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MIMM 314 > Lecture 8 - T Cell Development 1 > Flashcards

Lecture 8 - T Cell Development 1 Flashcards

1
Q

what do immune cells arise from?

A

multi-potent hematopoietic stem cells

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2
Q

describe the progression from mutli-potent hematopoietic stem cells to specific immune cells

A

series of differentiation steps with increasingly restricted differentiation potential to make specific cells

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3
Q

what happened when adult mice had their thymus removed?

A

nothing

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4
Q

what happened when neonatal mice had their thymus removed?

A

in sterile environment –> nothing

in non-sterile environment –> died from infection

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5
Q

what did they know about the thymus and lymphocytes before the discovery of thymus?

A

knew that lymphocytes existed but didn’t realize there were T and B cells

initially thought thymus was “graveyard” for lymphocytes

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6
Q

describe the difference of T cells in mice and humans in terms of when mature T cells can be detected

A

mature T cells can be found at birth in mice

mature T cells can be found at 14-15 weeks of pregnancy in humans

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7
Q

what would happen if you remove the thymus from a human at birth?

A

they would be fine –> since mature T cells are there by 14-15 weeks of pregnancy, the baby would have T cells

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8
Q

what are the intermediate cells in T cell development?

A

DN1 –> DN2 –> DN3 –> DN4 –> DP –> SP

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9
Q

describe a flow cytometry plot of CD4 vs CD8 for T cell precursors

A

looks like a bird

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10
Q

how can we gate the DN population of cells to look at DN1 vs DN2 vs DN3 vs DN4

A

Use CD44 vs CD25 to look at specific DN populations

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11
Q

what are the 6 general steps for T cell maturation?

A
  1. Progenitor seeding of the thymus
  2. T cell lineage commitment
  3. B-selection
  4. Positive selection
  5. CD4 vs CD8 cell fate choice
  6. Impact of basal self-reactivity on T cell function
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12
Q

what cells seed the thymus? where do they come from in fetal vs adult mice?

A

multi-potent HSCs seed the thymus

  • in fetal mice from liver
  • in adult mice from BM
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13
Q

describe T cells in fetus vs adult mouse

A

fetus: invariant with less diverse Ag receptor

adult: more specific and very diverse T cell population

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14
Q

why are fetal T cells more invariant and less specific?

A

they don’t have fully functioning immune system so fast-acting, non-specific cells (more immune-like) are better to get rid of infection

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15
Q

what allows for the different types of T cells in fetal vs adult mice?

A

Lin28 and let-7 interaction

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16
Q

where is Lin28 expressed?

A

Lin28 is expressed in HSC progenitors in fetal liver

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17
Q

what is the role of Lin28?

A

Lin28 is an RNA-binding protein that inhibits the maturation of the let-7 miRNA

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18
Q

Let-7 levels in fetal vs adult mice

A

fetus has Lin28 so low let-7 levels

adult has no Lin28 so high let-7 levels

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19
Q

what happens when there’s forced expression of Lin28 in adult bone marrow?

A

it is sufficient to cause differentiation of SOME fetal T cell subtypes

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20
Q

what happens when fetal progenitors are transferred into an adult thymus?

A

can support SOME fetal T cell subtypes

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21
Q

what’s the next step once progenitors have been seeded in the thymus?

A

commitment to the T cell lineage

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22
Q

what is known as the “master regulator” of T cell fate?

A

Notch

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23
Q

4 Notch receptors and 5 Notch ligands

A

RECEPTORS:
- Notch1
- Notch2
- Notch3
- Notch4

LIGANDS:
- Jagged 1
- Jagged 2
- Delta 1
- Delta 4
- Delta 3

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24
Q

how does Notch signaling work? (3 steps)

A
  1. Thymic epithelial cell has notch ligand which binds notch receptor on thymocyte
  2. Interaction forces intracellular domain of notch to be released
  3. drives T cell fate
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25
Q

4 main functions of Notch signaling

A
  1. cell fate choice
  2. cell survival
  3. proliferation
  4. stem cell maintenance
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26
Q

Notch is _______ and ________ to determine T cell fate

A

Notch is NECESSARY and SUFFICIENT to determine T cell fate

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27
Q

what happens with GAIN OF FUNCTION NOTCH in lymphoid progenitor cell?

A

lymphoid progenitor cell makes T cells but NOT B cells

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28
Q

what happens with INACTIVATION OF NOTCH in lymphoid progenitor cell? why?

A

lymphoid progenitor cell makes B cells but NOT T cells because notch is necessary

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29
Q

what is the difference between the notch ligands?

A

most are redundant with the same roles

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30
Q

what is the main notch ligand in the thymus? what cells is it expressed on?

A

delta 4

found on cortical thymic epithelial cells

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31
Q

what happens to multipotent progenitor cells that become bone marrow cells without notch ligand?

A

B cells are produced

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32
Q

what happens to multipotent progenitor cells that become bone marrow cells with notch ligand?

A

T lineage cells are produced

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33
Q

progenitor cells that seed the thymus are ______

A

progenitor cells that seed the thymus are MULTIPOTENT

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34
Q

what does it mean for progenitor cells that seed the thymus to be multipotent?

A

make T or B cells depending on the signal

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35
Q

describe production of B and NK cells from DN1 vs DN2 vs DN3 cells with or without notch

A

with notch:
- cannot make B cells!
- can make some NK cells but less as progenitor cells are more differentiated

without notch:
- can make B and NK cells
- but lose ability to make either B or NK cells as progenitor cells are more differentiated

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36
Q

describe production of DP T cells from DN1 vs DN2 vs DN3 cells with or without notch

A

T cells can only be made when notch is present

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37
Q

what is a way that therapeutic T lineage cells can be made? what is the issue with this?

A

can use a bead with notch ligand on its surface to interact with notch receptor on hematopoietic progenitor

easy to reach the DP stages but hard to reach mature T cells

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38
Q

what happens once the progenitor cells have committed to the T lineage?

A

Beta selection! make beta chain for TCR

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39
Q

at what intermediate cell does TCRbeta chain rearrangement begin?

A

DN2

40
Q

how does TCRbeta chain rearrangement work?

A

select 1 V, D, J, and C to make unique combination

41
Q

what gene causes TCR gene rearrangement?

A

recombination activating genes (RAG) 1 and 2

42
Q

what allows for the most TCR diversity?

A

terminal deoxynucleotidyl transferase (TdT) adds non-template nucleotides at junctions of the gene segments

43
Q

what percent of TCR diversity is due to TdT?

A

95%

44
Q

is TCRalpha made at the same time as TCRbeta?

A

no

45
Q

what is the preTCR?

A

preTCR = rearranged TCRbeta chain paired with invariant preTCRalpha chain

46
Q

3 things that a functional TCRbeta chain leads to:

A
  1. transient downregulation of RAG, allelic exclusion
  2. proliferation
  3. differentiation
47
Q

why is RAG downregulated?

A

so that only 1 beta chain is rearranged

48
Q

what are the 2 hypotheses for how beta-selection works?

A

either ligand DEPENDENT or ligand INDEPENDENT

49
Q

what does it mean for beta selection to be ligand independent?

A

only requires rearranged beta chain for beta selection

50
Q

evidence that preTCR signaling/beta-selection is ligand independent (2)

A
  1. delete MHC, differentiation from DN to DP is unaffected
  2. delete extracellular domain of preTCR (where ligand would bind), differentiation from DN to DP is unaffected
51
Q

evidence that preTCR signaling/beta-selection is ligand dependent (3)

A

preTCR can bind to a peptide presented by MHC I and an absence of this interaction leads to:
1. decreased beta-selection efficiency
2. decreased TCR repertoire diversity
3. abnormal thymocytes

52
Q

describe the binding of preTCR to peptide presented by MHC I and what this allows for

A

binds in diff conformation than mature TCR which means more beta chains can bind less specifically to peptide presented by MHC I

53
Q

which of the following plots of thymocytes would we get if there was no functional RAG? why?

A

C

without RAG, there will be no differentiation to DP

54
Q

what happens after beta selection?

A

positive selection

55
Q

what happens during positive selection?

A

DN4 becomes DP as TCR is sampling environment

56
Q

when does TCRalpha chain rearrangement occur?

A

at DP stage

57
Q

describe TCRalpha chain rearrangement

A

only V and J are rearranged

58
Q

does TCRbeta chain rearrangement occur during TCRalpha chain rearrangement?

A

no!! that has been completed

59
Q

what are the 3 outcomes for thymocytes with TCRs during positive selection

A
  1. death by neglect –> apoptosis
  2. positive selection –> survival and differentiation
  3. negative selection –> apoptosis
60
Q

what causes death by neglect?

A

TCR receives no signal from any peptides therefore will never be able to bind peptide

61
Q

what causes positive selection?

A

TCR binds with low affinity to a SELF PEPTIDE

62
Q

what causes negative selection?

A

TCR binds too strongly to self peptide, therefore too much self-reactivity

63
Q

are T cells self-reactive? why?

A

yes, all T cells are self-reactive

bc the low affinity interaction with self peptide is what allows for positive selection

64
Q

how many different TCR heterodimers are possible?

A

10^20

65
Q

what percent of DP thymocytes can pass positive selection?

A

only 5-10%

66
Q

describe positive selection and MHC restriction

A

positive selection ensures that the TCR has dual specificity for peptide + MHC (MHC restriction)

67
Q

what is MHC restriction?

A

TCR can only recognize peptide presented by self-MHC and will NOT recognize the same peptide presented by another MHC

68
Q

describe bone marrow chimera

A

mouse is irradiated and BM cells are killed –> donor BM cells from another mice are added to the irradiated mouse

69
Q

what cells are radiation resistant?

what cells are not radiation resistant?

A

thymic epithelial cells ARE radiation resistant

hematopoietic cells (BM cells) are NOT radiation resistant

70
Q

what happens to lymphocyte generation when WT mouse is donor for irradiated WT mouse?

A

positive selection can occur

71
Q

what happens to lymphocyte generation when MHCnull mouse is donor for irradiated MHCnull mouse?

A

no MHC = no positive selection = no CD4+/CD8+ DP cells

72
Q

what happens to lymphocyte generation when WT mouse is donor for irradiated MHCnull mouse?

A

no positive selection because the thymic epithelial cells don’t have MHC

73
Q

what does the WT –> MHCnull bone chimera tell us?

A

positive selection requires MHC expression in thymic epithelial cells

74
Q

specifically, what type of thymic epithelial cells are required for positive selection?

A

cortical thymic epithelial cells

75
Q

what allows cortical thymic epithelial cells to induce positive selection?

A

THYMOPROTEASOME

76
Q

what is the thymoproteasome?

A

beta5t proteasome subunit encoded by Psmb11 gene in cortical thymic epithelial cells allows for peptide fragments to be made for positive selection

77
Q

specifically, what is the thymoproteasome required for?

A

required for CD8+ T cell development to present peptides on MHC I

78
Q

what happens in beta5t KO? significance

A

reduced CD8+ cells –> therefore required for positive selection

79
Q

why are the peptides made by the thymoproteasome unique?

A

they bind to MHC I with lower affinity than normal peptides so they can fall off more easily –> this helps with the low affinity interactions with self-peptides

80
Q

what happens after positive selection?

A

CD4 vs CD8 T cell lineage choice

81
Q

what are the 2 previous models of CD4 vs CD8 T cell lineage choice?

A
  1. instructive model
  2. stochastic model
82
Q

describe the instructive model

A

when TCR on DP binds peptide on MHC, the QUALITY and QUANTITY of the signal instructs the DP to downregulate CD4 or CD8

ex. if there’s stronger signal when TCR binds MHC II with CD4, the cell knows to downregulate CD8

83
Q

describe the stochastic model

A

when TCR on DP binds peptide on MHC, one of the co-receptors is randomly downregulated, then there’s a second step to ensure the TCR specificity and remaining co-receptor match

84
Q

what is the current model of CD4 vs CD8 T cell lineage choice?

A

kinetic model

85
Q

describe the kinetic model

A

when TCR on DP binds peptide on MHC, CD8 is downregulated and depending on whether the signal continues or is interrupted will instruct whether it is CD4 or CD8 involved in TCR binding

if signal persists –> CD4
if signal is lost –> CD8

86
Q

what is the master regulator of CD4 vs CD8 T cell lineage choice?

A

ThPOK

87
Q

what happens with ThPOK overexpression in DP cells?

A

only CD4 cells are produced

88
Q

what happens with ThPOK KO in DP cells?

A

only CD8 cells are produced

89
Q

ThPOK is ________ and ________ to make ___ T cells

A

ThPOK is NECESSARY and SUFFICIENT to make CD4 T cells

90
Q

what happens with ThPOK if TCR signal is persistent with CD8 downregulation?

A

if there is a persistent signal in cells with MHC II when CD8 is downregulated, ThPOK is turned on leading to:
- expression of CD4-specific genes
- inhibition of cytokines for CD8 development

91
Q

if MHC II expression is restricted to DCs, what type of T cells will be found in the thymus?

A

Cortical thymic epithelial cells require MHC I and MHC II to make CD4 and CD8 –> so if MHC II is only on DCs, will only make CD8!

92
Q

what happens once T cells become single positive?

A

thymic egress

93
Q

are T cells finished maturing once they leave the thymus?

A

no, they continue differentiating until they become fully mature T cells

94
Q

what are T cells called once they leave the thymus but before they become mature T cells?

A

recent thymic emigrants

95
Q

where do recent thymic emigrants mature?

A

in peripheral lymphoid organs

MIMM 314 (23 decks)

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