****This web page was produced as an assignment for an undergraduate course at Davidson College.****
The CD3 Complex
My Favorite Immunological Protein
An Introduction to CD3
The CD3 complex is a group of cell surface molecules that associates with the T-cell antigen receptor (TCR) and functions in the cell surface expression of TCR and in the signaling transduction cascade that originates when a peptide:MHC ligand binds to the TCR. CD3 is composed of invariant subchains belonging to the immunoglobulin superfamily. It is found on the surfaces of T cells and of thymocytes (Janeway et al., 2005).
The CD3 Subchains
CD3 is composed of three distinct subchains: γ, δ, and ε. These subchains are distinguishable by their molecular weights (Table 1).
Chain |
Molecular Weight (kDa) |
Gamma |
25-28 |
Delta |
20 |
Epsilon |
20 |
Table 1 . Molecular weights of B3 subchains. Adapted from Janeway et al., 2005.
Structurally, each CD3 subchain is composed of an extracellular immunoglobulin-like domain, a single-spanning transmembrane domain, and a cytoplasmic tail expressing a single ITAM (Janeway et al., 2005). The transmembrane domain of each subchain contains two acidic residues (Call et al., 2002).
The CD3 subchains are encoded in adjacent loci, and these genes are regulated as a unit (Janeway et al., 2005).
The TCR Complex
The TCR complex consists of a TCR, a B3 complex, and a two ζ subchains, which form a homodimer. The TCR itself consists of an α chain and a β chain that form a disulfide-linked heterodimer expressing one single antigen-binding site. Each CD3 complex contains one CD3γ subchain, one CD3δ subchain, and two CD3ε subchains. One of the CD3ε subchains forms a heterodimer with the CD3γ subchain, while the other CD3ε subchain forms a heterodimer with the CD3δ subchain (Call et al., 2002).
Covalent binding does not occur between the CD3γ subchain and the CD3δ subchain, and CD3γγ, CD3δδ, and CD3εε homodimers do not form (Sun et al., 2001).
The exact arrangement of the CD3 complex is a matter of intense debate in immunological circles because it is uncertain whether the six accessory chains associate with one or with two α:β heterodimers (Fernandez-Miguel et al., 1999).
Competing Theories
Based upon the results from a TCR stoichiometry experiment in mice, Gemma Fernandez-Miguel and others present a two TCRα:β heterodimer model. Assays of spleen T cells from transgenic mice with alleles for two different human TCRβ subchains with different variable regions demonstrate that each of the two subtypes of TCRβ subchains is present in a single TCR complex. This indicates that two TCR α:β heterodimers are present in each TCR complex. The researchers are cautious, however, concerning the universality of their experimental system (Fernandez-Miguel et al., 1999).
Matthew E. Call, among others, presents a one α:β heterodimer model in which the one of the two acidic residues present in the transmembrane domain of each CD3 subchain interacts with one of the three basic residues in the transmembrane domain of each TCR subchain. The model proposes that each single-spanning transmembrane domain is configured in an α-helical structure, which configuration allows the creation of trimeric interfaces between one basic residue in the transmembrane domain of a TCR subchain and one acidic residue in the transmembrane domain of each of two CD3 subchains. In this model, the TCRα subchain associates with the CD3δ:ε heterodimer, and the TCRβ subchain associates with the CD3γ:ε heterodimer. In order for its subchains to associate into a functional TCR complex, each TCR subchain must express a lysine in its transmembrane region (Call et al., 2002).
The Assembly of the TCR Complex
The TCR complex is assembled within the endoplasmic reticulum (ER). The genes coding for the CD3 subchains are transcribed and translated before the genes encoding for the TCR subchains are transcribed and translated, leading researchers to believe that the TCR α and β subchains associate with the CD3 complex only after all four of its subchains have been assembled successfully. The formation of the disulfide bridge between the two TCR subchains happens subsequently to the α and β subchains binding individually to the CD3 complex. Researchers have demonstrated that the formation of this bond is inefficient in the absence of the CD3 complex, leading them to believe that the CD3 complex actually facilitates the formation of the disulfide bridge. After the TCR complex has assembled completely, it is exported to the Golgi apparatus, where it undergoes oligosaccharide processing to become a mature TCR complex. Only then does the cell express the TCR complex on its surface (Alarcon et al., 1988).
The Function of the CD3 Complex in T Cell Activation
The CD3 complex must be present on the T-cell surface for TCRs to be expressed by the cell. Individuals lacking or with mutant versions of the CD3γ subchain or of the CD3ε subchain express low levels of TCRs on their cell surfaces and are also deficient in their T-cell response (Sun et al., 2001).
Once a peptide:MHC ligand has bound to the TCR, the CD3 complex functions in the transduction of the signal into the cytoplasm, which initiates a signaling cascade that ultimately activates the T cell. Two SRC-family tyrosine kinases known as Lck and Fyn, which associates with the cytoplasmic tails of both the CD3ε subchains and the ζ subchains of the TCR complex, phosphorylate and bind to the CD3ε subchains' ITAM regions. This binding allows for the binding of ZAP-70 to the ζ subchains, which propagates the remainder of the signaling cascade (Janeway et al., 2005).
The Fate of the TCR Complex in Activated and in Resting T Cell
While the binding of a peptide:MHC ligand to the TCR promotes significant downregulation of TCR complexes, the rate of TCR internalization does not increase significantly. This trend suggests that the binding of the ligand blocks TCR complexes from returning to the cell surface as opposed to increasing the rate of internalization of TCR complexes expressed on the cell surface. Research indicates that the TCR complexes are long-lived but constantly recycled by Rab4+ early endosomes. Most TCR complexes remain in endosomes without being degraded. Of the internalized TCR complexes that are degraded, approximately equal amounts are degraded within lysosomes and by proteosomes in the cytosol.
Further research suggests that the carboxy terminals of the ζ subchains play an important role in maintaining the cell surface expression of a TCR complex. Once a peptide:MHC ligand has bound to a TCR, the conformation of the TCR complex changes, releasing the ζ:ζ homodimer from the complex. This might indicate that the ζ:ζ homodimer covers a motif in the CD3 complex that, once exposed, allows for the internalization of the TRC complex by endocytosis (Vignali, 2003).
The Role of CD3 in Human Severe Combined Immunodeficiency (SCID)
Currently, researchers have identified at least ten distinct genes that encode for mutant proteins causing SCID in humans. Five of these genes encode mutant forms of polypeptides that are present in antigen-receptor accessory proteins, including the CD3δ and the CD3ε subchains. Mutations resulting in a premature stop codon alter the subchains' conformations to such an extent that they are not expressed on the T-cell surface. The absence of either of these subchains completely halts T cell development. Apparently, the absence of the CD3ε subchain blocks T cell development in the pre-TCRα double-negative stage, while the absence of the CD3δ subchain blocks development slightly later in the intermediate single-positive stage just before the double-positive stage. A deficiency in the CD3γ subchain, however, does not appear to affect T cell development (de Saint Basile et al., 2004).
The Role of the CD3 Complex in Prolonging Allograft Survival
Experimental data indicates that the immunotoxin FN18-CRM9, which consists of the anti-CD3 monoclonal antibody FN18 and a mutated diphtheria toxin designated CRM9, might prolong allograft survival in transplant patients. Preclinical trials on purified rhesus monkey T cells indicate that, although FN18 and FN18-CRM9 are equally efficacious in triggering the monoclonal antibody-mediated cross-linking of the TCR complex, FN18-CRM9 is more potent in triggering the phosphorylation of tyrosine residues and thereby triggering the T cells to internalize the CD3 complex (Hamaway et al., 2001). Without the CD3 complex, the TCR will not be expressed, and the T cell therefore will not activate.
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Literature Cited
Alarcon, B., Berkhout, B., Breitmeyer, J., Terhorst, C. 1988. Assembly of human T cell receptor – CD3 complex takes place in the endoplasmic reticulum and involves intermediary complexes between the CD3γ,δ,ε core and single T cell receptor α or β chains. J. of Biological Chemistry. 263(6): 2953-2961.
Call, M. E., Pyrdol, J., Wiedmann, K.W., Wucherpfennig, K. W. 2002. The organizing principle in the formation of the T-cell receptor – CD3 complex. Cell. 11(7): 967-979.
de Saint Basile, G., Geissmann, F., Flori, E., Uring-Lambert, B., Soudais, C., Cavazzana-Calvo, M., Durandy, A., Jabado, N., Fischer, A., Le Deist, F. 2004. Severe combined immunodeficiency caused by deficiency in either the δ or the ε subunit of CD3. J. of Clinical Investigation. 114(10): 1512-1517.
Fernandez-Miguel, G., Alarcon, B., Iglesias, A., Bluethmann, H., Alvarez-Mon, M., Sanz, E., de la Hera, A. 1999. Multivalent structure of an α:β T cell receptor. Proceedings of the National Academy of Scence USA. 96: 1547-1552.
Hamawy, M. M., Tsuchida, M., Cho, C. S., Manthei, E. R., Fechner, J. H., Knechtle, S. J. 2001. Immunotoxin FN18-CRM9 induces stronger T cell signaling than unconjugated monoclonal antibody FN18. Transplantation. 72(3): 496-503.
Janeway, C. A., Travers, P., Walport, M., Sclomchik, M. J. 2005. Immunobiology: the immune system in health and disease, 6th ed. Garland Science Publishing, NY, pp. 214- 216.
Sun, Z. J., Kim, K. S., Wagner, G., Reinherz, E. L. 2001. Mechanisms contributing to T cell receptor signaling and assembly revealed by the solution structure of an ectodomain fragment of the CD3ε:γ heterodimer. 2001. Cell. 105(7): 913-923.
Vignali, D. D. A. 2003 Apr. Recycling and downmodulation of the TCR:CD3 complex. <http://www.stjude.org/faculty/0,2512,407_2030_4255,00.html>. Accessed 2006 Mar 16.
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Contact me. (Summerlee Walter) suwalter@davidson.edu)