another one.

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Establishment of lysogeny-a bigger Establishment of lysogeny-a bigger viewview1.1.PPRR and and PPLL is constitutive promoters is constitutive promoters that promote transcription once the that promote transcription once the phage enter the cells. phage enter the cells. 2.2.PPRR directs the synthesis of both Cro directs the synthesis of both Cro and CII proteins. Cro favors lytic and CII proteins. Cro favors lytic development while CII favors development while CII favors lysogentic growth by activating the lysogentic growth by activating the synthesis of synthesis of repressor. repressor.3.3.The efficiency with which CII directs The efficiency with which CII directs transcription of cI gene (transcription of cI gene ( repressor) repressor) is critical in deciding the lysogeny.is critical in deciding the lysogeny.

??? What determines CII efficiency???? What determines CII efficiency?

8. The number of phage particles 8. The number of phage particles infecting a given cell affects whether infecting a given cell affects whether the infection proceeds lytically or the infection proceeds lytically or lysogenically.lysogenically.When more CII proteins are made from more When more CII proteins are made from more infected phages, there is a larger chance to infected phages, there is a larger chance to produce enough produce enough repressor to produce lysogeny. repressor to produce lysogeny.

9. Growth conditions of 9. Growth conditions of E. coli E. coli control control the stability of CII protein and thus the stability of CII protein and thus thethelytic/lysogenic choice.lytic/lysogenic choice. Infection of Healthy and growing vigorously Infection of Healthy and growing vigorously bacterial cells >>CII is unstable >>propagates bacterial cells >>CII is unstable >>propagates lytically because.lytically because.When conditions are poor for bacterial growth When conditions are poor for bacterial growth >>CII becomes stable >> form lysogens and sit >>CII becomes stable >> form lysogens and sit tight.tight.[CII is degraded by a specific protease FtsH][CII is degraded by a specific protease FtsH]

10. Transcriptional Antitermination 10. Transcriptional Antitermination in in development: examples of development: examples of regulation after transcription regulation after transcription initiation. initiation.

Two Two phage regulatory proteins N phage regulatory proteins N and Q, called antiterminators, and Q, called antiterminators, prevent the termination at some prevent the termination at some termination sites and promotes the termination sites and promotes the transcription of the early late and transcription of the early late and late genes for the lytic growth of late genes for the lytic growth of the phage. the phage.

Gene expression in Gene expression in lytic growthlytic growth

Three phases: Immediate early:

Transcription starts at PR and PL that flank the cI and stops at the -dependent terminators (t) after the N and cro genes;

Delayed early: Transcription begins at the same promoters, but bypasses the terminators by virtue of the N gene product, N, which is an antiterminator;

Late: Transcription begins at a new promoter P’R ; it would stop short at the t without the Q gene product, Q, another antiterminator.

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N proteins binds to the RNA

Q proteins bind to the QBE DNA site.

Antitermination by N proteinAntitermination by N protein

The gene The gene surrounding N surrounding N are depicted are depicted along with the along with the leftward leftward promoter (Ppromoter (PLL) and ) and operator (Ooperator (OLL), the ), the terminator and terminator and the the nut nut site.site.

Transcription in Transcription in the absence of N the absence of N

Transcription in Transcription in the presence of Nthe presence of N

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1.Principles of gene regulation. (1) who regulate? (2) where to target? (3) How to regulate?

2.Regulation of transcription initiation in bacteria: the lac operon, alternative factors, NtrC, MerR, araBAD operon.

3.The case of phage--layers of regulation: repressor and Cro and their binding; control of the lytic and lysogenic growth; lysogenic induction; control of the decision to lytic or lysogenic growth by CII; Antiterminators.

Key points of the chapter

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