Lecture #15: Bacterial Gene Regulation

Question 1

Gene transfer and recombination
Genetic material transferred between bacteria via 3 mechanisms:

Answer 1

• Transformation
• Transduction
• Conjugation

Question 2

Transformation

Answer 2

• taking up of dead material

Question 3

Transduction

Answer 3

• genetic material is transferred via a virus

Question 4

Conjugation

Answer 4

• bacteria is actively trying to transfer DNA to another bacteria (transferring of genes)

Question 5

Genetic recombination + Homologous recombination

Answer 5

• cutting DNA molecules and putting them back together (insertion, etc.)
• Homologous recombination: recombination between sequences that are similar.

Question 6

Components of Conjugation

Answer 6

• Conjugation is formed when the bacteria makes a bridge to another bacteria in order to transfer DNA.
• Two bacterias in every situation (donor/recipient cell)
• F factor: plasmid carried by bacteria that enables it to be a donor in conjugation.
• what does the plasmid have that allows it to be a donor? Genes on the plasmid code for pilus and specialized DNA handling machinery.
• becomes an F+ cell from an F- cell

Question 7

Conjugation: F+ to F- cells

Answer 7

• (2) special kind of DNA replication
• The pilus connects the two cells in the diagram

Question 8

Conjugation: Hfr to F-

Answer 8

• High frequency recombination
• Trasnferring of bacterial chromosome through functional versions of a,b,c, and d genes.
• The F plasmid gets incorporated into the bacterial chromosome, so it becomes an Hfr cell. The groups become nonfunctional
  -At some, the pilus is formed, and the bacterial chromosome (Hfr) can transfer the functional b group into the new bacteria.

Question 9

Bacterial Gene Regulation

Answer 9

• Gene expression regulation equals transcriptional regulation
  → regulation of gene expression is important to control when genes are transcribed (therefore the protein product made), in relation to environmental or cellular conditions. (Ex. make enzymes for metabolizing certain substrates (lactose) when it is present, or when the product you want is absent (tryptophan).)
  → genes can be turned on (activated), or turned off (repressed)

Question 10

Promoter

Answer 10

• DNA sequence required for RNA polymerase recruitment

Question 11

Bacterial transcriptional unit organization

Answer 11

• In prokaryotes the RNA polymerase binds to the promoter directly
• We also need the transcriptional stop site
• Before the promoter there are regulatory sequences and regulatory factors: work together with protein factors to regulate specific gene expression.

Question 12

Operon

Answer 12

• functional unit of DNA containing a cluster of genes under the control of a single regulatory system. Genes in an operon are transcribed as one unit
• Regulatory factor is produced by a gene that is outside.

Question 13

Lactose Metabolism

Answer 13

• The enzyme that breaks down the disaccharide (galactose) is LacZ-B-galatosidase (only want to make it when lactose is around)
• lacY is a permease is involved in letting lactose into the cell
• Not all bacteria genes are in operons (like the regulatory gene) (key)

Question 14

Inducible Lac Operon + Lactose absent

Answer 14

Use a brake and accelerator analogy to look at two levels of regulation
- Lac repressor is the brake system, binds to the operator

Question 15

Lactose present

Answer 15

• RNA polymerase can bind to the promoter because the allolactose changes the shape of the repressor
• B-galactosidase- converts lactose to glucose and galactose
  –>But also converts lactose to allolactose at low rate.
  –>Senses allolactose (proxy for lactose levels) to know that we have to turn off the repressor.

Question 16

2nd level of regulation at the lac operon

Answer 16

• System can also be modulated by the presence of other sugars
• If Glucose is present IT’S MORE EFFICIENT TO USE GLUCOSE, so if it’s present we want to use it and not lactose.
• It will do this through a catabolite activator protein (CAP): binds to a CAP site 5’ of the promoter in the presence of cAMP (adenosine monophosphate that has been cyclized) + increases transcription
• Increased glucose leads to low cAMP levels

Question 17

lactose is present

Answer 17

• cAMP is bound to the CAP site

Question 18

glucose is present, and lactose is present

Answer 18

• Because cAMP is not there the CAP cannot bind to the cap site (thus, taking the foot off the accelerator)

Question 19

Lactose absent, glucose absent

Answer 19

-Regulatory activity: lacI is repressing, CAP activating
- LacZ production: +/- (a little bit of lac is made)

Question 20

Lac absent, glu present:

Answer 20

• Regulatory activity:: lacI is repressing, CAP not activating
• LacZ production: +/-

Question 21

Lac pres, glu absent:

Answer 21

• Regulatory activity: LacI not repressing, CAP activating
• LacZ production: ++/++

Question 22

Lac pres, glu, pres:

Answer 22

• Regulatory activity: LacI is not repressing, CAP not activating
• LacZ:+

Question 23

Horizontal gene transfer

Answer 23

• E.Coli goes from friendly to pathogenic through transfer + regulation of a gene sequence called a pathogenicity island
• The genes confer the ability to be pathogenic on the E.Coli. The regulation of these genes is somewhat like what we see in lac.