Transcriptional Regulation (W8)

Why do bacteria regulate gene expression?

To conserve energy and materials by producing proteins only when they are useful under current conditions.

What is the difference between constitutive and regulated gene expression?

• Constitutive genes are expressed continually for essential functions

• Regulated genes change expression in response to conditions

What is an operon?

• A cluster of nearby genes that work together and are controlled together by regulatory proteins

• The genes are usually copied into one mRNA and often make proteins (products) that are used in the same pathway

What are the main parts of a bacterial operon?

• A promoter

• An operator or activator-binding site

• Structural/functional genes

• A separate regulatory gene often makes the repressor or activator protein that controls the operon.

What is the promoter?

A DNA sequence where RNA polymerase binds to begin transcription.

What is the role of RNA polymerase in operon expression?

It binds the promoter and synthesizes RNA from the DNA template.

• Regulators can block or help this process

What are structural (functional) genes?

Genes that encode proteins that carrying out the cellular function or pathway. The operon controls whether these genes are expressed

What is a regulatory gene?

A gene that encodes a regulatory protein, such as a repressor or activator, which controls transcription of another gene or operon.

What is an operator?

A regulatory DNA site, usually near or overlapping a promoter, where a repressor can bind and block transcription.

What is an activator-binding site?

A regulatory DNA site, often upstream of the promoter, where an activator binds to help RNA polymerase initiate transcription.

Compare negative and positive transcriptional control.

• Negative control uses a repressor to reduce or prevent transcription

• Positive control uses an activator to increase transcription

How does a repressor regulate an operon?

It binds an operator and prevents RNA polymerase from binding effectively or moving through the DNA to transcribe the genes.

How does an activator regulate an operon?

It binds an activator-binding site and helps recruit or stabilize RNA polymerase at the promoter.

What is an effector molecule in transcriptional regulation?

A small molecule that binds a regulatory protein and changes its shape and DNA-binding activity.

What is a repressible operon?

An operon usually on that is turned off when its end product is abundant

• These commonly control biosynthetic pathways

• Example: tryptophan synthesis

How does the trp operon respond when tryptophan is abundant?

• Tryptophan acts as a corepressor by binding the trp repressor

• This now-active repressor binds to the operator and blocks transcription

How does the trp operon respond when tryptophan is scarce?

• The trp repressor is inactive and cannot bind the operator

• So, RNA polymerase continues to transcribe genes needed to synthesize tryptophan

Why is the trp operon repressible?

Tryptophan synthesis is costly and is needed only when environmental tryptophan is unavailable, so having an abundant amount of tryptophan / product shuts off its own biosynthetic genes.

What is a corepressor?

A small molecule, such as tryptophan, that binds a repressor and enables it to bind DNA and inhibit transcription.

What is an inducible operon?

An operon normally off/low that is turned on when an inducer (often the pathway substrate) is present

• These commonly control catabolic pathways

• Example: lac operon

How is the lac operon regulated when lactose is absent?

The LacI repressor binds the operator and blocks RNA polymerase, subsequently blocking transcription of genes for lactose use.

How is the lac operon regulated when lactose is present?

• Allolactose, derived from lactose, binds the lac repressor and changes its shape

• lac repressor then releases from the operator, allowing transcription to proceed at a slow rate

Why is the lac operon inducible?

Lactose-related molecules induce expression only when lactose is available to be used as a nutrient.

What happens when arabinose is present, and why is this a positively regulated operon?

• araC binds arabinose and becomes an activator

• Helps RNA polymerase turn on the araBAD genes for arabinose metabolism, so the operon is positively regulated

How does AraC regulate the araBAD operon when arabinose is absent?

• AraC binds sites that create a DNA loop and prevent effective RNA polymerase action

• It acts as an anti-activator, functionally like a repressor

A cell makes a toxinase enzyme that’s designed only to break down a (pretend) toxic molecule called toxininium. How should toxinase expression respond to toxininium?

It should be inducible: toxininium should inactivate or remove a repressor so toxinase is produced only when the toxin is present.

What is quorum sensing?

Cell-cell communication in which microbes use signaling molecules to coordinate gene expression according to population density.

What is an autoinducer?

• A small quorum-sensing signal molecule produced and released by cells

• It can diffuse or be transported between cells and accumulates as the population grows

  • More autoinducer makes cells secrete more autoinducer and transcribe quorum sensing regulated genes

What happens to autoinducer concentration at low versus high cell density?

• At low density, autoinducer is diluted or diffuses away

• At high density, production by many cells causes it to accumulate to a threshold concentration

Why is quorum sensing useful for costly group behaviors?

• It prevents individual cells from wasting energy on a behavior that is ineffective at low population density

• Activates it only when enough cells can produce a useful effect

What is an example of costly behavior that quorum sensing is helpful for?

Light production (i.e. bioluminescence)

• Uses many ATP, NADPH, and numerous steps

• Solution? → Only make light when there are enough cells to count

How does the Vibrio fischeri lux system work?

• LuxI synthesizes an autoinducer

• At high cell density, the autoinducer binds LuxR

• Activated LuxR promotes transcription of lux genes, including luminescence genes and more luxI.

Why does Vibrio fischeri produce light mainly at high cell density in its squid host?

• Bioluminescence is energetically expensive and too weak to benefit the host when few bacteria are present

• Quorum sensing coordinates light production after enough cells accumulate

What are common outcomes controlled by quorum sensing?

• Biofilm and extracellular polymeric substance production

• Secreted enzymes or factors

• Virulence-related behaviors

• Symbiosis functions

• Developmental changes

In what situation is quorum sensing especially likely to be used?

• When a secreted product or cooperative behavior works only after many cells contribute enough of it

• Examples: biofilm formation, host colonization, or extracellular enzyme production.