Initially, in the hypothetical, on the basis of cell phenomenology, and then in an experimentally well-documented form using molecular biology methods, it was found that the T-cell receptor recognizes not actually a foreign antigen, but its complex with proteins controlled by the main histocompatibility complex (MNS).
Basic evidence of dual recognition: molecules I and II classes of MCS and associated antigenic peptide associated with them, R. Zinchernagel and P. Dodokhti were obtained. Studies of these scientists were noted by awarding the Nobel Prize for Medicine for 1997.
The structure of T-cell antigensic receptors
There are two types of T-cell antigensic receptors: TCRAB and TCRYB (from English - T Cell Receptor, TCR). The latter is expressed on the minor subpopulation of T-cells (TYB), which in a small amount are represented in thymus and on the periphery - in the spleen, blood. In ontogenesis, they precede T-cells with TCRAB (Tab). Thymus is not required to mature them, they are capable of self-reproduction, take part in antibacterial protection, reacting to carbohydrate components. Phylogenetically TYB preceded TAB.
Attempts to identify TCR with the help of anti-immunoglobulin antibodies, as was done in the search for antigensnial structures in B cells, turned out to be unsuccessful.
The identification of TCR managed only with the use of monoclonal antibodies (MAT) and cloned T-cell lines. Some clone-specific mat reacted only with clones isolated from pre-immunized animals. Making such clones in the culture of such clones corresponding to the specificity of the mat, suppressed the ability of cloned T-cells to recognize the antigen used for immunization. The presence of such antigensical mats provided a full-fledged study of antigensnial structures of T cells.
Each functionally mature T cell has about 3 104 TCR. They are a heterodimer built for most cells from OS- and (3-chains covalently interconnected by cysteine \u200b\u200bbridge. Each chain consists of a variable V-domain and a constant C-domain, homologous to the relevant immunoglobulin domains. The TCR structure also features a hinge Domain with cysteine \u200b\u200bresidue, which forms a disulfide bridge, combining A- and B-chaining a single molecule.
The TCR cell membrane is held with a hydrophobic transmembrane sequence of amino acid residues. A characteristic feature of a transmembrane domain is the presence of positively charged amino acid residues in it. Ends each chain with a short cytoplasmic tail, immersed in the cytoplasm. Available structural differences between TCRU BCR cannot be considered defining, since the main property is to build an active antigensnial area due to the processes of recombination and interaction of two V-domains - remains common.
Genetic control of the structure of the T-cell antigensnicational receptor
The organization of genes encoding the A- and B-chains of TCR is mainly homologous to the one that is known for the lungs and heavy chains of immunoglobulins. The V-domain of the OS-chain, like a light chain of immunoglobulins, is monitored only by V- and J-gene segments. At the same time, the formation of a V-domain B-chain, as well as a heavy immunoglobulin chain, is provided with a complete set of V-, D-, J-gene segments.
The T-cell genome has more than 100 V-genes for the TCR A-chain, which is two and a half times less than the amount that is known for the light chains of immunoglobulins.
Each such gene includes two exon - one for a leader (L) sequence that is absent in the mature A-chain, but presented in the parcel at the time of its transport from the endoplasmic reticulum to the cell surface, and the second - for encoding the TCR V-domain itself. The J-gene segments for the A-chain are significantly more than for a light chain of immunoglobulins (100 against 4). The constant area of \u200b\u200bthe A-chain is controlled by C-gene, comprising separate exons for C-domain and a hinge and one common exone - for transmembrane and tail parts of the molecule.
The number of V-genes for the B-chain is 30. In addition, there are two DJC clusters. Each cluster includes one D- and six active J-gene segments. Functional differences between clusters are unknown. The C-gene for the constant region of the B-chain includes four exon for constant, hinge, transmembrane and tail sections of the polypeptide. The processes of recombination, transcription, splicing and translation of the genetic material for A- and P-chains in the formation of TCR in T cells are similar to those providing synthesis of immunoglobulins in B cells.
As well as in the case of immunoglobulins and immunoglobulin receptors, the TCR variability depends on the random interaction of the gene segments in the process of recombination of the genetic material encoding V-domains: Vj - for A-chains and VDJ - for B-chains, as well as at the expense of those Additional changes, which, as in the case of BCR, accompany recombination. The exception is the lack of somatic mutagenesis in V-genes. The calculation of the variability of TCR V-domains, which is carried out in the same way as for immunoglobulins, shows an extremely high level of diversity of these antigensnuclear structures. Thus, only the presence in the genome of non-V-, D- and J-gene segments gives potentially (excluding modifications during reorganization) 2.8,106 options.
Immunoglobulins and immunoglobulin B-cell receptors recognize native antigenic epitopes. In this regard, the individual sections of the antigensnial center have equal chances of variability. The situation with TCR is somewhat different, since this receptor recognizes the complex of an antigenic peptide with MNC molecules.
A variety of TCR is associated to a large extent with a third loop of a V-domain formed by the third hypervariable site - CDR3 (from English - Compledenarity Determining Region). When the antigen-binding center CDR3 is found in its inner part. The first and second loops - CDR1 and CDR2 - occupy the periphery of the center respectively. In such conformational construct, there is a completely defined biological meaning associated with the adaptation of TCR to the form of the immunogen with which it interacts. Antigenic peptides fill the space (gap) formed by A-spiral structures of MMC molecules, and thus turn out to be in the middle of the antigenic complex peptide: MHC. Such a complex is characterized by a huge set of antigenic specificities associated with peptides, and limited diversity characteristic of MNS molecules. In connection with such an organization, an immunogenic complex should be expected to expect an increased variability of CDR3 and a smaller variability of CDR1 and CDR2. The study of the genetic organization of genes for TCR confirms a similar point of view. Thus, TCR has a significantly less compared to immunoglobulin, the number of V-genes defining the specificity of CDR1 and CDR2, but the increased number of J-CER-cops participating in coding CDR3 catering receptors and related proteins in the process of activation T cells
TCR, like membrane antigensnient immunoglobulin B cells, has a very short cytoplasmic tail. In this regard, the signal from the interaction of TCR with the peptide complex: MCS molecules cannot be transmitted inside the cell. Transmissive function is performed by invariant, low molecular weight associated with TCR proteins that have received a common name - CD3. The CD3 complex includes five proteins: CD3Y, CD3B and CD3E proteins are represented on cell surface and have aid homology with immunoglobulins, CD3 cytoplasmic proteins? And CD3N do not have such homology.
Proteins, homologous immunoglobulins, are expressed on cell surface in the form of CD3E6 and CD3EY heterodimers, their connection with TCR is carried out by means of electrostatic attraction. Negatively charged transmembrane sections of CD3 chains interact with the carrier's positive total charge of TCR transmembrane sections. The presence of a long tail allows them to interact with cytoplasmic proteins-transductors after the antigenic signal is obtained.
Two other polypeptide - CD3T and CD3N are also included in the complex in the form of SS dimers or CN-about 80% TCR associated with homodimer and only 20% with a heterodimeter. Functional differences between them are unknown. The main domain of these proteins, in contrast to other POPs, is in the cytoplasm. It is the head, and not the tail of C and n interacts in a cytoplasm with transductor proteins.
In addition to the signal-transmission function, the CD3 protein is responsible for TCR transport to the cell surface. In mutant cells, in which there is no synthesis of U-, B-or E-chains, the expression of TCR is completely suppressed, although the intracellular synthesis of these receptors is not disturbed. When mutations of the C-chain gene, the yield of TCR on the cellular surface occurs to a lesser extent compared to the norm. Both transussive and transport functions of CD3 proteins are homologous to that characteristic of IGA and IGP-proteins of the immunoglobulin, antigensnial complex.
In the activation of T-cells that recognized the antigen, the CD4 and CD8 CD8 dealers are also involved - T-cell differentiation markers. As already noted, the first of them is a marker of CD4 + T-cells, the second - cytotoxic T-lymphocytes (CD8 + T cells). For a long time, the function of these proteins remained unknown. It turned out that they take the most direct involvement in the process of interaction between TCR with the corresponding ligand as a coreceptor.
CD4 is a single-stranded molecule consisting of four immunoglobulin-like domains (Fig. 4.9). D1 and D2 domains, as well as d3 and d4 form pair, tight-packed, rigid structures. These pairs are connected by a flexible hinge section. The tail part of the CD4 molecule has sufficient length to interact with cytoplasmic transductor proteins. The TCR and CD4 cell surface are presented independently of each other. Their meeting occurs in the process of forming an antigen response. After the TCR recognition of the antigenic complex, CD4 with a molecule II class II occurs. The reaction of the interaction is carried out between the B2-domain of the MNS II molecule and the first CD4 domain. We also assume a weak inclusion in the interaction and second domain D2.
A similar pattern is observed in recognizing the antigenic complex with cytotoxic T-lymphocytes (CTL). The current participants in the interaction - TCR of cytotoxic T-lymphocytes, a complex of peptide with a molecule I class I and a marker of cytotoxic T-lymphocytes - CD8. CD8, although it performs the CD4 function of the coreboard, is structurally different from the T-helper marker. It is a heterodimer, each chain of which includes one immunoglobulin-like domain and a sufficiently long associated with a diaphragm section of a chain, which is subject to significant conformational change. As well as CD4, CD8 is represented on the cell membrane independently. Its function of the corescape is implemented in the process of antigenic recognition. After the interaction of TCR with antigenic ligand, the A- and P-domains of the CD8 with A3 domain of the IM CD molecule arrive. The resulting molecular complex is the condition of transmission through the CD8 CD8 cores inside the cell.
Intracellular events that determine the activation of T cells are similar to those that occur in cells after antigenic stimulation. The aggregate of antigen-representing molecules of MNS, the T-cell receptor complex comprising CD3 of the molecule, and CD4 or CD8 molecules provokes the intracellular interaction of various tyrosine kinases with the cytoplasmic part of the polypeptides. Among the CD3 proteins, the highest binding activity has the highest activity, which presented in the cytosole is not tail, and the head part. Activated as a result of the interaction of kinase provide a cascade of reactions, the consequence of the induction of specific gene transcriptions. Among the genes entered into the transcription process, they have a special place that encode the synthesis of T-dependent cytokines (in particular, IL-2). Ultimately, the chain of events from the interaction of TCR with an antigenic complex and the formation of a complex molecular unit to intracellular reaction transformations leads to proliferation.
And T cells have independent antigensic receptors belonging to the same superfamily immunoglobulin. Antigensnic C-cell receptors (BCR) are a monomer form of Igm, modified by an additional sequence of amino acid residues in the C-terminal part of the molecule. This sequence is a transmembrane and tail segment of a heavy chain. T-cell antigensnical receptor (TCR) consists of two polypeptide chains, each of which includes two domains: V and C. The mechanism of genetic control of V-domains both BCR and TCR is generally similar and includes the process of random recombination of gene segments (V, D, J). Despite the fact that the functional purpose of antigensic receptors of the two cell types is the same (alien recognition), the implementation of such a function of V- and T-cells is carried out in different ways. While SLG B cells recognizes the antigenic determinant itself without any additional conditions, TCR T cells can recognize only the complex of antigenic determinants with its own I or II Class II molecule.
T-cell receptors (eng. TCR) - surface protein complexes of T-lymphocytes responsible for recognizing processed antigens associated with the molecules of the main histocompatibility complex (eng. MHC) on the surface of antigen-representing cells. TCR consists of two subunits, borrowed in the cell membrane and are associated with a multi-suspension CD3 complex. The TCR interaction with MHC and the antigen associated with it leads to the activation of T-lymphocytes and is a key point in the launch of the immune response.
TCR is a heterodimeric protein consisting of two subunits - α and β or γ and δ, presented on the cell surface. Subunits are fixed in the membrane and are connected with each other disulfide bond.
According to its structure, the TCR subunit belongs to the superfamily of immunoglobulins. Each of the subunits is formed by two domains with a characteristic immunoglobulin laying, a transmembrane segment and a short cytoplasmic segment.
N-terminal domains are variable (V) and are responsible for binding an antigen presentable by molecules of the main histocompatibility complex. The composition of the variable domain contains a hypervariable portion (CDR) characteristic of immunoglobulins. Due to the extraordinary diversity of these areas, various T cells are able to recognize the widest range of various antigens.
The second domain is constant (c) and its structure is the same in all subunits of this type in a particular individual (with the exception of somatic mutations at the level of genes of any other proteins). On the plot between the C-domain and the transmembrane segment there is a substrate residue, with which a disulfide connection is formed between two TCR circuits.
The TCR subunits are aggregated with the CD3 membrane polypeptide complex. CD3 is formed by four types of polypeptides - γ, δ, ε and ζ. The subunits γ, δ and ε are encoded closely with the clutch genes and have a close structure. Each of them is formed by one constant immunoglobulin domain, transmembrane segment and long (up to 40 amino acid residues) by the cytoplasmic part. The chain ζ has a small extracellular domain, a transmembrane segment, and a large cytoplasmic domain. Sometimes instead of a chain ζ, the complex includes a chain η - a longer product of the same gene obtained by alternative splicing.
Since the structure of proteins of the CD3 complex is invariant (no variable sections), they are not able to determine the specificity of the receptor to the antigen. Recognition is exclusively a TCR function, and CD3 provides signal transmission into a cell.
The transmembrane segment of each of the CD3 subunits contains a negatively charged amino acid residue, and TCR is positively charged. Due to electrostatic interactions, they are combined into a common functional complex T-cell receptor. On the basis of stoichiometric studies and measurements of the molecular weight of the complex, the most likely composition is (αβ) 2 + γ + δ + ε2 + ζ2.
TCR consisting of αβ chains and γδ-chains are very close in structure. These forms of receptors are presented in different ways in various tissues of the body.
The structure of the T-lymphocyte receptor largely resembles the structure of the antibody molecule. T-cell receptor molecules (TCR) consist of two chains - a and r. Each of them contains V- and C-domains, their structure is fixed by disulfide bonds. Variable domains of A- and P-chains have not 3-4, like antibodies, and at least 7 hypervariable sites that form an active receptor center. For C-domains, near the membrane, there is a hinge area of \u200b\u200b20 amino acid residues. It provides a compound of a- and p-chains using disulfide bonds. A transmembrane hydrophobic domain of 22 amino acid residues is located behind the hinge area, it is associated with a short intracytoplasmatic domain of 5-16 amino acid residues. The recognition of the T-cell receptor of the represented antigen occurs as follows. MNS molecules ClassP, as well as T-lymphocyte receptors, consist of two polypeptide chains - a and r. Their active center for binding represented antigenic peptides has the form of "gap". It is formed by spiral areas of the a- and p-chains, connected at the bottom of the "slit" with each other an unbeaten area formed by the segments of the other chain. In this center (slit), the MTC molecule joins the processed antigen and thus represents its T cells (Fig. 63). The active center of the T-cell receptor is formed by hypervariable sections of A- and P-chains. It also represents a kind of "gap", the structure of which corresponds to the spatial structure of the imaginary molecule molecule of the antigen's peptide fragment class to the same extent as the structure of the active center of the antibody molecule corresponds to the spatial structure of the determinant of the antigen. Each T-lymphocyte carries receptors only for one peptide, that is, it is specific to a specific antigen and connects the processed peptide of only one type. The addition of the imagined antigen to the T-cell receptor induces the transmission of the signal from it on the cell genome.
It requires its contact with the CD3 molecule to operate any TCR. It consists of 5Subedinits, each of which is encoded by its genome. CD3 molecules have all subclasses of T-lymphocytes. Due to the interaction of the T cell receptor with the CD3 molecule, the following processes are provided: a) TKR removal on the surface of the T-lymphocyte membrane; b) giving the appropriate spatial structure of the molecule of the T-cell receptor; c) receiving and transmitting the signal T-cell receptor after its contact with the antigen in the cytoplasm, and then in the T-lymphocyte gene through the phosphatidality cascade with the participation of intermediaries.
As a result of the interaction of the MNC molecule, a class support of the antigenic peptide, with a peptide T-lymphocyte receptor, as it were, is embedded in the "gap" of the receptor, which is formed by hypervariable sections of A- and P-chains, in contact with both chains
The organization of genes encoding A- and (3-chains of TCR is mainly homologous to the one that is known for light and heavy immunoglobulin chains. V-domain A-chain, like a light chain of immunoglobulins, is monitored only by V- and J-gene segments. In The same time, the formation of a V-domain (3-chains, as well as a heavy immunoglobulin chain is provided with a complete set of V-, D-, Jennic segments (Fig. 3.14).
The T-cell genome has more than 100 V-genes for A-chains.
TCR, which is two and a half times less than the amount that is known for the light chains of immunoglobulins. Each such gene includes two exon - one for a leader (L) sequence that is missing in a mature A-chain, but presented in this chain at the time of its transport from the endoplasmic reticulum to the cell surface, and the second - for encoding the TCR V-domain itself. The J-gene segments for the A-chain are much larger than for a light chain of immunoglobulins (50 against 4). The constant area of \u200b\u200bthe A circuit is controlled by the C-genome, including individual exons for the C-domain, the hinge and one common exone - for transmembrane and tail parts of the molecule.
The number of V-genes for (3-chains is 30. In addition, there are two DJC clusters. Each cluster includes one D- and six J-gene segments. Functional differences between clusters are unknown. C-gene for the constant area | 3-chains includes Four exons for constant, hinge, transmembrane and tail sections of the polypeptide.
The processes of recombination, transcription, splicing and broadcasting of the genetic material for A- and (3-chains during the formation of TCR in T cells are similar to those providing synthesis of immunoglobulins in B cells.
In the same way as in the case of immunoglobulins and immunoglobulin receptors, the TCR variability depends on the random interaction of the gene segments in the process of recombination of the genetic material encoding V-domains: Vj - for the circuits and VDJ (3-chains. Calculation of variability of V-domains TCR, which is carried out in the same way as for immunoglobulins (see HL.
- , shows an extremely high level of diversity of these anti-
At the first stage of operation from mice immunized by a temonimal antigen (AG), a total, undifferentiated T-cell population containing a variety of clones (in fig. Figures 1 -6). The second stage was to release individual T-cell clones, among which were specific to the used antigen (in fig. As an example, four clones are given, one of which is clone 3, - specifically reacts with the antigen). The third stage of work included the production of monoclonal antibodies (MAT) to antigen-reactive clone. The task of this stage is to obtain monoclonal antibodies that can react only with a clone used for immunization. At the same time, the cross reaction mats about the overall specifics of the antigen-reactive clone and intransited clones (top table). The absence of cross-reactivity mat indicates the presence of a positively reactive adornal clone of particular specificity - presumably, antigensnical receptor. Confirmation of such an assumption is the reaction of the delay of interaction mat with a corresponding clone in the presence of the used antigen (bottom table). Obtaining mat to the antigensnial T-cell receptor created conditions for its full study
Fig. 3.13. The structure of YantngeshrvSooMP; YauczoShiro Resentshire T-fly *.
T-cell antigensnial receptor (TCR) is a heterodiment composed of a- and p-chains. Each chain includes two domains: variable (V) and constant (C). The interacting VA- and ur domains form an antigens of the TCR. In addition to the main V- and C-domains in the TCR structure, there is a hinge region with a cysteine \u200b\u200bresidue, forming a covalent bond between A- and P-chains, as well as transmembrane and short tail sectors
germinating structures (Table 3.2). With the general similarity of the organization and recombination of genetic material for immunoglobulins and TCR, it should be noted some features in controlling the specificity of these molecules.
Immunoglobulins and immunoglobulin 6-cell receptors recognize native antigenic epitopes. In this regard, the individual sections of the antigensnial center have equal chances of variability. The situation with TCR is somewhat different, since this receptor recognizes the complex of an antigenic peptide with MCS molecules.
A variety of TCR is associated with a large extent with a third loop of a V-domain A generated by the third hypervariable site - CDR3 (Soc. From English, "Compleventiarity Determining Region"). When the antigen binding center is formed by the V-houses, the CDR3 A- and 0-chains are found in the inside of this center. The first and second loop (CDR1 and CDR2, respectively) occupy the periphery of the center. In such a conformational place
on the
Fig. 3.14. Ortshnzatsp Geoon, controlling A- and R-CSECH T-Isyshugo Recents.
The principle of reorganization of gene segments controlling the A- and R-CPCs TKR, the tag is also as for immunoglobulins. The difference is that the locus for (3-chains has two identical clusters. What is the functional value of such duplication, is unknown
Table 3.2.
T-cell receptor variability in comparison with
immunoglobulin receptors and immunoglobulins 
researchum There is a completely defined biological meaning associated with the adaptation of TCR to the form of the antigen with which it interacts. As already noted, the antigenic peptides fill the space (gap) formed by A-spiral structures of MCS molecules and thus turn out to be in the middle of the antigenic complex of peptidgm. Such a complex is characterized by a huge set of antigenic specificities associated with peptides, and limited diversity characteristic of MNS molecules. In connection with such an organization, an immunogenic complex should be expected to expect an increased variability of CDR3 and a smaller variability of CDR1 and CDR2. The study of the genetic organization of genes for TCR confirms such a point of view. Thus, the TCR has a significantly less compared to immunoglobulin, the number of V-genes, which determine the specificity of CDR1 and CDR2, but, with an increased number of J-segments participating in CDR3 coding (see Table 3.2). In fig. 3.15 A simplified scheme is presented illustrating the interaction of soy with peptide and CDR1 and CDR2 with a-spiral sequence of MNS molecules.
Molecules related to the main histocompatibility complex (eng. MHC.) On the surface of antigen-representing cells. TCR consists of two subunits, borrowed in the cell membrane and are associated with a multi-suspension CD3 complex. The TCR interaction with MHC and the antigen associated with it leads to the activation of T-lymphocytes and is a key point in the launch of the immune response.
Structure
TCR is a heterodimeric protein consisting of two subunits - α and β or γ and δ, presented on the cell surface. Subunits are fixed in the membrane and are connected with each other disulfide bond.
According to its structure, the TCR subunit belongs to the superfamily of immunoglobulins. Each of the subunits is formed by two domains with a characteristic immunoglobulin laying, a transmembrane segment and a short cytoplasmic segment.
N-terminal domains are variable (V) and are responsible for binding an antigen presentable by molecules of the main histocompatibility complex. The composition of the variable domain contains a hypervariable portion (CDR) characteristic of immunoglobulins. Due to the extraordinary diversity of these areas, various T cells are able to recognize the widest range of various antigens.
The second domain is constant (c) and its structure is the same in all subunits of this type in a particular individual (with the exception of somatic mutations at the level of genes of any other proteins). On the plot between the C-domain and the transmembrane segment there is a substrate residue, with which a disulfide connection is formed between two TCR circuits.
The TCR subunits are aggregated with the CD3 membrane polypeptide complex. CD3 is formed by four types of polypeptides - γ, δ, ε and ζ. The subunits γ, δ and ε are encoded closely with the clutch genes and have a close structure. Each of them is formed by one constant immunoglobulin domain, transmembrane segment and long (up to 40 amino acid residues) by the cytoplasmic part. The chain ζ has a small extracellular domain, a transmembrane segment, and a large cytoplasmic domain. Sometimes instead of the chain ζ, the complex includes a chain η - a longer product of the same gene obtained by alternative splicing.
Since the structure of proteins of the CD3 complex is invariant (no variable sections), they are not able to determine the specificity of the receptor to the antigen. Recognition is exclusively a TCR function, and CD3 provides signal transmission into a cell.
The transmembrane segment of each of the CD3 subunits contains a negatively charged amino acid residue, and TCR is positively charged. Due to electrostatic interactions, they are combined into a common functional complex T-cell receptor. Based on stoichiometric studies and measuring the molecular weight of the complex, the most probable composition is (αβ) 2 + γ + δ + ε 2 + ζ 2.
TCR consisting of αβ chains and γδ-chains are very close in structure. These forms of receptors are presented in different ways in various tissues of the body.
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An excerpt characterizing T-cell receptor
- All Cossacks flaw. Cleaning the hut for Colonel, endured them. Pity to watch guys, "said Danceun. - They plunged them: so alive, believe whether it bursts something in his own way."And the pure people guys," said the first. - White, that's white birch, and there are brave, say noble.
- Do you think how? He from all ranks are scored.
"And nothing knows about our," Danceun said with a smile of bewilderment. "I'm telling him:" Whose crown? ", And he is a burst. Wonderful people!
"After all, then wisely, my brothers," the one who was surprised in whiteness was surprised, "the men were told under Mozhaisk, as they were to clean the battered, where she was, so that, he says, read the month lying the dead. Well, says, lies, says their own, how white paper, clean, nor blue powder smells.
- Well, from the cold, what about - asked one.
- Eca you're smart! By cold! It was hot because it was. You can also not rubley too. And then, says, come to our, all, says, rotted in the worms. So, he says, we will asshide the headscarves, yes, the title face, and tested; urine no. And their own, says how white paper; Neither blue powder smells.
Everyone was silent.
"Should, from food," said Feldfelf, "the gentleman's food erupted."
No one objected.
- the man told that this one, under Mozhaisk, where she was that, they were angry with ten villages, twenty days were drove, did not take all the dead, then. Wolves of these what, says ...
"That suffer was real," said the old soldier. - just was what to remember; And then everything after that ... so, only the people of torment.
- And that, uncle. The day before yesterday we came, so where they do not allow themselves. Vivid rifles left. On the knees. Pardon - says. So, only an example one. They told the Polyon itself, the boards took two times. Words do not know. I will take away: here on those in the hands will catch the bird, fly away, and it will fly away. And there is no position too.
- Eca to lie healthy you, Kiselev, I'll see for you.
- What to lie, though true.
"And I can be in my custom, I would be angry, and I would go to the ground." Yes Osinov Colom. And the fact that the people ruined.
"We will do all one end, will not walk," yawning, said the old soldier. "
The conversation is walked, the soldiers began to fit.
- Vish, stars, passion, so burn! Tell me, the cans were laid out, "the soldier said, admiring the Milky Way.
- This guys, to the crop year.
- The wood bar will still have.
- I'll get the back, and the belly frozen. Here is a miracle.
- Oh my God!
- What are you pushing something - about you one fire, or what? Hit ... Oblocked.
Because of the silence, the snoring of some of the fallen silence was heard; The rest turned and warmed, occasionally talking. From the distant, steps for a hundred, the fire was heard friendly, cheerful laughter.
"Vish, scream in the fifth company," said one soldier. - And the people that - passion!
One soldier rose and went to the fifth company.
"So then laughter," he said, returning. - Two Hranzuza stuck. One frown at all, and the other is so crushing, the bound! Songs playing.
- Oh about? Go to see ... - Several soldiers headed for the fifth company.
The fifth company stood beside the forest. The huge fire burned brightly in the midst of snow, lighting the trees aggravated by the branches.
In the middle of the night, the soldiers of the fifth company heard the steps in the forest in the forest in the snow and the chrysk bust.
"Guys, Veded," said one soldier. All raised their heads, listened, and from the forest, in the bright light of the fire, two, holding themselves for each other, human, weirdly dressed figures.
These were two french hiding in the forest. Hoarsely speaking something on incomprehensible soldiers, they approached the fire. One was higher than the growth, in an officer hat, and seemed quite weakened. Going to the fire, he wanted to sit down, but fell to the ground. Another, small, chunky, bangbed on the cheeks of the soldier, was stronger. He raised his comrade and, pointing to his mouth, said something. The soldiers were surrounded by the French, unkind the sore of the chinel and both brought cereals and vodka.
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The organization of human genes encoding the α-, β-and δ-chains of the T cell receptor is shown in Fig. 8.6. (Due to the complexity, the organization of γ-genes is not shown.) Several properties deserve special attention. First, α- and γ-chains are constructed from V- and J-gene segments, like L-circuits LG, while the β- and δ-chains are constructed from V-, D- and J-gene segments like n -Caps LG. Secondly, the locuses P and U are on different chromosomes, while the gene segments of α and δ locuses are located on one.
Fig. 8.6. Organization α-, β- and δ-genes of a person coding T-cell receptor
The genes encoding the δ-chain are limited from both sides (on 5 "- and 3" -concats) genes encoding the α-chain. Thirdly, there is more Vα- and Vβ genes in the germs than Vγ- and Vδ-genes (5-10). Notice also that there are two Cβ genes (Cβ1 and Cβ2), but these genes and their products are almost identical and any functional differences between them are unknown. Thus, they do not need to identify with antibody isotypes, in which the constant genes of the H-chain LG and their products differ significantly.
Separate TCR V-areas received rooms, for example Vα2 and Vβ7. Interestingly, the use of certain TCR V-regions in some cases has been associated with the response to specific antigens, especially the Superantigen - a number of antigens activating all T cells expressing a certain Vβ as a component of its TCR. For a person, Superantigenes are some bacterial toxins. The effect of these bacterial products on T cells can cause many reactions that often have clinical consequences.
The situation with TCR genes in some way is similar to this; β-, γ- and δ-genes TCR are subject to the rule of allelic exception, and α-genes are not. Thus, some T cells that use αβ as their TCR, have two different α-chains expressed with one β-chain, and therefore may have two different antigenic specificities. Up to 30% of human αβ-T-cells and the mouse express two α-chains, but their functional significance is still not clear.
Variety of T-cell receptors
The mechanisms of the occurrence of different T cell receptors are very similar to the mechanisms for the occurrence of B cell receptors. The basic principles of gene repairs are valid in the synthesis of the V- and C-regions of each chain of the T cell receptor (α, β, γ and δ). Recombinases and connecting sequences are used to connect the VJ or VDJ structures that ensure the specificity of the variable region of a certain TCR polypeptide chain. In the process of recombination in V- and T-cells, the same enzymes participate.In the activation of recombination genes in the early stages of differentiation of both V- and T-cells, two genes are played by two genes, called recombination activation genes (Rag-1 and Rag-2). Thus, as in the occurrence of the differences between LG, the differences in TCR are due to: 1) the presence of numerous V-genes in the embryonic line; 2) a random combination of chains; 3) the variability of connections and inserts. However, there is one important difference between the occurrence of differences in TCR and the LG molecules indicated earlier: in contrast to TCR LG after stimulation, the antigen is undergoing somatic hypermutagenesis.
The repertoire of various TCR is considered the same or even greater than the repertoire of LG molecules (estimated the number of possible differences in specificity for αβ is 1015, and for γδ-Tcr - 1018). The variability of compounds and inserts is an important component in the occurrence of various TCRs. It is due to it that it produces a huge number of different sequences of the hypervariable TCR section, known as CDR3. (In contrast, the CDR1- and CDR2-POCLEFOsTextric TCR does not occur during repaired, and the V-genome is encoded, which is in the embryo line.)
Crystallography data indicate that CDR3 is a section of the αβ-TCR binding center, which is in contact with the amino acids in the center of the peptide associated with the MNS molecule (see Fig. 9.3 and 9.4). Thus, a large number of different CDR3 sequences provides high TCR binding specificity with a peptide part of the Peptide MNC complex.
T-cell differentiation in thymus
Timus is a primary lymphoid body for the development of T-cells, similar to how the bone marrow is in mammals in the primary organ for differentiation of B cells. This emphasizes the absolute necessity of thymus for differentiation of immature predecessor cells into cells with the characteristics of T-lymphocytes. Unwanted consequences of the absence of thymus and, accordingly, the lack of mature T cells can be observed in children born without thymus (Di syndrome), and in mice with a genetically programmed absence of thymus (known as "naked" mice, because they also have no wool) .The differentiation of T-cells in the thymus occurs throughout the lifetime of the individual, but significantly decreases after the pubertal period. The size of the thymus itself in mammals decreases with the onset of puberty (thymus coaluation) predominantly due to the synthesis at this time of steroid hormones. In some species, especially in mice, the population of mature T cells is sharply depleted if the thymus is removed shortly after birth. In fact, this observation made it possible to identify the determining role of thymus in T-cell responses. The removal of thymus in animals at a later time has a much smaller effect on the population of mature T-cells.
T-cell differentiation in thymus is a complex multi-stage process. In the following subsections and in Fig. 8.7 We note a number of the main phases in the differentiation sequence.
Timocyte interaction with thymic nelmfoid cells
In fig. 8.7 It is shown that at each stage of ripening in the thymus (from the precursor cell to a mature T-cell), developing T-lymphocytes (Timocytes) are in contact and interact with a network formed by non-lifoid (stromal) cells of thymus. Timocytes move through the network of non-lifoid cells from the outer zone - the thymus bark to the inner - thymus cerebral substance.
Fig. 8.7. Ways of development of T-cells in Timus
The most important non-lifoid cells of the thymus are:1) Cortical epithelial cells; 2) Dendritic cells located primarily on the border of the crust and brainstant. The dendritic cells of the thymus occur from the bone marrow and enter the same family of cells, which presents the presentation of T-cell antigens in other tissues and organs.
Next, we will discuss in more detail how non-lifoid cells provide the main intercellular communications necessary for the development of ripening T-lymphocytes. They also produce cytokine IL-7, which induces proliferation (C-) T-lymphocytes in the early stages of development. Timus is the place of intensive proliferation of developing T cells, however, the overwhelming majority of these daily produced cells, estimated by about 95%, die without leaving it.
Rearander genes of T-cell receptors
Lymphoid predecessor cells penetrate into the outer sections of the thymus (subcapsular zone); At the same time, their TCR genes are in a non-repayable configuration (germline). It is usually believed that then the genes of γ-, δ- and β-chains take into reanigration almost simultaneously. Cells that productively rearant the γ- and δ-genes express γ- and δ-chains TCR on the cell surface. The sequence of processes in the early stages of the rearrangement of TCR genes is still not clear, and are not clear whether the cells expressing the γ- and δ-chains on their surface are capable of productively rearanate the β-chain gene.Despite this, observations suggest that cells expressing γ- and δ-chains as their TCR are separated from cells that will express α- and β-chains as their receptor, in the early stages of development in Timus, although the stage on which this happens, it is still not fixed. The cells expressing γδ as their TCR leaving the thymus and form a pool of peripheral γδ-T-cells.
Cells, productive rearantrating β-genes, express the β-chain TCR on the cell surface in the association with an invariant molecule known as pre-Tα. They are called pre-t-cells, and the combination of β-chain and pre-tα (together with CD3 and ζ) constitute the pre-T-cell receptor (PPE-TCR) is similar to pre-cells and pre-in-cell receptors.
Cells expressing the PPE-TCR are differentiated further. Similarly, the phases of the differentiation of pre-cells, the transmission of the signal through the PPE-TCR ceases to rearrange the TCR β-genes. This achieves that the cells express only one type of β-chain (allelic exception). In addition, the cells are proliferated, and the expression of pre-tα is suppressed in this extended population, α-genes begin to render and expression of CD4 and CD8-genes are allowed.
As indicated previously, the gene segments α and δ-loci TCR are located on the same chromosome, so the rearrangement of α-locus on a certain chromosome leads to the exclusion of δ-locus. (This ensures that the β-chain does not become a pair of 8-chain.) Thus, the next important stage of maturation of the αβ-line cell is the expression of CD4 and CD8 coreceptor molecules on its surface. Such αβ + - CD3 + CD4 + CD8 +Timocyte belonging to CD4 + CD8 +, or twice a positive cell, is found in the thymus cortex and forms most of the thymocytes in the thymus of young mammals.
Timic selection
Positive selection
Twice positive thymocyte passes through a multistage process of thymic selection (see Fig. 8.8). (There is a similar selection process before leaving the thymus γδ-T cell, is currently not clear.) At the first phase of the positive selection of TCR, twice positive thymocyte interact with MNC molecules expressed on epithelial cells in the thymus cortex.This interaction leads to survival and differentiation twice of positive cells; Those of them that do not participate in this important interaction and therefore are not selected, die by apoptosis. Positive selection also leads to the suppression of the expression of Rag-1 and Rag-2 genes and, thus, the cessation of further gene rearrangement. Therefore, since, as indicated earlier, the α-gene is not exposed to allele exception, positive selection ceases to further attempt to rearrange the α-chain repairs.
Another important property of positive selection is that the developing αβ-T cell becomes "trained" in relation to MNS molecules expressed by epithelial cells of the thymus bark. This means that the entire remaining T-cell life, even in the form of a mature cell that left the thymus will respond to the antigen only if it is associated with MNS molecules, with which the developing cell met in Timus. For this reason, the MNC molecules expressed in the Timus of the individual and the "trained" of its developing T cells belong to AUTU-MNS; All other types of MNS molecules for this person will be incompatible. This explains the emergence of phenomenon MHC restrictions or, more precisely, auto-MNS restriction, which is the main for T-cell response.
Negative selection
Since recombinations affecting the occurrence of TCR are more or less random, T-cells expressing TCRs specific about alien and own antigens can develop in thymus and pass positive selection. There is a chance that T cells with severe reactivity to their own components of the body will leave thymus and will interact with such antigens in the tissues, which can lead to unwanted autoimmune reactions. To prevent this twice, positive cells are subjected to the second phase of the selection - negative selection (Fig. 8.8).
Fig. 8.8. Positive and negative selection of αβ-TCR + CD4 + CD8 + T-cells in Timus
In fig. 8.8 shows a negative selection observed when twice positive cells interact with dendritic cells on the boundary of the cortical and brain layers. Molecules TCR, CD4 and CD8 interact expressed by twice positive Timocyte, and MNC molecules located on a dendritic cell. Since dendritic cells have peptides associated with MNC molecules, twice positive cells seem to interact with MNS and peptide expressed on the surface of the dendritic cell.
T-cells expressing TCRs that react with too high affinity with a combination of peptide and MNS are removed by apoptosis. A similar negative selection removes T cells expressing TCR with high reactivity to its own components.
Twice positive cells, survived negative selection, reduce expression both CD4 and CD8 by means of non-studied mechanism. All this leads to development or CD4 + -CD8-, or CD4 + -CD8 + -T-cells (monopositive). Two of these populations are the end point of the complex process of AP-TCR cell differentiation in the thymus. They leave the thymus and form peripherals (i.e., outside the thymus) of the mature CD4 + - and CD8 + -T-cell lines.
The role of peptides in thymic selection
A number of questions remain in the mechanisms involved in breeding. For example, what are the role and nature of peptides expressed by non-lifoid thymic cells at different stages of selection processes. These studies indicate that peptides expressed by cortical epithelial cells play a major role at the stage of positive selection. These peptides occur from autoantigen expressed in Timus or entered into it.Currently, it is not clear, however, as these peptides arising from autoantigen are selected T-cells with TCR, specific in relation to both incompatible and autologous antigens. In addition, it is not clear whether peptides are distinguished by cortical epithelial cells with positive selection, from those that are expressed by dendritic cells with negative selection.
Another unresolved question is how TCR interaction. Expressed by twice positive cells, with MNC molecules and peptides presented on cortical epithelial cells, leads to survival and differentiation twice of positive cells, while the interaction twice the positive cell with a dendritic thymus cell induces a negative signal (cellular death). These problems continue to study intensively.
Characteristics of T-cells leaving thymus
Differentiation in the thymus of T cells expressing αβ as its TCR, leads to the formation of the repertoire of peripheral CD4 + and CD8 + -T-cells capable of reacting to a huge number of alien antigens. These cells have two important characteristics.- They are characterized by auto-MNS restriction. They interact with peptides formed from intimidating antigens only when peptides are associated with the same set of MNS molecules, with which a developing T cell interacted during positive selection in Timus.
- They have an autotoolerance. CD4 + - and CD8 + -T cells do not respond to its own components.
