Lecture 10 - ras-Dependent Signaling Flashcards Preview

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Flashcards in Lecture 10 - ras-Dependent Signaling Deck (59):
1

What is the connection between ras-dependent signaling and oncogenes?

A lot of retrovirus oncogenes have downstream ras-dependent signaling pathways

2

What are the 2 types of oncogenes?

1. v-src: oncogene in a retrovirus
2. c-src: native proto-oncogene in cell

3

What is a proto-oncogene? What does it code for?

Normal gene that could become an oncogene due to mutations or increased expression. Proto-oncogenes code for proteins that help to regulate cell growth and differentiation.

4

What is a defective oncogenic virus?

Virus that has lost viral functions and takes advantage of an infection by helper viruses by infecting at the same time and inserting their genome into the helper virus package to propagate

5

What is Ras?

Monomeric super family of GTPases

6

What is the function of Ras?

Regulate cell growth through Ser/Thre protein kinases

7

What are the 5 subfamilies of Ras?

1. Ras
2. Rho
3. Arf
4. Rab
5. Ran

8

What is the function of Rho?

Reorganizes cytoskeleton through Ser/Thr protein kinases

9

2 other names for GTPase?

GTP binding protein = G protein

10

Compare the rate of the ras GTPase activity to that of the G alpha GTPase activity

Slower

11

Compare the rate of the ras GDP dissociation to that of the G alpha GDP dissociation

Same

12

What % of cancers have activating mutations of c-ras?

30%

13

How do ras GTPase make up for their slower rate of GTPase activity?

They recruit accessory proteins = GTPase activating proteins (GAPs)

14

What is the function of GAPs? How do they do it?

They markedly accelerate the intrinsic activity of c-ras by 10^4-10^5 by either activating the GTPase activity or accelerating GDP dissociation

15

What is the connection between GAPs and trimeric G proteins? What can we call this?

Trimer G proteins' substrates (eg: PLPC-beta) have GAP like activity resident on the G alpha subunit binding site so that when it binds, the substrate is activated but also it triggers the subunit to hydrolyze GTP

ANOTHER built-in OFF switch mechanism

16

What is the purpose of receptor tyrosine kinases recruiting GAPs to bind on phosphotyrosines?

These receptors are capable of inactivating ras by stimulating GTP hydrolysis

17

How is ras activated?

GDP dissociation + GTP binding

18

How is ras inactivated?

GTP hydrolysis

19

What proteins are responsible for activating ras? 3 names

How do they do this?

Guanine nucleotide exchange factor(GEF) = guanine nucleotide releasing protein (GNRP) = guanine nucleotide exchange protein (GNEP)

They markedly accelerate GDP dissociation

20

What are guanine nucleotide dissociation inhibitors (GDIs)?

They stabilize the c-ras-GDP state, stimulating c-ras inactivation

21

Inhibition of GAP: ras activation or inactivation?

Activation

22

Activation of GAP: ras activation or inactivation?

Inactivation

23

Inhibition of GEF: ras activation or inactivation?

Inactivation

24

Activation of GEF: ras activation or inactivation?

Activation

25

Inhibition of GDI: ras activation or inactivation?

Activation

26

Activation of GDI: ras activation or inactivation?

Inactivation

27

How is Rho activated/inactivated?

Same mechanisms as for Ras

28

Where are GEFs found? 2 places

1. Bound to receptor tyrosine kinases via a Grb-2 adaptor protein
2. In the nucleus

29

What controls if ras is activated or inactivated by receptor tyrosine kinases?

Whether GEFs or GAPs are recruited to bind to receptor tyrosine kinases

30

Once activated, what does ras do?

Induces the MAP-kinase cascade

31

Describe the 5 steps of the MAP-kinase cascade.

1. Ras binds MAP-kinase-kinase-kinase and activates PKC to phosphorylate it = activation
2. MAP-kinase-kinase-kinase phosphorylates MAP-kinase-kinase
3. MAP-kinase-kinase phosphorylates MAP-kinase
4. MAP-kinase dissociates from the scaffold to go into the nucleus
5. MAP-kinase phosphorylates substrates

32

What does MAP stand for?

Mitogen Activated Protein

33

When is MAP active?

Under conditions of cell division or growth

34

What are the 2 types of substrates that MAP-kinase phosphorylates?

1. Proteins
2. Gene regulatory proteins

35

What kinds of kinases are MAP kinases?

Ser/Thr EXCEPT for MEK, which is a Tyr kinase

36

What is another name for MAP kinase?

ERK = Extracellular Regulated Kinase

37

What is another name for MAP kinase kinase?

MEK

38

What are 2 other names for MAP kinase kinase kinase?

1. MEKK
2. raf

39

How do the MAP kinases recognize their substrates?

They do not recognize specific sequence but recognize specific conformations

40

What is PAK kinase?

MAP-kinase-kinase-kinase- kinase

41

Does each type of receptor/category of activity have its own MAPK cascade within a cell? Examples? What is a possible reason for this?

YUP

Examples: cell cycle arrest, glycerol synthesis, filamentous growth, cell wall remodeling, meiosis, sporulation (all examples in yeast)

Reason: substrates can enter the MAP kinase pathways at different points, so it would be hard to only have one cascade per cell

42

If one of the MAP kinases in one activity pathway fails, can a MAP kinase from another cascade come save this activity? Why/Why not?

NOPE because these kinases are held together by scaffolds and are functioning like a single unit

43

What are 4 different types of scaffolds holding MAP kinases cascades together?

1. MP1
2. JIP1
3. STE5
4. PBS2

44

What is sevenless?

Mutated membrane protein receptor tyrosine kinase that is activated by "bride of sevenless" and can then be bound by "son of sevenless", a GEF (with a Grb-2 adaptor, of course) which activates ras

45

What is another name for Grb-2?

Drk = downstream of receptor kinase

46

What are DUSPs? Function?

Dual-Specificity Protein Phosphatases that dephosphorylate MAP kinases (Ser/Thr OR Tyr)

47

What are the 2 types of genes activated through ras signaling? 2 names for each

1. Immediate early genes = primary response genes
2. Late response genes = secondary response genes

48

Timeframe of transcription induction for primary response genes?

Minutes

49

Is transcription induction of primary response genes dependent on protein synthesis? What does this mean?

NOPE

The proteins that are activating these genes already exist and do not need to be synthesized

50

Is the shutoff of transcription of primary response genes dependent on protein synthesis?

YUP

51

What is the mRNA half-life of primary response genes?

Short: 5-15 min

52

Over what time frame are secondary response genes expressed?

Hours

53

Is transcription induction of secondary response genes dependent on protein synthesis?

YUP

54

Describe the expression of primary response genes in non-active cells?

LOW

55

Describe the cascade of events that happens once the MAP kinase enters the nucleus.

1. Phosphorylates proteins, which are now activated
2. Activated proteins bind to primary response gene sequences to induce expression (protein synthesis independent)
3. mRNA is produced and then translated into primary response proteins
4a. Proteins are transcription factors that induce the expression of secondary response genes (protein synthesis dependent)
4b. Proteins destabilize the mRNA or turn off expression of the primary response gene (shut off is protein synthesis dependent)

56

When does the concentration of primary response mRNA drop?

When the concentration of secondary response mRNA rise

57

What are the 5 kinases that phosphorylate gene regulatory proteins?

1. PKA
2. CaM-K
3. PKC
4. MAP-kinase
5. PKB

58

How does MAP kinase phosphorylation allow it to dissociate from the scaffold to go into the nucleus?

Phosphorylation reveals its nuclear localization signals

59

What happens when the srn proto-oncogene is associated with the Rous sarcoma virus?

Src oncogene is often expressed at abnormally high levels, contributing to unregulated cell division and cancer