Microbiology 19: (Tanaka) Decoding NF-KappaB Signal

Question 1

NF-KappaB general function? Examples of stimuli and responses?

Answer 1

Transcription factor - central mediator of immune and inflammatory responses

Stimuli:
Infection, Antigen receptors, cytokines, genotoxic stresses

Responses:

• Immune response/regulation (immunoregulatory proteins)
• NF-kB regulation (Rel, IkB proteins)
• Proliferation, (cyclins, growth factors)
• apoptosis/cell survival (regulators of apoptosis)

Question 2

NF-kB structure?

Answer 2

Dimer

NF subunit, kB subunit (5 types of each subunit)

Question 3

IkB subtypes?

Answer 3

alpha, Beta, epsilon

Question 4

Mechanism of inhibition via IkBalpha on NF-kB?

Answer 4

IkBa bound to NF-kB protein

Ligand binding to receptor signals eventually lead to activation of IKK

IKK phosphorylates bound IkBa -> targets it for degradation via proteosome

After degradation, NF-kB free to act -> enters nucleus via NLS

NF-kB increases IkBa expression -> long delay negative feedback

Question 5

Hypothesis design: what is the point of having 3 subtypes of IkB? Experiment design?

Answer 5

1) Simulate signalling without Beta or Episilon
- predicted simulation shows long delay negative feedback oscillation of signal (expected)

2) Electrophoretic mobility shift assay: