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BIO 317 > 20 - Signaling in Time and Space > Flashcards

20 - Signaling in Time and Space Flashcards

1
Q

True or false: simple motifs can display complex dynamics

A

True: these motifs can be arranged into loops and self-regulation

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2
Q

True or false: segregation can lead to specificity

A

True: segregating signaling determinants can increase specificity

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3
Q

How can diffusion alter the rate of a signaling pathways?

A

Diffusion of substrates into different regions (kinases and phosphatases) can alter the rate of the signaling pathway

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4
Q

What happens when activated Erk is added to cells?

A

It can do some nerve-y things, but cannot mimic the full effects of NGF

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5
Q

What is the conclusion of adding activated Erk to cells?

A

NGF has branched signaling, where it doesn’t all go through Erk

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6
Q

What is another name for MAP kinase kinase kinase?

A

Raf

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7
Q

What is another name for MAP kinase kinase?

A

MEK

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8
Q

What is another name for MAP kinase?

A

Erk

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9
Q

What is another name for Raf?

A

MAP kinase kinase kinase

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10
Q

What is another name for MEK?

A

MAP kinase kinase

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11
Q

What is another name for Erk?

A

MAP kinase

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12
Q

True or false: different aspects of NGF go through different parts of the pathway

A

True: these effects are mediated in different stages of the pathway

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13
Q

Which part of the Ras pathway is responsible for the action potential?

A

The receptor

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14
Q

Which part of the Ras pathway is responsible for survival?

A

The adaptors and Ras

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15
Q

Which part of the Ras pathway is responsible for cell division?

A

Ras

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16
Q

Which part of the Ras pathway is responsible for neurites?

A

Erk

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17
Q

Which part of the Ras pathway is responsible for neurotransmitter synthesis?

A

Erk

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18
Q

What is the effect of the receptor in the Ras pathway?

A

The action potential

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19
Q

What is the effect of the adaptors in the Ras pathway?

A

Survival

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20
Q

What is the effect of Ras in the Ras pathway?

A

Cell survival and division

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21
Q

What is the effect of Erk in the Ras pathway?

A

Neurotransmitter synthesis and neurites

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22
Q

How can the steps of the Ras pathway be tested to tease out their effects?

A

Through the use of activated/inhibited forms for each step of the pathway

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23
Q

What is one of the alternative pathways found at the same p-Tyr of TrkA for NGF (not PI3K)?

A

Signaling through the Rap/B-raf pathway (which goes into the Erk pathway)

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24
Q

What is one of the alternative pathways found at the same p-Tyr of TrkA for NGF (not B-raf)?

A

Signaling through PI3K and Akt

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25
Q

What determines which signaling pathway will be activated by a p-Tyr?

A

Preferential binding and competition

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26
Q

What is one alternative pathway found at the different p-Tyr of TrkA for NGF?

A

The PLC-gamma / PKC pathway

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27
Q

True or false: gene expression is constant over time

A

False: there are early genes and late genes

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28
Q

Generally, what do early genes do?

A

Help transcribe later genes

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29
Q

True or false: gene transcription can cluster together over time

A

True: they can travel and cluster together over time for temporal resolution

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30
Q

True or false: there are functional effects to the timing of gene regulation

A

True: they can lead to varied phenotypes happening at precise times

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31
Q

What happens when NGF is added to PC12 cells?

A

They get neuronal like processes

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32
Q

What happens when EGF is added to PC12 cells?

A

They undergo rapid division

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33
Q

What types of changes does NGF lead to?

A

Long term changes

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34
Q

What type of changes does EGF lead to?

A

Transient changes

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35
Q

Which growth factor leads to long term changes?

A

NGF

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36
Q

Which growth factor leads to transient changes?

A

EGF

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37
Q

True or false: NGF and EGF signal through very different pathways

A

False: both go through a very similar Ras pathway

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38
Q

What does NGF and EGF have in common?

A

They both signal through a very similar Ras pathway

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39
Q

What happens to MEK and ERK under NGF activation?

A

MEK and ERK activation are sustained

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40
Q

What happens to MEK and ERK under EGF activation

A

MEK and ERK activation occurs very quickly in time

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41
Q

Why does MEK activation occur ERK activation?

A

Because MEK is upstream of ERK

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42
Q

Which occurs first: MEK activation or ERK activation?

A

MEK activation

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43
Q

Where does MEK/ERK go under NGF activation?

A

They translocate to the nucleus

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44
Q

How much time is needed for MEK/ERK to translocate to the nucleus?

A

> 15-20 min

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45
Q

What is the limit of EGF signaling?

A

15-20 min

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46
Q

How come EGF signaling cannot lead to MEK/ERK translocation into the nucleus?

A

The limit of EGF signaling is 15-20 min, which is the time needed for MEK/ERK translocation into the nucleus

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47
Q

What pathway allows for sustained signaling by NGF?

A

The Frs2/B-raf pathway

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48
Q

What does the Frs2/B-raf pathway do?

A

Allows for sustained signaling of MEK/ERK by NGF

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49
Q

What pathway allows for transient signaling by EGF?

A

The Ras/Raf pathway

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50
Q

What happens if B-raf is knocked out?

A

There will only be transient signaling

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51
Q

How come Frs2 doesn’t recognize the p-Tyr in EGFR?

A

There is a slightly different structure, which prevents binding of Frs2 to EGFR

52
Q

How is EGFR down-regulated?

A

Receptor-mediated endocytosis via clathrin coated pits

53
Q

What does clathrin do?

A

Aids in the formation of endocytotic pits

54
Q

What happens to EGFR endosomes?

A

They get degraded in lysosomes

55
Q

What is the structure of the clathrin coated pits (before forming the endosome)?

A

Similar to a soccer ball

56
Q

What is a clathrin triskelion?

A

An individual clathrin molecule that can aggregate to form the clathrin coated pit

57
Q

What does dynamin do?

A

Pinches off the clathrin coated pit

58
Q

What molecule pinches off the clathrin coated pit?

A

Dynamin

59
Q

How does dyanmin pinch off the clathrin coated pit?

A

Through GTP hydrolysis

60
Q

What type of molecules are Rab proteins?

A

Small GTP binding proteins

61
Q

What do Rabs do?

A

Determine which endosome pathway (lysosome vs MVB is preferred)

62
Q

What does MVB stand for?

A

Multivesicular body

63
Q

What type of endocytosis does EGFR go through?

A

Clathrin dependent endocytosis

64
Q

What type of endocytosis does Trk go through?

A

Clathrin independent endocytosis

65
Q

Where is Ras found in the cell?

A

At the cell surface

66
Q

Where is Rap found in the cell?

A

At the endosomes

67
Q

Where is B-raf found in the cell?

A

At the endosomes

68
Q

What does Rab7 do?

A

Brings endosome into the lysosome

69
Q

Would Rab7 be expected in EGFR endosomes or Trk endosomes?

A

EGFR endosomes, since it signals for degradation

70
Q

How does the MVB of Trk lead to sustained signaling?

A

Through stabilization and retrograde transport to the cell body

71
Q

What does Rab5 and Rab1 do?

A

Lead to sustained signaling of the MVB

72
Q

Would Rab5 and Rab1 be expected to remain in EGFR endosomes or Trk endosomes?

A

Trk endosomes, since it signals for sustained activation of ERK

73
Q

What is the signaling endosome hypothesis?

A

Endosomes were retrogradely transported to the cell body for signaling

74
Q

True or false: the signaling endosome hypothesis is fairly old

A

False: it is a fairly recent hypothesis

75
Q

Before the signaling endosome hypothesis, what was the prevailing function of endosomes?

A

Degradation and recycling (not signaling)

76
Q

How does NGF binding to TrkA in the axon lead to sustained ERK signaling in the nucleus?

A

The endosome is transported from the axon to the cell body

77
Q

What happens when endocytosis of EGFR is blocked?

A

The cell will experience prolonged EGF signaling

78
Q

How can endocytosis of EGFR be blocked?

A

By blocking dynamin

79
Q

How come blocking dynamin prolongs the EGF response?

A

The receptor stays at the cell surface, and is not down regulated by endocytosis

80
Q

What is the shape of PC12 cells?

A

Small round tumor cell balls

81
Q

What is the advantage of a signaling endosome?

A

No longer dependent on ligand binding at the cell surface for signaling, allowing for more efficient long-term signaling

82
Q

True or false: signaling endosomes allow for more efficient long-term signaling

A

True: they no longer depend on the ligand being constantly presented at the cell surface

83
Q

How does a signaling endosome increase spatial resolution?

A

It is a mobile signaling unit

84
Q

Why is a signaling endosome used over a random walk?

A

A signaling endosome takes significantly less time to reach the nucleus compared to a random walk

85
Q

What does Huntingtin do?

A

Mediates the retrograde transport of BDNF-TrkB signaling endosomes in dendrites of striatal neurons

86
Q

What is the effect of Huntingtin mutation (as seen in Huntingtin’s disease)?

A

It prevents retrograde transport of the BDNF-TrkB signaling endosome, leading to degradation of the striatal neurons

87
Q

How come neurons degenerate without the TrkB-BDNF signaling endosome?

A

No more sustained survival signaling

88
Q

How are in vivo studies hard to manipulate?

A

In terms of the context of other cells

89
Q

How are in vitro studies hard to manipulate?

A

In terms of space (no orientation, random processes)

90
Q

What is a Campenot chamber culture?

A

A device with a greasy teflon divider, and scratches of collagen in the middle

91
Q

What is the purpose of a Campenot chamber culture?

A

To separate the cell body from the dendritic and axonal compartments

92
Q

What is the purpose of the collagen scratches in the Campenot chamber culture?

A

Place to grow the cell

93
Q

What is the purpose of the greasy teflon divier in the Campenot chamber culture?

A

Prevent material in one compartment to diffuse into another compartment

94
Q

In a Campenot chamber culture, if a substrate was placed in the distal neurite area, and is found in the cell body, what conclusion can you make?

A

The substrate was endocytoses, and moved retrogradely along the neurite back towards the cell body

95
Q

What is the more recent, sophisticated form of a Campenot chamber culture?

A

Microfluidic chambers

96
Q

How do microfluidic chambers work?

A

They maintain a pressure difference to prevent material from diffusing between different compartments

97
Q

In a microfluidic chamber, what ensures that the media given to the distal neurites will not diffuse to the cell body?

A

A pressure gradient prevents the diffusion of the media (driving force in one direction)

98
Q

What is dynein?

A

A motor protein that moves vesicles from the periphery to the cell body

99
Q

What happens if dynein is blocked, and NGF is added to the distal neurites?

A

The cell dies, because it did not receive those survival signals

100
Q

How come blocking dynein causes neuron death, even if NGF is added to the distal neurites?

A

Dynein is needed to transport the NGF-TrkA endosome back to the cell body for sustained ERK / survival signaling

101
Q

How can middle axons be examined in a microfluidic system?

A

By placing “dividers” between the cell bodies, the middle axons, and the distal axons

102
Q

How can endosomes be tracked in the cell?

A

By using domain swapping

103
Q

How does domain swapping work (to track the endosome)?

A

Can stimulate it with one ligand, and see where the endosome (based on the intracellular domain) goes

104
Q

What is QDot605?

A

A fluorescent tag that can be added to EGF

105
Q

What happens when neurons are cultured with EGFR and EGF in the microfluidic chamber?

A

They die (no sustained ERK signaling)

106
Q

What happens when neurons are cultured with EGFR/TrkA chimeric receptor and EGF in the microfluidic chamber?

A

They survive (sustained ERK signaling)

107
Q

In the microfluidic chamber, where is ETrkB expressed?

A

All over the cell body

108
Q

In the microfluidic chamber, where is qEGF expressed?

A

In specific portions of the cell body

109
Q

In the microfluidic chamber, where is pERK expressed?

A

At the places where qEGF is expressed

110
Q

Why is ETrkB expressed all over the cell body in the microfluidic chamber?

A

The adenovirus inserts the gene, so it is expressed everywhere

111
Q

Why is pERK expressed only where qEGF is expressed?

A

The receptor bound to the ligand is needed to lead to pERK signaling

112
Q

Where did the qEGF and pERK come from in the microfluidic chamber?

A

Retrogradely transported signaling endosomes

113
Q

What is the conclusion of the microfluidic chamber experiments with qEGF?

A

The Trk receptor needs to bind to a ligand and be retrogradely transported back to the cell body for sustained ERK signaling

114
Q

After 2 hours, what happened to the qEGF/pERK endosomes in the microfluidic chamber?

A

They formed multivesicular bodies

115
Q

True or false: the EGFR endosome is not retrogradely transported

A

False: it is also retrogradely transported like the Trk endosome

116
Q

Where are EGFR endosomes retrogradely transported to?

A

Lysosomes

117
Q

Where are Trk endosomes retrogradely transported to?

A

MVBs (not lysosomes)

118
Q

What does MVB stand for?

A

Multivesicular body

119
Q

What do Trk endosome MVBs also contain?

A

Rab5

120
Q

What type of protein is Rab5?

A

A small GTPase

121
Q

What does Rab5 do?

A

Help determine where in the endosomal pathway a particular endosome will go

122
Q

What are the spatial effects of neurotrophin signaling?

A

They can signal at different places for different effects (dendritic branching, neurite growth, etc.)

123
Q

How can NGF endosomes be used in somal signaling?

A

They can phosphorylate soma derived TrkA receptors, which signals for them to be transported to the gowth cone

124
Q

What is the signal for soma derived TrkA receptors to be stimulated?

A

Phosphorylation by NGF endosomes

125
Q

What is kinesin?

A

A motor protein that moves endosomes from the cell body to the periphery

126
Q

Which motor protein aids in anterograde transport?

A

Kinesin

127
Q

Which motor protein aids in retrograde transport?

A

Dynein

BIO 317 (27 decks)

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