III. Signal transduction and cell cycle | 43. NFκB and TGFβ signaling Flashcards
I. NF-κB SIGNALING
1. What is NF-κB?
NF-κB = Nuclear factor kappa-light-chain-enhancer of activated B cells
NF-κB is a transcription factor, which will translocate from the cytosol to the nucleus when activated, where it regulates expression of more than 200 genes involved in immune response, inflammation, proliferation and apoptosis.
I. NF-κB SIGNALING
2. How can NF-κB be activated?
NF-κB can be activated by cytokines, growth factors, bacterial products, viruses and oxidative stress.
I. NF-κB SIGNALING
3A. What is the medical significance of NF-κB?
- NF-κB is broadly used by eukaryotic cells as a regulator of genes that control cell survival and proliferation.
=> misregulated NF-κB-signaling can lead to different pathological processes (diseases) - NF-κB also controls genes involved in inflammation, and is therefore chronically active in many inflammatory diseases (asthma, arthritis, sepsis etc.)
- Cancers can arise from sites of infection and inflammation. Many human tumors have misregulated, constitutively active NF-κB. It is either due to mutations in genes encoding the NF-κB TFs or in genes that control NF-κB activity
I. NF-κB SIGNALING
4. What is the structure of NF-κB?
A heterodimer with one small and one bigger subunit (50kDa and 65kDa respectively)
- p50: consists of NF-κB1 and NF-κB2
- p65: consists of RelA, RelB and c-Rel. This is the transcription activation domain
I. NF-κB SIGNALING
5. Why can NF-κB be inactive?
Without any stimulus, NF-κB is inactive and retained in the cytosol by IκB (Inhibitor
of κB). IκB masks the nuclear localization signals (NLS) of NF-κB proteins
I. NF-κB SIGNALING - NF-κB signaling pathway
6A. What are other components of NF-κB signaling pathway?
- IKK
- IκB kinase complex, a serine/threonine protein-kinase of 3 subunits: IKKα and IKKβ (protein kinases) and IKKγ (adaptor) - TAK1:
- a serine/threonine protein-kinase which phosphorylates (activates) IKKβ using the canonical route - NIK
- NIK: a serine/threonine protein-kinase which phosphorylates (activates) IKKα using the non-canonical route - Ubiquitin
- a small protein of 76 AAs.
- Ubiquitination = addition of ubiquitin to a protein.
- Polyubiquitination is a signal for degradation in the 26S proteasome, but can have other functions as well (protein assembling) - Ubiquitin-ligases:
- couple ubiquitin to proteins with high selectivity for the target
I. NF-κB SIGNALING - NF-κB signaling pathway
6B. What are the features of IKK?
IKK
- IκB kinase complex, a serine/threonine protein-kinase of 3 subunits: IKKα and IKKβ (protein kinases) and IKKγ (adaptor)
I. NF-κB SIGNALING - NF-κB signaling pathway
6C. What are the features of TAK1?
TAK1:
- a serine/threonine protein-kinase which phosphorylates (activates) IKKβ using the canonical route
I. NF-κB SIGNALING - NF-κB signaling pathway
6D. What are the features of NIK?
NIK
- NIK: a serine/threonine protein-kinase which phosphorylates (activates) IKKα using the non-canonical route
I. NF-κB SIGNALING - NF-κB signaling pathway
6E. What are the features of Ubiquitin?
Ubiquitin
- a small protein of 76 AAs.
- Ubiquitination = addition of ubiquitin to a protein.
- Polyubiquitination is a signal for degradation in the 26S proteasome, but can have other functions as well (protein assembling)
I. NF-κB SIGNALING - NF-κB signaling pathway
6F. What are the features of Ubiquitin-ligases?
Ubiquitin-ligases:
- couple ubiquitin to proteins with high selectivity for the target
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7A. What are the steps in Canonical route of NF-κB activation?
1) Inflammatory mediators (TNFα = tumor necrosis factor, IL-1β = interleukin 1 beta, LPS = lipopolysaccharide) bind to specific receptors, and together with adaptors, will recruit a scaffold protein and a ubiquitin-ligase from the cytosol
2) The ubiquitin-ligase polyubiquitinates the scaffold protein
3) The polyubiquitinated scaffold protein recruits the kinase TAK 1 (which activates IKKβ) and IKK complex (Inhibitor of κB kinase complex) [IKKα + IKKβ]
4) Kinase TAK1 phosphorylates and activates IKKβ
5) IKKβ phosphorylates IκB
6) The phosphorylated IκB is recognized and polyubiquitinated by a ubiquitin-ligase
7) The polyubiquitinated IκB is degraded in the 26s proteasome
8) IκB will disappear and the released NF-κB TF will translocate into the nucleus and activate transcription
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7B1. What are the Inflammatory mediators in Canonical route of NF-κB activation?
- TNFα = tumor necrosis factor
- IL-1β = interleukin 1 beta
- LPS = lipopolysaccharide
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7B2. What do the Inflammatory mediators in Canonical route of NF-κB activation?
Inflammatory mediators bind to specific receptors, and together with adaptors, will recruit a
scaffold protein and a ubiquitin-ligase from the cytosol
(1. TNFα = tumor necrosis factor
2. IL-1β = interleukin 1 beta
3. LPS = lipopolysaccharide)
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7C. What is the role of The ubiquitin-ligase in Canonical route of NF-κB activation?
The ubiquitin-ligase polyubiquitinates the scaffold protein
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7D. What does polyubiquitinated scaffold protein do in Canonical route of NF-κB activation?
The polyubiquitinated scaffold protein recruits the kinase TAK 1 (which activates IKKβ) and IKK complex (Inhibitor of κB kinase complex) [IKKα + IKKβ]
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7D. What does Kinase TAK1 do in Canonical route of NF-κB activation?
Kinase TAK1 phosphorylates and activates IKKβ
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7E. What does IKKβ do in Canonical route of NF-κB activation?
IKKβ phosphorylates IκB
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7F. How is the polyubiquitinated IκB degraded?
The polyubiquitinated IκB is degraded in the 26s proteasome
I. NF-κB SIGNALING - Canonical route of NF-κB activation
7G. The phosphorylated IκB is recognized and polyubiquitinated by _____
a ubiquitin-ligase
I. NF-κB SIGNALING - Non-conical route of NF-κB activation
8A. What are the steps of non-conical route of NF-κB activation?
1) Without stimulus, a complex of adaptors and ubiquitin ligases polyubiquitinates the NIK kinase. NIK is continuously degraded in the proteasome
2) Other inflammatory mediators bind to the receptors. The receptor recruits the complex which in turn polyubiquitinates itself, and this complex is degraded. The NIK kinase is accumulated and auto-activated
3) Active NIK phosphorylates and activates IKKα. The substrate of IKKα, p100 is a precursor of NF-κB and has an IκB-like domain with which it holds Rel in the cytosol
4) IKKα phosphorylates p100, which is then polyubiquitinated by a ubiquitin-ligase. Ubiquitinated p100 will undergo limited proteolysis: its IκB-like domain is lost, but the remaining p52 = mature NF-κB2 translocates to the nucleus with Rel
=> Induction of the IκB gene will switch NF-κB off
I. NF-κB SIGNALING - Non-conical route of NF-κB activation
8B. What will active NIK do in non-conical route of NF-κB activation
- Active NIK phosphorylates and activates IKKα.
- The substrate of IKKα, p100 is a precursor of NF-κB and has an IκB-like domain with which it holds Rel in the cytosol
I. NF-κB SIGNALING - Non-conical route of NF-κB activation
8B. What will IKKα do in non-conical route of NF-κB activation
- IKKα phosphorylates p100, which is then polyubiquitinated by a ubiquitin-ligase.
- Ubiquitinated p100 will undergo limited proteolysis: its IκB-like domain is lost, but the remaining p52 = mature NF-κB2 translocates to the nucleus with Rel
II. TGFβ SIGNALING
1. What is TGFβ?
- TGFβ (transforming growth factor β) is a first messenger, 25kDa polypeptide and forms a dimer.
- Its important effects are inhibition of cell proliferation, immunosuppression, regulation of extracellular matrix synthesis and induction of apoptosis.
II. TGFβ SIGNALING
2. What are the important effects of TGFβ?
Its important effects are
- inhibition of cell proliferation
- immunosuppression
- regulation of extracellular matrix synthesis and induction of apoptosis.
II. TGFβ SIGNALING
3A. What does the TGFβ superfamily consist of?
The TGFβ superfamily consists of 40 different cytokines divided into 2 major groups:
1. Bone morphogenic proteins (BMP)
2. TGFβ/ activin proteins
II. TGFβ SIGNALING
3B. In the TGFβ superfamily, what is the role of Bone morphogenic proteins (BMP)?
- Induce the formation of bone and cartilage
- Play important role in tissue architecture throughout the body
II. TGFβ SIGNALING
3B. In the TGFβ superfamily, what is the role of TGFβ/ activin proteins?
- TGFβ: includes endogenous growth inhibiting proteins. Has immunosuppressive functions
- Activin: enhances FSH biosynthesis and secretion,
regulation of menstrual cycle. Roles in wound repair, immune response, metabolism and so on
II. TGFβ SIGNALING - TGFβ receptors
4A. What are the 2 types of TGFβ receptors and their features?
- We have 2 types of TGFβ receptors: TGFβ receptor I and TGFβ receptor II
=> both are enzyme-coupled receptors that are single transmembrane domain proteins with serine/threonine kinase activity on the cytosolic side of the plasma membrane
II. TGFβ SIGNALING - TGFβ receptors
4B. What is the structure of TGFβ receptor I?
TGFβ receptor I has a GS domain where 5 residues can be phosphorylated
II. TGFβ SIGNALING - SMAD proteins
5A. What are SMAD proteins?
- SMAD proteins are a family of proteins that are the main signal transducers for the TGFβ pathways.
- There are 3 types of SMAD proteins
II. TGFβ SIGNALING - SMAD proteins
5B. What are the 3 types of SMAD proteins and their structures?
SMAD proteins are a family of proteins that are the main signal transducers for the TGFβ pathways
II. TGFβ SIGNALING - Signaling with TGFβ
6A. What are the 5 steps of Signaling with TGFβ?
1) TGFβ arrives at the cell surface and binds to type II receptor, which will eventually cause heterodimerization -> active receptor complex (type I & II receptor + TGFβ)
2) Type II receptor will phosphorylate the GS domain of type 1 receptor => activates type I receptor
3) SARA (SMAD anchor for receptor activation) will bind to the receptor and phosphorylate R-SMAD (receptor regulated SMAD)
4) The phosphorylated R-SMAD will meet co-SMAD (common SMAD) in the cytosol and form a dimer, which is translocated into the nucleus
5) It will activate gene expression and can express proteins which will restrain the cell division cycle
II. TGFβ SIGNALING - Signaling with TGFβ
6B. What does Type II receptor do in Signaling with TGFβ?
Type II receptor will phosphorylate the GS domain of type 1 receptor => activates type I receptor
II. TGFβ SIGNALING - Signaling with TGFβ
6B. What does SARA do in Signaling with TGFβ?
SARA (SMAD anchor for receptor activation) will bind to the receptor and phosphorylate R-SMAD (receptor regulated SMAD)
II. TGFβ SIGNALING - Signaling with TGFβ
6C. What does The phosphorylated R-SMAD do in Signaling with TGFβ?
The phosphorylated R-SMAD will meet co-SMAD (common SMAD) in the cytosol and form a dimer, which is translocated into the nucleus
II. TGFβ SIGNALING - Signaling with TGFβ
7. What is the role of TGFβ signaling?
- TGFβ signaling will inhibit cell proliferation, causes differentiation or induces apoptosis.
- The loss of it speeds up proliferation, helps dedifferentiation -> cancer
