Final: Ch 16 Tyrosine Kinases, Ras/MAPK Flashcards Preview

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Flashcards in Final: Ch 16 Tyrosine Kinases, Ras/MAPK Deck (88):
1

3 things transcription of genes is influenced by

Chromatin structure

Epigenetic modifications of histones

What TFs and proteins the cell contains

2

4 basic types of signal transduction pathway

Receptor-associated kinase (RTK/cytokine receptor)

Cytosolic kinase (GPCR)

Protein subunit dissociation

Protein cleavage

3

kinases phosphorylate what two types of kinases

tyrosine kinase (cancer)

serine/thr kinase

4

tyrosine kinases phosphorylate __________ as a part of activation

eachother

5

What 2 categories of receptor activate tyrosine kinases?

Receptor tyrosine kinase

Cytokine receptors

6

the target of a kinase is often a...

kinase

7

specifically, tyrosine kinases phosphorylate what?

specific tyrosine residues on target proteins

Autophosphorylate the receptor

8

what do the phosphorylated (by a tyrosine kinase) target proteins do?

activate one or more signaling pathways for cell proliferation, differentiation, survival, and metabolism

9

2 types of receptors that activate tyrosine kinases

receptor tyrosine kinases (RTK) - the tyrosine kinase is an intrinsic part of the receptor's pp chain (encoded by same gene)

cytokine receptors - receptor and kinase bound tightly together but encoded by different genes

10

for cytokine receptors, the tightly bound ______ is known as a...

kinase, JAK kinase (just another kinase)

11

are tyrosine kinase receptors dimers?

most are monomeric, but ligand binding induces conformational change to form a dimer receptor

ex. insulin receptor is already a dimer

12

2 ways kinases are inactivated

by a phosphatase - remove phosphate from target protein

by endocytosis - internalize the receptor

13

what types of signaling molecule activate RTKs?

soluble or membrane bound peptides or hormones

ex. growth factors EGF, NGF

14

3 domains of a RTK

extracellular ligand-binding domain

1 transmembrane alpha helix

cytosolic domain with protein tyrosine kinase activity

15

activation lip

flexible domain in cytosolic side of RTK

unphosphorylated without ligand binding - blocks kinase activity (or binding of ATP like in the insulin receptor)

after ligand binding, the receptor sometimes and one kinase phosphorylates a tyrosine residue in the activation lip of the other to activate its kinase activity

16

how does the conformational change in the phosphorylated activation lip activate kinase activity?

reduces the Km for ATP or the substrate

17

how does activation of epidermal growth factor receptor by EGF work?

EGF ligand binds and forms an asymmetric kinase dimer

the activator kinase binds the juxtamembrane segment of the receiver kinase

this causes a conformational change that removes the activation lip from the kinase active site of the receiver

the active kinase phosphorylates tyrosine residues in the receptor

18

_ RTSs participate in signaling by the members of the ___ family of signaling molecules

4, EGF

19

in humans, the _ members of the ___ family are...

4, HER (human epidermal growth factor receptor)

HER 1-4

HER1 - homodimerization

HER2- heterodimerization

20

Gene amplification of HER2

Error in DNA replication makes multiple copies of HER2

HER2 makes cells more sensitive to EGF

21

amplification of the HER2 gene

25% of breast cancer - overexpression of HER2 protein in tumor cells

makes the tumor cells sensitive to growth my low levels of any member of the EGF family

22

an effective therapy for HER2 overexpression

monoclonal antibodies for the HER2 protein

23

cytokines

small, secreting signaling molecules that control growth and differentiation of specific cells

ex. interleukins (proliferation and functioning of T cells and Ab producing B cells) --> TNF-alpha

24

prolactin

a cytokine that during pregnancy, causes the epithelial cells lining the mammary gland ducts to differentiate into acinar cells that produce milk

25

interferons

cytokines that are secreted after viral infection

act on nearby cells to induce enzymes that make the cells resistant to viral infection

26

many cytokines induce formation of important types of _____ cells

blood

ex. erythropoietin (Epo) - triggers production of RBC

27

binding of a cytokine to its receptor activates a tightly bound ___ protein tyrosine kinase

JAK

28

all cytokines evolved from a common ancestral protein and have a similar tertiary structure consisting of...

4 long alpha helices folded together

29

cytokine receptors do not posses _______ enzymatic activity

intrinsic - instead a JAK kinase is tightly bound to the cytosolic domain

30

JAK stands for

just another kinase

31

the 4 members of the JAK kinase family 3 domains...

N-terminal receptor binding domain

C-terminal kinase domain (poorly active)

middle domain that regulates kinase activity

32

how are JAK kinases activated?

just like in RTKs - ligand binding induces conformational change to form a dimer where the JAKs are close together so they can phosphorylate each other

this phosphorylation enhances ATP/substrate affinity to increase kinase activity

33

what purpose do the phosphorylated tyrosine residues serve?

binding sites for proteins that have phosphotyrosine binding domains

ex. SH2 domain

34

what determines which phosphotyrosine an SH2 will bind?

variations in the hydrophobic pocket In the SH2 domain

35

phosphotyrosine binding domain (PTB)

found on multidocking proteins

Bind phosphorylated tyrosines on the receptor

36

multidocking protein

serve as docking sites for multiple signal transduction proteins

ex. IRS-1 (insulin receptor substrate) is phosphorylated by the receptor and the tyrosines (on IRS-1) serve as docking sites for SH2-containing signaling proteins

37

JAK kinases activate ____ transcription factors

STAT TFs

38

3 domains of STAT

N-terminal DNA binding domain

SH2 domain that binds phosphotyrosines in a cytokine's cytosolic domain

C-terminal domain with a critical tyrosine residue

39

how does JAK-STAT work?

SH2 domain on STAT binds the activated cytokine receptor at a phosphotyrosine

JAK kinase phosphorylates STAT's C-terminal tyrosine

STAT dissociates from the receptor and dimerizea, exposing a nuclear-localization signal

the STAT dimer moves into the nucleus and binds specific enhancers to alter gene expression

40

are the genes available to be activated by any STAT different?

yes

in mammary glands, STAT5 induces transcription of genes encoding milk proteins

when STAT5 is activated in erythroid progenitor cells, it induces transcription of the bcl-xl gene that prevents apoptosis

41

prolonged treatment of cells with ligand often _____ the number of available cell-surface receptors so there is a less robust response to ligand than at the beginning of treatment

reduces (desensitization)

42

desensitization example HER1/EGF

in absence of EGF, HER1 receptors are long lived (receptor-mediated endocytosis occurs at a slow rate)

after EGF binding, endocytosis of HER1 is increased and only a few receptors return to the membrane, most are degraded in lysosomes

43

do HER1 mutants that lack kinase activity undergo accelerated endocytosis in the presence of ligand?

no

perhaps b/c activation of kinase activity in normal HER1 induces conformational change that exposes a sorting motif that signals endocytosis

44

can internalized receptors signal from within endosomes before their degradation?

yes

evidence: binding signaling proteins like Grb-2 and Sos

45

2 things that can happen to internalized receptors

returned to cell membrane

degraded in lysosomes

46

ubiquitination

addition of the small protein ubiquitin to a lysine in a protein signals it for degradation

enzyme c-Cbl does this

47

SHP1

phosphatase that negatively regulates signaling from cytokine receptors

inactivates JAK protein by binding to active receptor

has 2 SH2 domains

48

SHP1 in inactive state

one SH2 domain of SHP binds the catalytic site so it is inactive

49

SHP1 in active state

The blocking SH2 domain binds a phosphotyrosine in the active receptor

Catalytic site placed near phosphate in JAK activation lip and removes

50

SOCS proteins

terminate signaling from cytokine receptors by negative regulation

SH2 domain binds activated receptor to block binding of other SH2 containing proteins (competitive inhibition)

51

SOCS-1 mechanism

binds critical phosphotyrosine in activation lip of activated JAK2 kinase

52

SOCS box

domain that recruits ubiquitin ligases

Ubiquitin is a signal for degradation in proteasomes

53

almost all RTK and cytokine receptors activate which pathway

Ras/MAP

54

Ras protein

small monomeric G protein

member of the GTPase superfamily of switch proteins

55

what does activated Ras promote at the membrane

formation of signal transduction complexes containing 3 sequentially acting protein kinases (kinase cascade)

The kinase cascade activates MAP kinases

56

what do members of the MAP kinase family do

translocate into the nucleus and phosphorylate many proteins

ex. TFs that regulate expression of proteins for the cell cycle

57

Ras activation is accelerated by

guanine nucleotide factor (GEF)

binds the Ras*GDP complex to cause dissociation of GDP

58

what deactivates Ras

GTPase activating protein (GAP)

catalyzes hydrolysis of GTP to GDP to inactivate Ras (Ras has a low intrinsic hydrolysis rate)

59

does Ras operate downstream of most RTKs and cytokine receptors?

yes

60

the compound eye of the fly is composed of 800 individual eyes called _______

ommatidia

each ommatidium consists of 22 cells, 8 of which are photosensitive neurons called retinula or R cells (R1-R8)

61

Sevenless (Sev)

an RTK that regulates development of R7 cell in an ommatidium

62

flies with mutant Sev gene

R7 cell does not form, flies can't see UV light

63

Boss (bride of Sevenless)

protein expressed on the surface of R8 cells - is the ligand for the Sev RTK on the surface of the R7 precursor cell

no Boss, no R7 cells

64

fruit fly experiment with Sev + Ras

temperature sensitive Sev mutant formed R7 cells at an intermediate temp

2nd mutation in 1/3 other important proteins in the pathway produced no R7 cells (SH2 adapter, Sos-a GEF, Ras)

65

always active Ras in Sev mutant fly

produces R7 cells even though no Sev

proves that activation of Ras is a critical step in intracellular signaling by most if not all RTKs and cytokine receptors

66

RTKs and JAK kinases are linked to Ras by _______ proteins

adapter - serve as a link, not catalytic

GRB2 - links activated receptor (SH2) and SoS (SH3)

SoS (Son of Sevenless) - a GEF that speeds up GTP binding to Ras

67

SoS (Son of Sevenless)

a guanine nucleotide exchange protein (GEF) that catalyzes conversion of inactive GDP-Ras to active GTP-Ras

68

binding of SoS to inactive Ras causes a conformational change that triggers..

dissociation of GDP so GTP can bind and activate Ras

69

signals pass from activated Ras to a cascade of protein _______, ending with ____ _______

kinases, MAP kinase

70

mitogen

a ligand that stimulates growth and division (cell cycle)

71

MAP kinase (MAPK mitogen activated protein kinase)

cytosolic kinase

gets phosphorylated by MEK and activated, then amplifies the signal by phosphorylating nuclear TFs

72

Melanoma

Always active B-Raf stimulates MEK/MAP to make skin cancer

73

Ras/MAPK pathway

active Ras*GTP binds N-terminal domain of Raf kinase

hydrolysis of Ras*GTP to Ras*GDP releases active Raf

Raf phosphorylates MEK which phosphorylates MAP

MAP phosphorylates nuclear TFs

74

mutant Raf kinase without N-terminal regulatory domain

always active and induce cultured cells to proliferate in absence of stimulation by growth factors

also tumor cells

75

does MAPK dimerize?

yes, after phosphorylation a conformational change occurs and it dimerizes

the dimer is translocated into the nucleus where it phosphorylates TFs

76

what does quiescent mean

non-growing

77

early response genes

induced well before cells enter S phase

ex. one of these genes encodes the TF c-Fos

78

c-Fos

TF that induces expression of many genes encoding proteins required for cells to move through the cell cycle

79

most RTKs that find growth factors utilize the MAPK pathway to activate genes encoding proteins like _-__, which propel the cell through the cell cycle

c-Fos (a TF)

80

the enhancer that regulates the c-Fos gene contains a _______ _______ ______

serum response element - an enhanced that binds many TFs

81

MAPK induces transcription of the c-Fos gene by direct activation of which TF, and indirect activation of another TF

direct: ternary complex factor

indirect: serum response factor

82

association of phosphorylated ternary complex factor with two molecules of phosphorylated serum response factor forms... I

an active trimeric factor that activates gene transcription

83

_____ transmit signals to MAPK in yeast mating pathways

GPCR

G-betagamma triggers a kinase cascade like Ras/MAPK cascade

84

_____ proteins separate multiple MAPK pathways

scaffold

allow the kinases of one pathway to interact with one another, but not with kinases from other pathways

ex. Ksr (kinase suppressor of Ras) binds MEK and MAPK

85

basic idea of scaffold proteins

coordinates responses by organizing free floating proteins

ex. MAPK on scaffold activated in Ras/MAPK pathway

MAPK-P can enter nucleus and phosphorylate TFs

86

mitogens up...

cell proliferation

cell cycle

active in cancer

87

weird note about nutrients for animal cells

90% defined medium, 10% fetal calf serum (growth factors)

eliminate 10% = serum starved cells

look at signaling pathways activated when it is added back

88

Dominant-negative mutation

A loss of function in cells with wild-type copies

Mutant prevents normal protein from functioning