Lecture 5 - Gene Regulation in Prokaryotes Pt.1

Why must genes be regulated?

To turn genes ON when needed and OFF when no longer needed.

At what levels can gene expression be regulated?

DNA (transcriptional), RNA (post-transcriptional), and protein (post-translational).

What are regulatory elements?

DNA sequences near a gene that regulate expression but are not transcribed.

What is positive control of gene expression?

Regulation that increases transcription.

What is negative control of gene expression?

Regulation that decreases transcription.

What are DNA-binding proteins?

Proteins that bind DNA to regulate gene expression.

What is a DNA-binding domain?

The part of a protein that binds DNA via backbone interaction or base hydrogen bonding.

Why is DNA-binding protein regulation reversible?

Proteins can bind and unbind dynamically.

What is Helix Loop Helix?

Protein structure that has a secondary shape that is well recognized as DNA-binding protein.

What is zinc finger and leucine zippers?

DNA-recognition element - recognizes a specific sequence on the DNA and binds (controls positive or negative)

What impacts the expression of genes?

The Binding impacts gene expression.

What is an operon?

A cluster of genes controlled by a single promoter and operator.

Only _____ have operons.

Prokaryotes

What is the regulator gene?

A gene that encodes a regulatory protein; usually not physically part of the operon.

What is the promoter?

DNA sequence where RNA polymerase binds.

What is the operator?

DNA sequence where regulatory proteins bind.

What is the structural gene?

Everything after the regulator gene, they generate enzymes that are needed by the cell

What is in front of the structural gene?

the operator

What does the operator bind to?

Regulatory protein

What can the regulatory protein be?

An activator or repressor.

What creates the regulatory protein?

The regulator gene

What is next to the operator?

The promoter (promoter binds polyermase)

What regulatory protein is used in negative control?

Repressor.

What does a repressor do?

Inhibits transcription.

What regulatory protein is used in positive control?

Activator.

What does an activator do?

Stimulates transcription.

What is an inducible operon?

Normally OFF (due to active repressor); must be turned ON by a signal.

What is a repressible operon?

Normally ON (inactive repressor); must be turned OFF by a signal.

If there is an activator on the operator and it leaves what happens?

No transcription

If there is a repressor on the operator and it leaves what happens?

Transcription occurs.

What are cis-acting factors?

DNA elements that affect only the DNA molecule they are on.

Example of a cis-acting factor

Promoter or operator sequence.

What happens if one gene copy has a mutated promoter?

Only that copy is affected; the other still works.

What is a main difference between cis and trans operating factors?

• Cis stays in one spot, trans can float around

• Cis is made up of DNA elements, trans is made up of protein

What are trans-acting factors?

Diffusible molecules (usually proteins) that regulate genes on different DNA molecules.

Examples of trans-acting factors

Activators and repressors - they are moving around looking to bind.

What are the structural genes in lac operon?

lacZ, lacY, lacA.

What does lacZ encode?

β-galactosidase (breaks lactose into glucose + galactose)

What does lacY encode?

Permease (imports lactose into the cell)

What does lacA encode?

Transacetylase (function unknown).

What regulatory elements are part of the lac operon?

Promoter and operator (cis-acting).

What does LacI encode?

The lac repressor protein.

Is lacI cis- or trans-acting?

Trans-acting.

Is lacI gene constitutive (making the regulator)?

Yes — it is always making the regulator protein.

What happens in the absence of lactose?

Repressor binds operator → no transcription.

What happens when lactose is present?

Lactose → allolactose → inactivates repressor → transcription occurs.

What is LacP?

Lactose promoter

What is LacO?

Lactose Operator

What is allolactose?

An inducer formed from lactose.

What does allolactose do?

Binds the repressor and prevents it from binding the operator.

Who discovered the lac operon mechanism?

Jacob and Monod.

What is a partial diploid (merozygote)?

A cell with both chromosomal and plasmid copies of lac genes.

How are genotypes written in partial diploids?

Chromosome / plasmid (e.g., I⁺ P⁺ O⁺ Z⁺ Y^- (copy1 ) / I⁻ P⁺ O⁺ Z⁺ Y^- (copy 2))

How is the merozygote made?

A secondary lac operon is inserted to create a merozygote.

What is lacI⁻?

Nonfunctional repressor → constitutive expression.

Is lacI⁻ recessive or dominant?

Recessive.

What is Lacl+?

Funtionally normally. 1 copy of normal Lacl was sufficient to restore function because its trans acting element.

What is lacIˢ (superrepressor)?

Repressor that cannot bind allolactose.

What is the effect of lacIˢ?

Always represses transcription, even with lactose present.

Is lacIˢ dominant or recessive?

Dominant.

What is lacOᶜ?

Operator mutation that cannot bind repressor.

What is the effect of lacOᶜ?

Constitutive transcription. (can’t stop transcribing B-gal)

Is lacOᶜ cis- or trans-acting?

Cis-acting.

What is lacO^-?

Operator has a mutation that prevents it from binding the repressor.

What is the difference between lacO^c and lacO^-?

They function the same but have different sequences.

What is lacP⁻?

Promoter mutation preventing RNA polymerase binding.

What is the effect of lacP⁻?

No transcription, regardless of lactose.

What is lacZ⁻?

β-galactosidase is nonfunctional.

What is lacY⁻?

Permease is nonfunctional.

Do lacZ⁻ or lacY⁻ affect regulation?

No — they affect protein function, not transcription.