L16 Virulence: Mechanisms of Gene Regulation I

Question 1

Before ingestion, intestinal pathogens often reside in water - describe the conditions of the water.

Answer 1

1. Low temperature
2. Low ionic strength
3. Low [organic nutrients]
4. Neutral pH

Question 2

After ingestion, intestinal pathogens must adapt to its new environment. Describe this environment.

Answer 2

1. Higher temperature
2. Higher osmotic strength
3. Low pH in the stomach
4. High pH, bile salts, lack of oxygen, abundant organic nutrients in SI
5. Sequestration of iron by the host

Question 3

To survive this journey, the pathogen must rapidly express a variety of gene products - what are 5 examples?

Answer 3

1. Proteins to tolerate the low stomach pH
2. Flagella and chemotaxis proteins for migration to a suitable niche
3. Adhesins that permit colonization
4. Toxins and invasins to elicit disease
5. Iron chelators (siderophores) to scavenge for ironW

Question 4

What is a virulence factor?

Answer 4

Any bacterial property required for entry, growth, or survival in a host

Question 5

What are 4 examples of virulence factors?

Answer 5

1. Capsule (inhibits complement killing)
2. Adhesins (adherence to host cells)
3. Acid tolerance factors (adapt to stomach)
4. Enzymes (synthesize unavailable nutrients)

Question 6

What is a virulence gene?

Answer 6

Any gene that encodes a virulence factor

Question 7

___% of genes in V. cholerae and S. typhimurium encode virulence factors.

Answer 7

5-10

Question 8

Where are virulence genes often located?

Answer 8

1. Mobile genetic elements (plasmids or phages)

2. Pathogenicity islands (large, localized regions of the chromosome)

Question 9

True or false - pathogenicity islands are often absent in closely related non-pathogenic strains

Answer 9

True

Question 10

Bacteria organize their genes in ___.

Answer 10

Multicistronic operons

Question 11

What is a cistron?

Answer 11

A sequence of DNA that encodes a polypeptide

Question 12

What is an operon?

Answer 12

A unit of transcription that includes more than one cistron

Question 13

What is multicistronic mRNA?

Answer 13

The mRNA that results from transcription of a multicistronic operon

Question 14

What are the 4 components of a typical operon?

Answer 14

1. Promoter
2. Operator
3. Cistrons
4. Terminator

Question 15

What is RNAP?

Answer 15

RNA polymerase; a biological machine that transcribes DNA into mRNA

Question 16

What is the promoter?

Answer 16

The site at which RNAp binds

Question 17

What is sigma?

Answer 17

A subunit of RNAP that specifically recognizes and binds the promoter

Question 18

What is the closed complex?

Answer 18

The product of the RNAP/DNA interaction

Question 19

What is a holoenzyme?

Answer 19

RNAP + sigma factor

Question 20

What are the steps of transcription initiation?

Answer 20

1. Formation of closed complex
2. Formation of open complex
3. Initiation and elongation

Question 21

Why do we need an open complex?

Answer 21

RNAP causes the double strand of DNA to open, creating the open complex. Strand separation allows for synthesis of the mRNA strand complementary to the DNA sense strand

Question 22

When the RNA polymerase is the only thing involved in transcription, this is known as ___.

Answer 22

Basal/constitutive transcription

Question 23

Describe the relationship between glucose and cAMP in E. Coli and organisms like it.

Answer 23

Inverse (as glucose decreases, cAMP increases and vice versa)

Question 24

Describe the process of repression, using lac as an example.

Answer 24

In this example, glucose is present and cAMP is low. Transcription of lacR is basally active, leading to the repressor protein. The repressor binds to the operator, which blocks RNAP from binding the promoter sequence.

Question 25

Cells regulate transcription primarily at its ___. They make the decision to do so by setting the ___ of initiation. This depends upon the ability of ___ to bind the promoter.

Answer 25

Initiation; frequency; RNAP

Question 26

What are the two types of regulator proteins that control the ability of RNAP to bind the promoter?

Answer 26

1. Repressor | 2. Inducer

Question 27

What does the repressor do?

Answer 27

Binds the operator and prohibits binding of RNAP to the promoter

Question 28

What does the inducer do?

Answer 28

Binds the repressor, changes the conformation so that it can no longer bind DNA

Question 29

What is an activator?

Answer 29

Interacts with RNAP, increasing the ability of RNAP to bind the promoter

Question 30

Describe the process of induction, using lac as an example.

Answer 30

In this example, glucose is absent and cAMP is high. Lactose is present. Lactose, the inducer, binds to the repressor, preventing it from binding to the DNA. RNA polymerase binds its activator, cAMP-CRP, which helps it bind to the promoter better. Transcription of lac occurs.

Question 31

RNAP binds to the ___.

Answer 31

Promoter

Question 32

The repressor binds to the ___.

Answer 32

Operator

Question 33

By binding the operator, the repressor reduces ___.

Answer 33

RNAP binding affinity

Question 34

When the inducer binds the repressor, what happens?

Answer 34

It decreases the repressor binding affinity for the operator.

Question 35

The activator interacts with RNAP and does what to its binding affinity?

Answer 35

Increases

Question 36

cAMP is a co-activator - what does it do?

Answer 36

Binds the activator, increasing the activator binding affinity

Question 37

The probability of transcription initiation increases as the stability of the closed complex ___.

Answer 37

Increases

Question 38

What increases stability of the closed complex? What decreases it?

Answer 38

Activators; repressors

Question 39

What is the core enzyme, and what is it comprised of?

Answer 39

RNA polymerase; 2 alpha, 1 beta, 1 beta prime subunits

Question 40

What is a holoenzyme?

Answer 40

RNA polymerase + sigma

Question 41

What does sigma do?

Answer 41

Recognizes the promoter

Question 42

What are the two domains of alpha and what do they do?

Answer 42

NTD and CTD; NTD helps assemble the core enzyme; CTD binds DNA involved in closed complex formation and interacts with sigma in the open complex formation

Question 43

When CRP is bound by cAMP, what happens?

Answer 43

CRP binds DNA

Question 44

What are the 5 interactions that enhance transcription initiation?

Answer 44

1. Sigma/DNA 2. DNA/alpha-CTD 3. Sigma/alpha-CTD 4. CRP/DNA 5. CRP/alpha-CTD

Question 45

What is a regulon?

Answer 45

A group of operons subject to the control of a common regulator

Question 46

Describe the general sequence of events in a regulon.

Answer 46

1. Cell senses a stimulus 2. Sensor signals to activate/deactivate a regulator 3. Regulator binds to several operons 4. Binding turns some on/some off 5. Gene products respond to original stimulus 6. Gene products exert feedback control on their own expression