L17 Virulence: Mechanisms of Gene Regulation II

Question 1

What is cholera?

Answer 1

Life-threatening diarrheal disease

Question 2

What is the causative agent of cholera?

Answer 2

Vibrio cholerae

Question 3

What are the physical characteristics of V. cholerae?

Answer 3

1. Highly motile
2. Uni-flagellated
3. Gram-negative
4. Curved rod
5. Extracellular pathogen

Question 4

What must occur for V. cholerae to cause disease?

Answer 4

1. Be ingested via contaminated food or water
2. Survive passage through the gastric acid barrier of the stomach
3. Colonize the upper SI
4. Produce and excrete toxin
5. Disseminate in a watery diarrhea

Question 5

What is the main virulence factor of cholera?

Answer 5

Cholera toxin (mutants lacking toxin genes are much less virulent)

Question 6

What are the two subunits of the cholera toxin?

Answer 6

A and B

Question 7

The ctxA ctxB operan is carried by the ___.

Answer 7

Lysogenic CTX prophage

Question 8

Describe how the 1 A subunit exerts its effects.

Answer 8

1. A is cleaved to yield an active A1 subunit.
2. A1 catalyzes ADP-ribosylation of the regulatory G protein, G-alpha-s
3. G-alpha-s activates adenylyl cyclase
4. Adenylyl cyclase increases cAMP levels
5. cAMP activates PKA
6. PKA phosphorylates transporters, causing efflux of ions and water
7. Unregulated ion transport causes water to leak into the intestinal lumen, leading to severe, watery diarrhea

Question 9

What are the 5 B subunits required for?

Answer 9

Secretion of A1 toxin out of the bacterial cell and interaction with the host cell surface receptor, GM1 glycoprotein

Question 10

Describe how a single polar flagellum functions as a virulence factor in colonization.

Answer 10

Motility is used to reach the site of colonization. Flagellar genes are expressed when toxin genes are not; motility genes are turned off at the site of colonization and toxigenic genes are turned on. Note that non-motile and hyper-motile strains are less virulent.

Question 11

What are the virulence factors required for adherence to host epithelial cells?

Answer 11

1. TCP (toxin co-regulated pilus)

2. ACF (accessory colonization factors)

Question 12

What happens when there are mutations in the TCP or ACF genes?

Answer 12

Colonization decreases

Question 13

Where are the TCP and ACF genes located?

Answer 13

On a large pathogenicity island termed the TCP-ACF element

Question 14

What is ToxR?

Answer 14

Protein that acts as the global virulence regulator

Question 15

Activation of the ToxR regulon permits what?

Answer 15

Synthesis of virulence factors

Question 16

What is ToxS?

Answer 16

Protein that acts as the environmental sensor

Question 17

ToxR and ToxS form an operon. How is transcription of these genes regulated?

Answer 17

Via temperature

Question 18

What happens to the toxRS operon at low temperatures?

Answer 18

At low temperatures (outside the host), the operon is on. ToxR and ToxS are synthesized and incorporated into the membrane.

Question 19

What happens to the toxRS operon at high temperatures (in the upper GI tract)?

Answer 19

ToxRS is off; the proteins are not synthesized, but the old proteins are still present in the membrane

Question 20

What happens to the toxRS operon at high temperatures (intestine)?

Answer 20

ToxS receives a signal unique to the intestine environment. It communicates this information to ToxR. ToxRS activates ToxT synthesis.

Question 21

What does ToxT do?

Answer 21

ToxT autoregulates and increases transcription of itself. ToxT also activates transcription of the TCP-ACF pathogenicity island. Finally, ToxT regulates transcription of the ctxA ctxB operon carried by the lysogenic CTX prophage, leading to the Ctx toxin.

Question 22

What are the components of the ToxR regulon?

Answer 22

1. Stimulus: unknown (lumenal)
2. Sensor: ToxS
3. Regulator: ToxR (also ToxT, technically)
4. Member genes: toxT, tcp, acf, ctxab