T6-PROKARYOTIC CONTROL OF GENE EXPRESSION

What is gene expression

Overall process by which the information encoded in a gene is converted into an observable phenotype (most commonly production of a protein)

Why regulate gene expression?

1. To adjust to sudden changes

2. To conserve energy

3. To save resources

Process of gene expression

DNA to mRNA to Protein

Why is gene expression regulation important?

To ensure the right proteins are produced at the right time and amount based on cellular needs.

Why doesn’t a cell produce all proteins at once?

Because different proteins are needed in different amounts or only under certain conditions.

How does gene regulation help during stress or changing conditions?

It allows rapid production of specific proteins needed to respond to environmental changes.

How does gene regulation affect energy use in the cell?

It conserves energy by only producing necessary proteins.

What are 6 types of RNA

mRNA, rRNA, tRNA, snRNA, snoRNA, other noncoding RNA

What is the function of mRNA

messenger RNAs, code for proteins

What is the function of rRNA

ribosomal RNAs, form the basic structure of the ribosome and catalyze protein synthesis

What is the function of tRNA

transfer RNAs, central to protein synthesis as adaptors between mRNA and amino acids

What is the function of snRNA

small nuclear RNAs, function in a variety of nuclear processes, including the splicing of pre-mRNA

What is the function of snoRNA

small nucleolar RNAs, used to process and chemically modify rRNAs

What is the function of other noncoding RNAs

function in diverse cellular processes, including telomere synthesis, X-chromosome inactivation, and the transport of proteins into the ER

Why is gene expression regulated in cells?

Because cells have the same DNA, but only certain genes are expressed all the time while others are switched on or off as needed.

What is an operon?

A group of genes in prokaryotes that are transcribed together.

Where are operons found?

Only in prokaryotes.

What happens when RNA polymerase binds an operon promoter?

All genes in the operon are transcribed together.

What is the function of operons?

They control important biochemical processes in prokaryotes.

Who discovered operons?

Jacob, Monod, and Lwoff through studies in E. coli.

What is the lac operon?

A set of genes involved in lactose processing in bacteria.

What does the lac operon control?

The breakdown and utilization of lactose.

What are the four scenarios of the production of β-galactosidase?

1. When glucose is present and lactose is absent the E. coli does not produce β--galactosidase.

2. When glucose is present and lactose is present the E. coli does not produce β--galactosidase

3. When glucose is absent and lactose is absent the E. coli does not produce β--galactosidase.

4. When glucose is absent and lactose is present the E. coli does produce β--galactosidase

What is the function of β-galactosidase?

It hydrolyzes lactose into glucose and galactose.

What does lactose get broken down into?

Glucose and galactose.

When is β-galactosidase NOT produced in E. coli?

When glucose is present (regardless of lactose).

When is β-galactosidase produced in E. coli?

When glucose is absent and lactose is present.

What is the preferred energy source for E. coli?

Glucose.

Why is β-galactosidase not produced when both glucose and lactose are absent?

Because the operon cannot be switched on.

Why is β-galactosidase produced when glucose is absent but lactose is present?

Lactose is used as an alternative sugar source.

What happens to the lac operon when glucose is absent but lactose is present?

1. The repressor is removed,

2. allowing RNA polymerase to bind and

3. β-galactosidase to be produced so

4. lactose can be metabolized.

What is the function of a repressor protein?

regulatory gene that codes for a protein that inhibits expression of target genes by binding to an operator or silencer to inhibit gene expression and turn genes off.

What is the role of a promoter and how does a suppressor gene affect it?

A promoter is a DNA sequence where RNA polymerase binds to start transcription; a suppressor gene can inactivate or block the promoter, preventing transcription.

What happens to the lac operon when lactose is absent vs present?

1. When lactose is absent, a repressor protein binds the operator and blocks RNA polymerase from the promoter, preventing transcription of lac genes.

2. When lactose is present, the lacI gene produces a repressor protein that prevents RNA polymerase binding, so genes like β-galactosidase are not expressed.

What happens to the lac operon when lactose is present?

Allolactose binds the repressor, causing a shape change that prevents it from binding the operator, allowing transcription.

What is allolactose?

A lactose-derived molecule that acts as the natural inducer of the lac operon.

What is the role of allolactose in the lac operon?

It inactivates the repressor, allowing gene expression for lactose metabolism.

What happens when the lac operon is OFF (no lactose present)?

The repressor binds the operator and blocks RNA polymerase, stopping transcription of lacZ, lacY, and lacA.

What is the effect of allolactose binding to the repressor?

It causes a conformational change that prevents the repressor from binding the operator.

What genes are blocked when the lac operon is OFF?

lacZ, lacY, and lacA.

What happens to the lac operon when both glucose and lactose are present?

The operon is OFF or only minimally expressed due to catabolite repression.

What is catabolite repression?

A regulatory mechanism where glucose prevents strong transcription of the lac operon even if lactose is present.

Why is the lac operon weakly expressed when both sugars are present?

RNA polymerase binds weakly and is unstable, causing poor transcription.

What happens to RNA polymerase binding when glucose and lactose are both present?

It can bind the promoter but is unstable and frequently falls off.

What role does allolactose play when both glucose and lactose are present?

It inactivates the repressor, but transcription is still limited due to glucose effect.

Why does lactose alone not guarantee lac operon activation?

Because glucose presence triggers catabolite repression, reducing transcription efficiency.

What happens to the lac operon when glucose is absent and lactose is present?

The operon is fully active with high transcription of lac genes.

What role does the activator protein (CAP) play in the lac operon?

It stabilizes RNA polymerase binding to increase transcription.

When is the activator protein (CAP) active?

When glucose is absent.

What is the effect of CAP–cAMP on the lac operon?

It enhances RNA polymerase binding, leading to strong transcription.

Why is lac operon expression highest when glucose is absent and lactose is present?

Because the repressor is removed and CAP–cAMP activates transcription.

Describe the

• activator protein,

• repressor protein,

• rna polymerase and

• lac operon of

+GLUCOSE +LACTOSE

1. not bound to DNA,

2. lifted off operator site,

3. keeps falling of promoter site,

4. no transcription

Describe the

• activator protein,

• repressor protein,

• rna polymerase and

• lac operon of

+GLUCOSE -LACTOSE

1. not bound to DNA,

2. bound to operator site,

3. blocked by the repressor,

4. no transcription

Describe the

• activator protein,

• repressor protein,

• rna polymerase and

• lac operon of

-GLUCOSE -LACTOSE

1. bound to DNA

2. bound to operator site,

3. blocked by the repressor,

4. no transcription

Describe the

• activator protein,

• repressor protein,

• rna polymerase and

• lac operon of

-GLUCOSE +LACTOSE

1. bound to DNA,

2. lifted of operator site,

3. sits on promoter site,

4. transcription

Describe concept 1: gene regulation in bacteria

• Bacteria adapt to environmental changes by using regulatory proteins to turn genes on or off,

• tightly controlling gene expression at the transcriptional level

• so only needed genes are expressed depending on conditions.

Describe concept 2: The Lactose Operon (Jacob-Monod model)

• The lactose (lac) operon is a group of bacterial genes controlled by a single promoter,

• where the regulatory gene (lacI) produces a repressor protein that regulates transcription by

  • controlling RNA polymerase binding at the promoter site.

Describe Concept 3: The lac Operator

• The lac operator is a short DNA sequence near or overlapping the promoter that binds the lac repressor protein (lacI).

• When the repressor is bound, it blocks RNA polymerase and stops transcription (OFF state).

• When lactose is present, allolactose binds the repressor causing it to detach, allowing RNA polymerase to transcribe lacZ, lacY, and lacA,

  • producing a polycistronic mRNA that enables lactose metabolism.

What is the lac operon an example of?

An inducible system.

What does “inducible system” mean in the lac operon?

It is normally OFF and turned ON only when lactose (allolactose) is present.

What does lacZ encode?

β-galactosidase, which breaks lactose into glucose and galactose.

What does lacY encode?

Lactose permease, which transports lactose into the cell.

What does lacA encode?

Thiogalactoside transacetylase, which transfers acetyl groups to β-galactosides.

Explain the Jacob and Monod model of transcriptional regulation of the lac operon by lac repressor.

1. Lac repressor binds operator (O) → blocks RNA polymerase, transcription OFF

2. Lactose binds repressor → conformational change, repressor detaches from operator

3. RNA polymerase binds promoter (P) → transcription ON, producing polycistronic mRNA for lacZ, lacY, lacA

What happens in lac operon repression (no lactose)?

Repressor binds operator, blocks RNA polymerase, no transcription (operon OFF).

What triggers induction of the lac operon?

Lactose (allolactose) binding to the repressor.

What is the effect of allolactose on the repressor?

It causes a conformational change and releases the repressor from the operator.

What happens when the repressor is removed?

RNA polymerase binds the promoter and transcription begins.

What type of mRNA is produced from the lac operon?

A polycistronic mRNA containing lacZ, lacY, and lacA.

What proteins are produced from the lac operon mRNA?

β-galactosidase, lactose permease, and thiogalactoside transacetylase.

Describe Concept 4: The lac Regulatory Gene

• The lac regulatory gene (lacI) is located upstream of the lac operon and is

  • continuously expressed at a low level to produce a lac repressor protein.

• This repressor binds the operator and blocks RNA polymerase, preventing transcription of the structural genes.

• Although separate from the operon, lacI controls it and allows rapid response to lactose availability.

Describe Concept 5: The Lac Repressor Protein

• The lac repressor protein (produced by lacI) is active in the absence of lactose and binds to the operator, blocking RNA polymerase from transcribing lacZ, lacY, and lacA.

• This prevents unnecessary gene expression and conserves energy in E. coli.

Describe Concept 6: The Effect of Lactose on the lac Operon

• When lactose is present, it is converted to allolactose, which binds to the lac repressor at an allosteric site and causes a conformational change.

• This prevents the repressor from binding the operator, allowing RNA polymerase to bind the promoter and transcribe the lac operon, producing lacZ, lacY, and lacA.

Describe Concept 7: The lac Inducer: Allolactose

• Allolactose is the inducer of the lac operon, formed from lactose inside the cell.

• It binds to the lac repressor at an allosteric site, causing a conformational change that prevents the repressor from binding the operator.

• This allows RNA polymerase to transcribe the lac genes.

• In the absence of allolactose, the repressor remains bound and the operon stays OFF to conserve energy.

Describe Concept 8: Feedback Control of the lac Operon

• When lac operon enzymes break down lactose and allolactose, the inducer is removed and the repressor becomes active again, stopping transcription.

• lac mRNA also degrades quickly, so protein production stops.

• This negative feedback mechanism shuts the operon OFF once lactose is depleted, preventing waste of energy and resources.

Describe Concept 9: Energy Source Preferences of E. coli

• E. coli prefers glucose as its main energy source, so when glucose is present it is metabolized first and lac genes are only weakly expressed due to low cAMP and inactive CAP.

• When glucose is depleted, cAMP increases, CAP–cAMP binds DNA, stabilizes RNA polymerase, and strongly activates transcription of lac genes so lactose can be used efficiently.

What energy source does E. coli prefer?

Glucose

What happens to lac operon expression when glucose is present?

It is weakly expressed or inhibited.

Why is the lac operon weakly active when glucose is present?

Low cAMP prevents CAP activation, so RNA polymerase is not strongly recruited.

What happens when glucose is depleted?

cAMP increases and CAP–cAMP activates transcription of lac genes.

What is the role of CAP–cAMP?

It binds DNA and stabilizes RNA polymerase to strongly activate transcription.

Why does E. coli use glucose before lactose?

Because glucose is the most efficient and preferred energy source.

Describe Concept 10: The Effect of Glucose and Lactose on the lac Operon

1. When both glucose and lactose are present, the lac operon is weakly transcribed because RNA polymerase binds inefficiently without CAP–cAMP activation.

2. Maximum transcription occurs only when lactose is present (removes repression) and glucose is absent (high cAMP forms CAP–cAMP), allowing strong activation of the lac operon through both positive control and relief of negative control.

Describe Concept 11: The Effect of Glucose and Cyclic AMP on the lac Operon

1. Glucose and cAMP are inversely related in E. coli.

2. High glucose causes low cAMP, keeping CAP inactive and reducing lac operon transcription.

3. When glucose is low, cAMP increases, binds CAP to form the CAP–cAMP complex, which activates transcription by stabilizing RNA polymerase at the promoter, ensuring the lac operon is expressed only when glucose is absent.

How does glucose affect cAMP levels in E. coli?

High glucose lowers cAMP, while low glucose increases cAMP.

What is the role of cAMP in the lac operon?

It binds CAP to form CAP–cAMP, which activates transcription.

What happens to the lac operon when glucose is high?

cAMP is low, CAP is inactive, and transcription is low.

What happens to the lac operon when glucose is low?

cAMP increases, CAP–cAMP forms, and transcription is activated.

What is the function of the CAP–cAMP complex?

It binds near the promoter and stabilizes RNA polymerase to increase transcription.

Why is cAMP considered an energy signal?

It reflects glucose availability and regulates whether lactose is used for energy.

Describe Concept 12: The Effect of Lactose in the Absence of Glucose on the lac Operon

1. When lactose is present and glucose is absent, cAMP levels increase and bind to CAP (catabolite activator protein/CRP).

2. The CAP–cAMP complex binds near the lac promoter and enhances RNA polymerase binding, leading to strong transcription of the lac operon.

3. This provides positive control that fine-tunes gene expression when glucose is unavailable.

What happens to the lac operon when lactose is present and glucose is absent?

The lac operon is highly activated with strong transcription of lac genes.

What happens to cAMP when glucose is absent?

cAMP levels increase in the cell.

What does cAMP bind to in the lac operon system?

It binds to CAP (catabolite activator protein/CRP).

What is the function of the CAP–cAMP complex?

It binds near the lac promoter and enhances RNA polymerase binding, increasing transcription.

Why is transcription of the lac operon high when glucose is absent?

Because CAP–cAMP activates RNA polymerase while lactose removes repression.

What type of regulation does CAP–cAMP represent?

Positive control of gene expression.

Summarize lac operon

1. No inducer → repressor binds operator → operon OFF

2. Regulatory gene (lacI) produces repressor; promoter/operator are binding sites

3. Inducer (allolactose) inactivates repressor → transcription ON

What is the inducer of the lac operon?

Lactose in the form of allolactose

What is the role of allolactose in the lac operon?

It acts as an inducer that triggers transcription of lac genes.