8 - T cell Development Flashcards
Main points
Early phase:
1) commitment of hematopoietic precursors to the T cell lineage
2) initiation of antigen receptor gene rearrangement
3) expansion of cells that have successfully rearranged one of their T cell receptor genes
Second phase:
1) Positive selection (selection for those cells whose TCR engage self-MHC)
2) negative selection (selection against those cells whose TCR binds too strongly to self-peptide/MHC combinations)
3) Lineage commitment: commitment of thymocytes to effector cell lineages, including CD4+ helper or CD8+ cytotoxic populations, as well ad CD4+ regulatory T cells
Early development overview
begins with the arrival of small numbers of immature lymphocyte precorsors from the bone marrow, thorugh the blood, and into thymus. Here, they proliferate, differentiate, and undergo selection processes that reduce their numbers, generating CD4+ and CD8+ T cells.
the thymocyte precursors (aka thymus-settling progenitors, TSPs) are directed to the thymus via chemokine receptors. Upon arrival, they are still able to differentiate into B, NK, DC, and some other myeloid cells. It is only at the late DN2 stage of development that they become fully commited.
The commitment is dependent on Notch, a receptor that regulates several desicions during TCD (T cell development), including the early choice of whether to be a T or B cell. If a constitutively actove version of Notch1 is overexpressed in hematopoietic stem cells, T cells develop. If not really expressed, B cells. This happens in the bone marrow.
T cells movement through the thymus
T cells arrive in the thymus in the coticomedullary junction, where they encountre Notch ligands (abundantly expressed in the thymic epithelium).
The T cell precursors then travel to the outer thymic cortex, where they proliferate and begin to express their TCR.
T cells that survive the testing here mature and go to the thymic medulla, before exiting through the corticomedullary junction.
Thymocytes progress htrough 4 DN stages
thymocytes mature in thymus (1-3 weeks). Pass through several stages
The earliest T cells lack detectable CD4 and CD8, and are therefore referred to as DN. These can be further subdivided into 4 subsets based on the presence or absence of other cell surface molecules, including
- cKit (CD117), the receptor for stem cell growth factor
- CD44, an adhesion molecule
- CD25, the α chain of the IL-2 receptor
DN1: c-Kit ++ CD44+ CD25- (ja, dobbel pluss). Still multipotent, but recieve Notch signals immediatly upon entering thymic enviroment, beginning to restrict them to T cell lineage.
DN2: c-Kit++ CD44+ CD25+. the genes for γ, δ and β chains begin to rearrange. the TCR α chain locus however, remains inaccessible to the recombinase machinery for now. at the late DN2 stage, the T cell precursors fully commit, and reduce exression of both c-Kit and CD44
DN3 : c-Kit+, CD44-, CD25+. during transition from DN2 to DN3, the TCR βγδ chains continue to rearrange, and make the first major desicion in T cell development: whether to join the αβ or γδ lineage. Those DN3s that successfully rearrange their β chain typically commit to the αβ lineage. They lose expression of CD25, halt proliferation, and enter the DN4 stage
DN4: (c-Kit low/-, CD44-, CD25-), mature directly to CD4+ CD8+ DP thymocytes.
Thymocytes express either αβ or γδ receptors
αβ are the dominant participators in the adaptive immune response in secondary lymphoid organs. γδ also play an important role, particularly in protecting barrier tissues from outside infection.
The coice of whether αβ or γδ develops is largely due to how quickly the genes that encode each of the four receptor chains successfully rearrange.
rearrangement of βγδ begins during DN2. to become a αβ, the cell must generate a β protein chain (single in-frame V(D)J rearrangement, becoming γδ takes two such rearrangements. Probability therefore favors the former. 3x more likely.
most γδ do not go through the DP stage, but leaves the thymus as a DN. many emerge from the thymus with the ability to secrete cytokines, which αβ only can do after they encounter Ag. γδ express receptors that are not as diverse, and they seem to recognize unconventional MHC molecules
DN thymocytes undergo β selection, which results in proliferation and differentiation
β selection = identification and expandation of DN thymocytes with successful β rearrangement. The process involves the pre-Tα chain (protein uniquely expressed at this stage). this acts like a surrogate for the α chain (yet to rearrange) and assembles with the β chain + CD3 complex. This total complex = pre-TCR, and act like a sensor, initiating a transduction pathway, triggering the following events:
1) maturation to the DN4 stage
2) rapid proliferation in the subcapsular cortex of the thymus
3) suppression of further rearrangement of TCR β chain genes, resulting in allelic exclusion of the β chain locus
4) development to the DP stage
5) cessation of proliferation
6) initiation of TCR α chain rearrangement
Note that the proliferative phase enhances receptor diversity considerably by generating clones of cells with the same TCR β arrangement. Each of these cells can then rearrange a different α chain. the α chain rearrangement does not happen until the DP stop proliferating.
allelic exclusion = once one allele has a successful β chain rearrangement, rearrangement of the other allele is shut down.
Once a successful α chain has been produced, it takes the place of the surrogate pre-TCR
Positive and negative selection
MHC restriction = T cells only rec antigen peptides in complex with self-MHC.
self-tolerance = T cells have some affinity for self-MHC, but will not attack self-MHC/self-peptide complexes.
Most T cells are eliminated by failing positive selection.
Affinity model: DP thymocytes have one of three fates depending on the affinity of theri TCR for self-MHC/self-peptide complexes expressed on thymic stromal cells.
ALthough the affinity model does not explain all aspects of negative and positive selection, it remains a useful starting point for understanding these processes.
Negative selection can occur both in cortex and medulla, and is mediated by multiple cell types in these tissue compartments. Positive selection occurs in the cortex. Positively selected thymocytes up-regulate the CCR7 chemokine receptor, which allows them to migrate from the thymic cortex to the thymic medulla. Semimature thymocytes, in transition from the DP to the SP stage, can then be negatively selected for tissue-specific antigens presented by medullaty epithelial cells.
Positive selection
in the thymic cortex, newly generated DPs first browse the surfaces of cortical thymic epithelial cells (cTECs), which are unique in their ability to mediate positive selection. If the TCR of the DP rec the MHC-peptide (all self) complex on the cTEC, it will undergo positive selection (induces survival and differentiation of DPs). a maturation program is triggered, and it sends the cells to medulla and initiates their commitment to either CD4+ or CD8+ SP lineages. Positive selection requires the thymocyte specific signaling molecule, Themis.
The cells that do not engage the complexes undergo death by neglection.
negative selection (central tolerance) ensures self-tolerance
most neg selection occurs via clonal deletion, where high-affinity TCR interactions directly induce apoptosis. clonal deletion is mediated by a variety of cells in the thymus. The same interactions that activeta meture T cells are known ot induce apoptosis in immature DP T cells. thymic DC and macrophages are ideal for negative selection mediation, as are medullary thymic epithelial cells (mTECs), which express high levels of the costimulatory ligands CD80 and CD86. mTECs express AIRE, a unique TF that allows mTECs to express, process and present proteins that are ordinarily found in other tissues (like insulin, f.ex).
The selection paradox: why don’t we delete all cells we positively select?
Two models: affinity and altered peptide models. Not mutually exclusive.
according to the affinity model for selection:
the differences in the strength of TCR mediated signals recieved by thymocytes undergoing positive and negative selection determine the outcome of the interaction. DP thymocytes that recive low-affinity signals are positively selected, whereas those who recieve high-affinity (autoreactive) signals are negatively selected. No signal? die by neglect.
altered peptide model:
suggests that the cTECs (which induce positive selection) make peptides that are unique and distinct from peptides made by thymic cells that induce negative selection. Thus, those thymocytes selected on these unique peptide/MHC complexes would not automatically be negatively selected when they browse the medulla and other negatively selecting cells.
cTECs do indeed process peptides differently, and present a different array of peptides to developing T cells. The proteasome ecpressed by cTECs (thymoproteasome) has a unique component, and cTECs also express a unique version of the cathepsin protease. These features allow cTECs to produe a population of peptides no other cells can generate.
Lineage commitment
lineage commitment requires changes in genomic organization and gene expression that result in
1) silencing of one coreceptor gene (CD4 or CD8)
2) expression of genes associted with a specific lineage function.
Several models:
1) instructive model
TCR/CD4 and TCR/CD8 co-engagement generate unique signals that directly initiate distinct developmental programs. if a thymocyte randomly generates a TCR with affinity for MHC I molecules, the TCR and CD8 would bind a MHC I together, and generate a signal that specifically initiated a program that silenced CD4 expression, and induced expression of genes specific for cytotoxic T cell lineage functions.
2) stochastic model
a positively selected thymocyte randomly down-regulated CD8 or CD4.
3) strength of signal model
the strength and dureation of the TCR/coreceptor signal experienced in a thymocyte play a more important role in dictating cell fate than the specificity of either MHC I or II.
4) kinetic signaling model (most probable)
proposes that thymocytes commit to the CD4+ lineage if they recieve a continous signal in response to TCR/coreceptor engegament, but commit to the CD8 lineage if the TCR signal is interrupted.
We know that DP cells downregulate CD8 in response to positive selection. Given this response, only MHC II-restricted T cells will maintain continous TCR/CD4/MHC II interaction, and therefore develop CD4 lineage. However, with the loss of CD8 expressoin, MHC I-restricted T cels will lose the ability to maintain TCR/CD8/MHC I interactions.
TFs Th-POK and Runx3 play key roles in CD4+ helper cell and CD8+ cytotoxic cell commmitment, respectively. They work in part by reciprocally regulating each other. They both supress genes involved in differentiation to the other lineage.
DP thymocytes may commit to other types of lymphocytes
for a small percentage of thymocytes, high-affinity TCR interactions do not lead to deletion, but instead drive development to specialized cell lineages, including NKT cells, IELs (intraepithelial lymphocyte) , and regulatory CD4+ T cells.
Exit from the thymus and final maturation
maturing thymocytes initiate a cellular program that enhances their survival and up-regulates receotirs that facilitate their migration from the thymus and through the blood.
thymocytes deficient for the CCR7 chemokine receptor failed to enter the medulla, but they could still leave the thymus directly from the cortex, suggesting that at least one other receptor controlls thymus exit. This receptor is S1P1 (sphingosine 1-phosphate receptor 1).
The exit of SP thymocytes from the thymus depends on the expression of the S1P1 and its interaction with S1P at the corticomedullary junction.
Mature thymocytes that have just left the thymus are referred to as recent thymic emigrants. They do not yet function optimally, and undergo a post-thymic phase of maturation in secondary lymphoid tissue that fully licences them as naïve T cells.
T regulatory cells
Treg cells negatively regulate immune responses by inhibiting the proliferation of other T cell populations both directly and indirectly. They express surface CD4 as well as CD25, the α chain of the IL-2 receptor, and FoxP3 (master trc regulator, expression of this is necessary and sufficient to induce differentiation to the Treg lineage).
Most thymocytes failing negative selection dies, but some become Tregs, and leave the thymus to patrol the body and thwart autoimmune reactions. not fully understood what determines if they die or become Treg. Treg precursors + IL-10 and IL-15 -> Treg
Tregs developed in thymus = tTregs.
Tregs developed from conventional mature T cells exposed to Ag in the context of TGF-β and IL-10 cytokines = pTregs (peripheral).
Exactly how Tregs quell immune responses is still debated, but they probably do so via a variety of means:
- directly inhibiting an APCs ability to activate T cells
- directly killing T cells
- indirectly inhibiting T cell activity by secreting inhibitory cytokines IL-10 and TGF-β
- depleting the local envorinment of stimulatory cytokines such as IL-2
Peripheral mechanisms of tolerance also protect against autoreactive thymocytes
autoreactiev T cells can escape from the thymus. many Ags are “hidden” from these T cells bc only a subset of pAPCs express the right costimulatory molecules needed to initiate the immune respnse.
peripheral tolerance enforce T cell tolerance in the petiphery. Tregs contribute to this tolerance, as does the strict requirement for costimulatory interactions in order to activate a T cell. Costimulation can be provided only by professional APCs, whose activity is highly regulated.
