Lecture 7 - Virulence Gene Regulation

Question 1

Why regulate virulence genes?

Answer 1

It is very rare for pathogens to move directly from a specific host environment to another host environment.
Must be able to adapt to an intermediate environment.

Question 2

Phase variation

Answer 2

When a gene is either turned on or off

Question 3

Example of phase variation

Answer 3

Opa protein in Neisseria Gonorrhoeae

(CTCTT)n repeat between start of gene and variable regions of gene

DEpending on number of (CTCTT) repeats, different V regions of gene are in frame or out of frame

This allows the bacterium to express different adhesins

Question 4

Example of regulation at level of DNA

Answer 4

Slipped-strand misrepair during DNA replication

Question 5

Example of regulation at the level of transcription

Answer 5

Repressor protein binding to operator region

Question 6

Example of regulation at the level of translation

Answer 6

Operon encoding several genes on one mRNA

Question 7

Basic idea behind transcriptional regulation

Answer 7

Regulating function of RNA polymerase enzyme

Question 8

Which part of gene does RNA polymerase bind to?

Answer 8

Promoter

Question 9

Operon

Answer 9

Several genes encoded in a single mRNA transcript

Question 10

Regulon

Answer 10

Several promotors controlled by the same regulator

Question 11

Example of regulon

Answer 11

Listeria monocytogenes Prf regulation

Question 12

Listeria monocytogenes Prf regulation
1)
2)

Answer 12

1) prfA binds to prfA gene regulator, upregulates prfA expression
2) prfA also binds to many other promoters, one of which encodes a virulence gene

Question 13

Lac operon role

Answer 13

Expression of b-galactosidase under low glucose

Question 14

Gene encoding b-galactosidase

Answer 14

LacZ

Question 15

Lac operon structure (upstream to downstream)
1)
2)
3)
4)
5)
6)
7)

Answer 15

1) LacI gene encodes repressor protein
2) cAMP activator protein binding site
3) Promoter region
4) Operator region
5) LacZ
6) LacY
7) LacA

Question 16

Lac operon function
1)
2)
3)

Answer 16

1) When lactose is absent, repressor protein binds operator and prevents RNA polymerase function
2) When glucose is present cAMP activator protein leaves cAMP activator protein binding site and RNA polymerase functions at a far lower rate
3) When lactose is present and glucose is absent repressor protein is absent and cAMP activator protein is present. Translation of operon occurs

Question 17

DtxR role

Answer 17

Dimerises with Fe2+ to inhibit diphtheria toxin expression

Question 18

Bacterium causing diphtheria

Answer 18

Corynybacterium diphtheriae

Question 19

What causes expression of diphtheria toxin?

Answer 19

Low Fe2+

Question 20

Where does RNA polymerase stop transcribing in bacteria?

Answer 20

When it reaches a transcription terminator

NOT when it reaches a stop codon

Question 21

How does DtxR prevent toxin expression?

Answer 21

Dimerises with Fe2+, binds to operator region of tox gene

Question 22

Which bacteria are two component systems common in?

Answer 22

Gram negative

Question 23

Basic structure of a two component system

1)
2)
3)

Answer 23

1) Sensor (S) protein is in the periplasm/external environment, attached to a regulator (R) protein in the cytoplasm
2) Activation of S results in dimerisation, cross-phosphorylation of R.
3) R phosphorylates a second protein in the cytoplasm which dimerises, activates/represses a gene

Question 24

Example of a two component system
1)
2)
3)
4)

Answer 24

1) Sensor is activated
2) Histidine on R is cross phosphorylated
3) Phosphoryl transfer to aspartic acid on response regulator
4) Response regulator dimerises, activates gene

Question 25

Bordatella pertussis bvg regulation
1)
2)
3)

Answer 25

1) S region detects stimulus
2) On R region, histidine, then aspartic acid, then histidine is phosphorylated
3) bvgA protein is phosphorylated, activates genes

Question 26

Ways to regulate transcription
1)
2)
3)
4)

Answer 26

1) Two component systems
2) Different sigma factors
3) Repressor proteins
4) DNA topography

Question 27

A way that bacteria can alter DNA topography

Answer 27

Histone-like proteins

EG: H-NS

Question 28

What does H-NS stand for?

Answer 28

Histone-like nucleoid-structuring protein

Question 29

Function of H-NS
1)
2)
3)

Answer 29

1) H-NS binding sites exist in genes
2) Two bound copies of H-NS bind together
3) RNA polymerase is prevented from transcription

Question 30

Basic idea of quorum sensing

Answer 30

Bacteria release autoinducers

When autoinducer levels become high enough, autoinducers reenter bacteria and stimulate response

Question 31

Can unrelated bacteria participate in quorum sensing?

Answer 31

Yes

Question 32

Common autoinducer in Gram negative bacteria

Answer 32

Acyl homoserine lactone

AI-1

Question 33

Why are autoinducers different for Gram positive and negative bacteria?

Answer 33

Gram negative autoinducers need to be more lipophilic as they need to pass through two membranes

Question 34

AI-1 mode of action
1)
2)
3)
4)
5)

Answer 34

1) LuxI (AI synthase) makes AI-1
2) AI-1 released from bacterium
3) When [AI-1] is high enough, reenters cell
4) Binds LuxR response regulator
5) LuxR/AI-1 complex binds to gene

Question 35

Vibrio fischeri quorum sensing in squid
1)
2)
3)

Answer 35

1) Baby squid secretes gel which attracts bacteria
2) When V. fischeri have high enough numbers in squid, quorum sense and express luciferase gene
3) Bacteria emit light which squid uses to hunt

Question 36

Autoinducers used by Vibrio fischeri

Answer 36

AI-1 and AI-2

Question 37

Squid colonised by Vibrio fischeri

Answer 37

Bobtail squid

Question 38

Which bacterium is prf present in?

Answer 38

Listeria monocytogenes

Question 39

Which bacterium is opa protein present in?

Answer 39

Neisseria gonorrhoeae

Question 40

WHich bacterium is bvg present in?

Answer 40

Bordatella pertussis

Question 41

Which bacterium is DtxR present in?

Answer 41

Corynybacterium diphtheriae