14-3-3 Binding Phosphoproteome

Question 1

How do 14-3-3 proteins bind to proteins?

Answer 1

Through specific tandem pairs of phosphosites

Question 2

What is the dual-docking conformation?

Answer 2

14-3-3 will act as a singalling intergrator when its two binding sites on a target are phosphorylated by different kinases

Question 3

How can 14-3-3 binding affect its binding protein?

Answer 3

Can mask a domain
If bound to a disordered region, it can force a disorder-to-order transition which creates a new functional site in the target
Alter the interactions between the domains in a protein

Question 4

What is the 14-3-3 structure?

Answer 4

As homo- and hetero- dimers with phosphopeptides in them

Question 5

Where are 14-3-3 located?

Answer 5

In every eukaryotic cell

Question 6

What is an ‘AND’ gate?

Answer 6

A coincidence detector

Needs 2 inputs before it can generate an output

Question 7

Name a protein which 14-3-3 masks

Answer 7

Q96RKO CAPICUA

Question 8

How does 14-3-3 mask Capicua?

Answer 8

HMG is straddled in Capicua

the DNA interaction site is masked since HMG is phosphorylated twice and therefore 14-3-3 bind to and masks it

Question 9

What is ASK1?

Answer 9

Apoptosis signal-regulating kinase 1

Member of MAP3K5

Question 10

How does 14-3-3 inhibit MAP3K5?

Answer 10

It is phosphorylated on linker region which binds to the central groove of 14-3-3. This pulls the kinase domain against the back end of 14-3-3 and inhibits the kinase activity since ATP accessibility is removed

Question 11

What is ZNRF2?

Answer 11

A RING-E3 Ubiquitin ligase

Question 12

What is the structure of ZNRF2?

Answer 12

It is tethered to the intracellular membrane through its lipid tail

Question 13

What is the function of ZNRF2?

Answer 13

Facilitates the ubiquitylation of ion channels and regulates the trafficking of ions when it is tethered to the membrane

Question 14

What phosphorylates ZNRF2?

Answer 14

PKA
PKB
PKC

Question 15

What does ZNRF2 phosphorylation induce?

Answer 15

14-3-3 binding

Question 16

What happens when 14-3-3 binds to ZNRF2?

Answer 16

It is soluble and inhibited - dissociated from the membrane and goes into the cytoplasm

Question 17

What is GNF-2?

Answer 17

Memetic myristoyl

Binds to BCR-Abl myristoyl pocket

Question 18

How does GNF-2 impact ZNRF2?

Answer 18

It stops ZNFR2 from leaving the membrane, even when phosphorylation has occured

Question 19

What do the kinases which create phosphosites do and what is specific for their phosphorylation?

Answer 19

Phosphorylate a serine or threonine

They never have a proline in the +1 position

Question 20

What does insulin do in terms of 14-3-3?

Answer 20

Generates phosphosites for 14-3-3 to bind in the spleen, kidney, pancreas, adipose and gastro

Question 21

How does insulin drive glucose into skeletal muscles?

Answer 21

Through the translocation of GLUT4 in intracellular vesicles to the plasma membrane

Question 22

How does 14-3-3 aid GLUT4 translocation in response to insulin?

Answer 22

AS160 is phosphorylated and 14-3-3 docks onto it -> inhibits Rab-GAP activating protein -> gets loaded with GTP -> Activate Rab bring GLUT4 to the plasma membrane

Question 23

What is AS160?

Answer 23

a 14-3-3 binding protein which has an active site of Rab-GAP

Question 24

What are Rabs involved in?

Answer 24

Specific vesicle trafficking events

Question 25

what disease is associated with AS160 mutation?

Answer 25

Type 2 diabetes

Question 26

What is the phenotype of AS160 mutated type 2 diabetes? And what does this suggest?

Answer 26

Heavy pigmentation behind the neck and knees | Involved in trafficking melanin containing vesicles in the skin cells

Question 27

What happens when there is no amino acids in the heart available for it to function?

Answer 27

The heart can use glucose from AS160

Question 28

What is the function of SPEG?

Answer 28

Can bind to SERCA2A and phosphorylates it in response to insulin

Question 29

Where is SPEG found?

Answer 29

The heart

Question 30

What is SERCA2A?

Answer 30

A calcium pump

Question 31

What happens if SPEG is mutated at phosphorylated sites?

Answer 31

Impaired Ca2+ reuptake into SR and cardiac dysfunction

Question 32

What processes does insulin-induced 14-3-3 binding downregulate?

Answer 32

Lipolysis Gluconeogenesis Autophagy

Question 33

What processes does insulin-induced 14-3-3 binding upregulate?

Answer 33

``` Glucose uptake Amino acid uptake K+ and PO43- uptake Glycogen synthase Glycolysis in the heart Lipid synthesis Esterification of fatty acids ```

Question 34

What physiological functions does insulin-induced 14-3-3 binding control?

Answer 34

Arterial muscle tone relaxation Endothelial permeability and inflammation Cytoskeletal remodelling HCl production by parietal cells of the stomach Bone remodelling Feedback control of pancreatic function Brain functions

Question 35

If you have diabetes, what are common chronic secondary diseases?

Answer 35

renal disease | cardio myopathy

Question 36

How does the brain respond to insulin?

Answer 36

A huge dephosphorylation of 14-3-3 binding proteins | Inhibits cerebral PKA

Question 37

What protein in the brain is associated with Alzheimer's?

Answer 37

TAU

Question 38

How can diabetes cause Alzheimer's?

Answer 38

An increase in the insulin levels causes neurofibril tangles in the brain

Question 39

In fasted animals, what does 14-3-3 bind to?

Answer 39

PhosphoSer? | PhosphoSer948

Question 40

Ser858 is phosphorylated in fasted animals, what does this do?

Answer 40

Activates Rac1 GAP domain

Question 41

In fasted animals, what does phosphoFAM122A - 14-3-3 complex do?

Answer 41

It is translocated to the nucleus, which inhibits dephosphorylation of TAU by PP2A

Question 42

How is sleep linked to insulin resistance?

Answer 42

1 night of bad sleep can lead to insulin resistance to a higher degree compared to a high fat diet for several months