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V(D)J recombination is the mechanism of somatic recombination that occurs only in developing
lymphocyte A lymphocyte is a type of white blood cell in the immune system The immune system is a network of biological processes that protects an organism In biology, an organism (from Ancient Greek, Greek: ὀργανισμός, ''organismos'' ...

lymphocyte
s during the early stages of T and B cell maturation. It results in the highly diverse repertoire of
antibodies/immunoglobulins
antibodies/immunoglobulins
and
T cell receptor The T-cell receptor (TCR) is a protein complex found on the surface of T cells, or T lymphocytes, that is responsible for recognizing fragments of antigen as peptides bound to major histocompatibility complex (MHC) molecules. The binding be ...
s (TCRs) found in
B cell #REDIRECT B cell 3D rendering of a B cell B cells, also known as B lymphocytes, are a type of white blood cell White blood cells (WBCs), also called leukocytes or leucocytes, are the cells of the immune system The immune system is a ne ...
s and
T cell A T cell is a type of lymphocyte A lymphocyte is a type of white blood cell in the immune system The immune system is a network of biological processes that protects an organism In biology, an organism (from Ancient Greek, Greek: ...
s, respectively. The process is a defining feature of the
adaptive immune system The adaptive immune system, also referred as the acquired immune system, is a subsystem of the immune system The immune system is a network of biological processes that protects an organism In biology, an organism (from Ancient Greek, ...
. V(D)J recombination in mammals occurs in the primary lymphoid organs (
bone marrow Bone marrow is a semi-solid tissue Tissue may refer to: Biology * Tissue (biology), an ensemble of similar cells that together carry out a specific function * ''Triphosa haesitata'', a species of geometer moth found in North America * ''Triphos ...
for B cells and
thymus The thymus is a specialized primary Lymphatic system#Structure, lymphoid organ of the immune system. Within the thymus, T cell, thymus cell lymphocytes or ''T cells'' mature. T cells are critical to the adaptive immune system, where the body ad ...

thymus
for T cells) and in a nearly random fashion rearranges variable (V), joining (J), and in some cases, diversity (D) gene segments. The process ultimately results in novel
amino acid Amino acids are organic compound In , organic compounds are generally any s that contain - . Due to carbon's ability to (form chains with other carbon s), millions of organic compounds are known. The study of the properties, reactions, a ...

amino acid
sequences in the antigen-binding regions of immunoglobulins and TCRs that allow for the recognition of
antigen In immunology Immunology is a branch of biology that covers the study of immune systems in all organisms. Immunology charts, measures, and contextualizes the Physiology, physiological functioning of the immune system in states of both health ...
s from nearly all pathogens including
bacteria Bacteria (; common noun bacteria, singular bacterium) are ubiquitous, mostly free-living organisms often consisting of one Cell (biology), biological cell. They constitute a large domain (biology), domain of prokaryotic microorganisms. Typ ...

bacteria
,
viruses A virus is a wikt:submicroscopic, submicroscopic infectious agent that Viral replication, replicates only inside the living Cell (biology), cells of an organism. Viruses infect all life forms, from animals and plants to microorganisms, incl ...
,
parasites Parasitism is a Symbiosis, close relationship between species, where one organism, the parasite, lives on or inside another organism, the Host (biology), host, causing it some harm, and is adaptation (biology), adapted structurally to this w ...
, and
worms The World Register of Marine Species (WoRMS) is a taxonomic database that aims to provide an authoritative and comprehensive list of names of marine organisms. Content The content of the registry is edited and maintained by scientific specialists ...
as well as "altered self cells" as seen in
cancer Cancer is a group of diseases involving abnormal cell growth with the potential to invade or spread to other parts of the body. These contrast with benign tumor A benign tumor is a mass of cells Cell most often refers to: * Cell (biolo ...

cancer
. The recognition can also be
allergic Allergies, also known as allergic diseases, are a number of conditions caused by hypersensitivity Hypersensitivity (also called hypersensitivity reaction or intolerance) refers to undesirable reactions produced by the normal immune system, i ...

allergic
in nature (''e.g.'' to
pollen Pollen is a powdery substance consisting of pollen grains which are microsporophytes of seed plants The spermatophytes (; ), also known as phanerogams (taxon Phanerogamae) or phaenogams (taxon Phaenogamae), comprise those plant Plant ...

pollen
or other
allergen An allergen is a type of antigen In immunology Immunology is a branch of biology that covers the study of immune systems in all organisms. Immunology charts, measures, and contextualizes the Physiology, physiological functioning of the im ...
s) or may match host tissues and lead to
autoimmunity Autoimmunity is the system of immune responses of an organism against its own healthy cells, tissues and other body normal constituents. Any disease that results from such an aberrant immune response is termed an "autoimmune disease An autoimmu ...
. In 1987,
Susumu Tonegawa is a Japanese scientist who was the sole recipient of the Nobel Prize for Physiology or Medicine in 1987, for his discovery of the Genetics, genetic mechanism that produces antibody diversity. Although he won the Nobel Prize for his work in immun ...
was awarded the
Nobel Prize in Physiology or Medicine The Nobel Prize in Physiology or Medicine is awarded yearly by the Nobel Assembly , native_name_lang = , image = Nobel Assembly at Karolinska Institutet.jpeg , size = , motto = , formation = 190113 March 1978(as a forma ...
"for his discovery of the genetic principle for generation of antibody diversity".


Background

Human
antibody An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein used by the immune system to identify and neutralize foreign objects such as pathogenic bacteria and Viral disease, viruses. The antibody recognizes a unique mo ...

antibody
molecules (including
B cell receptor The B cell receptor (BCR) is a transmembrane protein on the surface of a B cell. A B cell receptor includes both CD79 and the antibody, immunoglobulin. The cell membrane, plasma membrane of a B cell is indicated by the green phospholipids. The B c ...
s) are composed of heavy and light chains, each of which contains both ''constant'' (C) and ''variable'' (V) regions, genetically encoded on three
loci Locus (plural loci) is Latin for "place". It may refer to: Entertainment * Locus (comics), a Marvel Comics mutant villainess, a member of the Mutant Liberation Front * Locus (magazine), ''Locus'' (magazine), science fiction and fantasy magazine ...
: *The immunoglobulin heavy locus (
IGH@ Immunoglobulin heavy locus, also known as IGH, is a region on human chromosome 14 Chromosome 14 is one of the 23 pairs of chromosome A chromosome is a long DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni( ...
) on chromosome 14, containing the gene segments for the
immunoglobulin heavy chain An antibody molecule. The two heavy chains are colored red and blue and the two light chains green and yellow. The immunoglobulin heavy chain (IgH) is the large polypeptide subunit of an antibody An antibody (Ab), also known as an immunoglobul ...
. *The immunoglobulin kappa (κ) locus (
IGK@ Immunoglobulin kappa locus, also known as IGK@, is a region on human chromosome 2 that contains genes for the kappa (κ) immunoglobulin light chain, light chains of antibodies (or immunoglobulins). In humans the κ chain is coded for by IGKV, V (va ...
) on chromosome 2, containing the gene segments for part of the
immunoglobulin light chain ] The immunoglobulin light chain is the small Peptide, polypeptide subunit of an antibody (immunoglobulin). A typical antibody is composed of two immunoglobulin (Ig) immunoglobulin heavy chain, heavy chains and two Ig light chains. In humans The ...
. *The immunoglobulin lambda (λ) locus (
IGL@ Immunoglobulin lambda locus, also known as IGL@, is a region on human chromosome 22 that contains gene In biology, a gene (from ''genos'' "...Wilhelm Johannsen coined the word gene to describe the Mendelian_inheritance#History, Mendelian u ...
) on chromosome 22, containing the gene segments for the remainder of the
immunoglobulin light chain ] The immunoglobulin light chain is the small Peptide, polypeptide subunit of an antibody (immunoglobulin). A typical antibody is composed of two immunoglobulin (Ig) immunoglobulin heavy chain, heavy chains and two Ig light chains. In humans The ...
. Each heavy chain or light chain gene contains multiple copies of three different types of gene segments for the variable regions of the antibody proteins. For example, the human immunoglobulin heavy chain region contains 2 Constant (Cμ and Cδ) gene segments and 44 Variable (V) gene segments, plus 27 Diversity (D) gene segments and 6 Joining (J) gene segments. The light chain genes possess either a single (Cκ) or four (Cλ) Constant gene segments with numerous V and J gene segments but do not have D gene segments. DNA rearrangement causes one copy of each type of gene segment to go in any given lymphocyte, generating an enormous antibody repertoire; roughly 3×1011 combinations are possible, although some are removed due to self reactivity. Most
T-cell receptor The T-cell receptor (TCR) is a protein complex found on the surface of T cells, or T lymphocytes, that is responsible for recognizing fragments of antigen as peptides bound to major histocompatibility complex (MHC) molecules. The binding be ...
s are composed of a variable
alpha chain The term alpha chain is normally used to indicate one of the subunits of a multi-subunit protein. The term "chain" is a general term given to any peptide sequence Peptides (from Greek language πεπτός, ''peptós'' "digested"; derived from π ...
and a beta chain. The T cell receptor genes are similar to immunoglobulin genes in that they too contain multiple V, D, and J gene segments in their beta chains (and V and J gene segments in their alpha chains) that are rearranged during the development of the lymphocyte to provide that cell with a unique antigen receptor. The T cell receptor in this sense is the topological equivalent to an antigen-binding fragment of the antibody, both being part of the immunoglobulin superfamily. An autoimmune response is prevented by eliminating cells that self-react. This occurs in the thymus by testing the cell against an array of self antigens expressed through the function of the
autoimmune regulator The autoimmune regulator (''AIRE'') is a protein that in humans is encoded by the ''AIRE'' gene. It is a 13kb gene on chromosome 21q22.3 that has 545 amino acids. AIRE is a transcription factor expressed in the Thymus#Medulla, medulla (inner part ...
(AIRE). The immunoglobulin lambda light chain locus contains protein-coding genes that can be lost with its rearrangement. This is based on a physiological mechanism and is not pathogenetic for leukemias or lymphomas. A cell persists if it creates a successful product that does not self-react, otherwise it is pruned via
apoptosis Apoptosis (from Ancient Greek Ancient Greek includes the forms of the Greek language Greek ( el, label=Modern Greek Modern Greek (, , or , ''Kiní Neoellinikí Glóssa''), generally referred to by speakers simply as Greek (, ) ...

apoptosis
.


Immunoglobulins


Heavy chain

In the developing
B cell #REDIRECT B cell 3D rendering of a B cell B cells, also known as B lymphocytes, are a type of white blood cell White blood cells (WBCs), also called leukocytes or leucocytes, are the cells of the immune system The immune system is a ne ...
, the first recombination event to occur is between one D and one J gene segment of the heavy chain locus. Any DNA between these two gene segments is deleted. This D-J recombination is followed by the joining of one V gene segment, from a region upstream of the newly formed DJ complex, forming a rearranged VDJ gene segment. All other gene segments between V and D segments are now deleted from the cell's genome. Primary transcript (unspliced RNA) is generated containing the VDJ region of the heavy chain and both the constant ''mu'' and ''delta'' chains (Cμ and Cδ). (i.e. the primary transcript contains the segments: V-D-J-Cμ-Cδ). The primary RNA is processed to add a
polyadenylated
polyadenylated
(poly-A) tail after the Cμ chain and to remove sequence between the VDJ segment and this constant gene segment.
Translation Translation is the communication of the meaning Meaning most commonly refers to: * Meaning (linguistics), meaning which is communicated through the use of language * Meaning (philosophy), definition, elements, and types of meaning discusse ...

Translation
of this mRNA leads to the production of the
IgM Immunoglobulin M (IgM) is one of several isotypes of antibody An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein used by the immune system to identify and neutralize foreign objects such as pathogenic bacte ...

IgM
heavy chain protein.


Light chain

The kappa (κ) and lambda (λ) chains of the immunoglobulin light chain loci rearrange in a very similar way, except that the light chains lack a D segment. In other words, the first step of recombination for the light chains involves the joining of the V and J chains to give a VJ complex before the addition of the constant chain gene during primary transcription. Translation of the spliced mRNA for either the kappa or lambda chains results in formation of the Ig κ or Ig λ light chain protein. Assembly of the Ig μ heavy chain and one of the light chains results in the formation of membrane bound form of the immunoglobulin IgM that is expressed on the surface of the immature B cell.


T cell receptors

During
thymocyte A Thymocyte is an immune cell present in the thymus The thymus is a specialized primary lymphoid organ of the immune system The immune system is a network of biological processes that protects an organism In biology, an organism (fr ...
development, the
T cell receptor The T-cell receptor (TCR) is a protein complex found on the surface of T cells, or T lymphocytes, that is responsible for recognizing fragments of antigen as peptides bound to major histocompatibility complex (MHC) molecules. The binding be ...
(TCR) chains undergo essentially the same sequence of ordered recombination events as that described for immunoglobulins. D-to-J recombination occurs first in the β-chain of the TCR. This process can involve either the joining of the Dβ1 gene segment to one of six Jβ1 segments or the joining of the Dβ2 gene segment to one of six Jβ2 segments. DJ recombination is followed (as above) with Vβ-to-DβJβ rearrangements. All gene segments between the Vβ-Dβ-Jβ gene segments in the newly formed complex are deleted and the primary transcript is synthesized that incorporates the constant domain gene (Vβ-Dβ-Jβ-Cβ). mRNA transcription splices out any intervening sequence and allows translation of the full length protein for the TCR β-chain. The rearrangement of the alpha (α) chain of the TCR follows β chain rearrangement, and resembles V-to-J rearrangement described for Ig light chains (see above). The assembly of the β- and α- chains results in formation of the αβ-TCR that is expressed on a majority of
T cell A T cell is a type of lymphocyte A lymphocyte is a type of white blood cell in the immune system The immune system is a network of biological processes that protects an organism In biology, an organism (from Ancient Greek, Greek: ...
s.


Mechanism


Key enzymes and components

The process of V(D)J recombination is mediated by VDJ recombinase, which is a diverse collection of enzymes. The key enzymes involved are recombination activating genes 1 and 2 (RAG),
terminal deoxynucleotidyl transferase Terminal deoxynucleotidyl transferase (TdT), also known as DNA nucleotidylexotransferase (DNTT) or terminal transferase, is a specialized DNA polymerase A DNA polymerase is a member of a family of enzyme Enzymes () are s that act as s ( ...
(TdT), and
Artemis Artemis (; grc-gre, Ἄρτεμις Artemis, ) is the Greek goddess Greek mythology is the body of myth Myth is a folklore genre consisting of narratives that play a fundamental role in a society, such as foundational tales or ori ...
nuclease, a member of the ubiquitous
non-homologous end joining Non-homologous end joining (NHEJ) is a pathway that repairs double-strand breaks in DNA. NHEJ is referred to as "non-homologous" because the break ends are directly ligated without the need for a homologous template, in contrast to homology directe ...
(NHEJ) pathway for DNA repair. Several other enzymes are known to be involved in the process and include DNA-dependent protein kinase (DNA-PK), X-ray repair cross-complementing protein 4 (XRCC4), DNA ligase IV,
non-homologous end-joining factor 1 Non-homologous end-joining factor 1 (NHEJ1), also known as Cernunnos or XRCC4-like factor (XLF), is a protein Proteins are large biomolecule , showing alpha helices, represented by ribbons. This poten was the first to have its suckture sol ...
(NHEJ1; also known as Cernunnos or XRCC4-like factor , the recently discovered Paralog of XRCC4 and XLF (PAXX), and DNA polymerases λ and μ. Some enzymes involved are specific to lymphocytes (''e.g.'', RAG, TdT), while others are found in other cell types and even ubiquitously (''e.g.'', NHEJ components). To maintain the specificity of recombination, V(D)J recombinase recognizes and binds to
recombination signal sequences Recombination signal sequences are conserved sequences of noncoding DNA that are recognized by the Recombination-activating gene, RAG1/RAG2 enzyme complex during V(D)J recombination in immature B cells and T cells. Recombination signal sequences g ...
(RSSs) flanking the variable (V), diversity (D), and joining (J) genes segments. RSSs are composed of three elements: a heptamer of seven conserved nucleotides, a spacer region of 12 or 23 basepairs in length, and a nonamer of nine conserved nucleotides. While the majority of RSSs vary in sequence, the consensus heptamer and nonamer sequences are CACAGTG and ACAAAAACC, respectively; and although the sequence of the spacer region is poorly conserved, the length is highly conserved. The length of the spacer region corresponds to approximately one (12 basepairs) or two turns (23 basepairs) of the DNA helix. Following what is known as the 12/23 Rule, gene segments to be recombined are usually adjacent to RSSs of different spacer lengths (''i.e.'', one has a "12RSS" and one has a "23RSS"). This is an important feature in the regulation of V(D)J recombination.


Process

V(D)J recombination begins when V(D)J recombinase (through the activity of RAG1) binds a RSS flanking a coding gene segment (V, D, or J) and creates a single-strand nick in the DNA between the first base of the RSS (just before the heptamer) and the coding segment. This is essentially energetically neutral (no need for
ATP hydrolysis 350px, Four possible resonance structures for inorganic phosphate ATP hydrolysis is the catabolic reaction process by which chemical energy Chemical energy is the potential of a chemical substance A chemical substance is a form of matter ...
) and results in the formation of a free 3'
hydroxyl group A hydroxy or hydroxyl group is a functional group with the chemical formula -OH and composed of one oxygen Oxygen is the chemical element Image:Simple Periodic Table Chart-blocks.svg, 400px, Periodic table, The periodic table of the ...

hydroxyl group
and a 5'
phosphate group In chemistry Chemistry is the science, scientific study of the properties and behavior of matter. It is a natural science that covers the Chemical element, elements that make up matter to the chemical compound, compounds composed of atoms ...

phosphate group
on the same strand. The reactive hydroxyl group is positioned by the recombinase to attack the
phosphodiester bond A phosphodiester bond occurs when exactly two of the hydroxyl groups A hydroxy or hydroxyl group is a functional group with the chemical formula -OH and composed of one oxygen atom Chemical bond , covalently bonded to one hydrogen atom. In or ...
of opposite strand, forming two DNA ends: a
hairpin A hairpin or hair pin is a long device used to hold a person's hair in place. It may be used simply to secure long hair out of the way for convenience or as part of an elaborate hairstyle A hairstyle, hairdo, haircut or coiffure refer ...

hairpin
(stem-loop) on the coding segment and a
blunt end Blunt end may refer to: * Sticky and blunt ends DNA ends refer to the properties of the end of DNA Deoxyribonucleic acid (; DNA) is a molecule File:Pentacene on Ni(111) STM.jpg, A scanning tunneling microscopy image of pentacene molec ...
on the signal segment. The current model is that DNA nicking and hairpin formation occurs on both strands simultaneously (or nearly so) in a complex known as a ''recombination center''. The blunt signal ends are flush ligated together to form a circular piece of DNA containing all of the intervening sequences between the coding segments known as a signal joint (although circular in nature, this is not to be confused with a
plasmid A plasmid is a small, extrachromosomal DNA Extrachromosomal DNA (abbreviated ecDNA) is any DNA that is found off the chromosome A chromosome is a long DNA molecule with part or all of the genetic material of an organism. Most eukaryo ...
). While originally thought to be lost during successive cell divisions, there is evidence that signal joints may re-enter the genome and lead to pathologies by activating
oncogene An oncogene is a gene In biology Biology is the natural science that studies life and living organisms, including their anatomy, physical structure, Biochemistry, chemical processes, Molecular biology, molecular interactions, Physio ...

oncogene
s or interrupting
tumor suppressor gene A tumor suppressor gene (TSG), or anti-oncogene, is a gene In biology, a gene (from ''genos'' "...Wilhelm Johannsen coined the word gene to describe the Mendelian_inheritance#History, Mendelian units of heredity..." (Greek language, Greek) ...
function(s) The coding ends are processed further prior to their ligation by several events that ultimately lead to junctional diversity. Processing begins when DNA-PK binds to each broken DNA end and recruits several other proteins including Artemis, XRCC4, DNA ligase IV, Cernunnos, and several DNA polymerases. DNA-PK forms a complex that leads to its
autophosphorylation File:Regulation of the Src-kinase by autophosphorylation.png, Fig. 2: Regulation of Src-kinase by autophosphorylation. Adapted from Frame (2002) Autophosphorylation is a type of post-translational modification of proteins. It is generally defined ...
, resulting in activation of Artemis. The coding end hairpins are opened by the activity of Artemis. If they are opened at the center, a blunt DNA end will result; however in many cases, the opening is "off-center" and results in extra bases remaining on one strand (an overhang). These are known as palindromic (P) nucleotides due to the palindromic nature of the sequence produced when DNA repair enzymes resolve the overhang. The process of hairpin opening by Artemis is a crucial step of V(D)J recombination and is defective in the severe combined immunodeficiency (scid) mouse model. Next, XRCC4, Cernunnos, and DNA-PK align the DNA ends and recruit terminal deoxynucleotidyl transferase (TdT), a template-independent DNA polymerase that adds non-templated (N) nucleotides to the coding end. The addition is mostly random, but TdT does exhibit a preference for G/C nucleotides. As with all known DNA polymerases, the TdT adds nucleotides to one strand in a 5' to 3' direction. Lastly, exonucleases can remove bases from the coding ends (including any P or N nucleotides that may have formed). DNA polymerases λ and μ then insert additional nucleotides as needed to make the two ends compatible for joining. This is a stochastic process, therefore any combination of the addition of P and N nucleotides and exonucleolytic removal can occur (or none at all). Finally, the processed coding ends are ligated together by DNA ligase IV. All of these processing events result in a
paratope A paratope, also known as an antigen-binding site, is the part of an antibody An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein used by the immune system to identify and neutralize foreign objects such as path ...
that is highly variable, even when the same gene segments are recombined. V(D)J recombination allows for the generation of immunoglobulins and T cell receptors to antigens that neither the organism nor its ancestor(s) need to have previously encountered, allowing for an adaptive immune response to novel pathogens that develop or to those that frequently change (''e.g.'',
seasonal influenza Flu season is an annually recurring time period characterized by the prevalence of an outbreak of influenza (flu). The season occurs during the cold half of the year in each hemispheres of Earth, hemisphere. Influenza activity can sometimes be ...
). However, a major caveat to this process is that the DNA sequence must remain in order to maintain the correct amino acid sequence in the final protein product. If the resulting sequence is out-of-frame, the development of the cell will be arrested, and the cell will not survive to maturity. V(D)J recombination is therefore a very costly process that must be (and is) strictly regulated and controlled.


See also

*
B cell receptor The B cell receptor (BCR) is a transmembrane protein on the surface of a B cell. A B cell receptor includes both CD79 and the antibody, immunoglobulin. The cell membrane, plasma membrane of a B cell is indicated by the green phospholipids. The B c ...
*
T cell receptor The T-cell receptor (TCR) is a protein complex found on the surface of T cells, or T lymphocytes, that is responsible for recognizing fragments of antigen as peptides bound to major histocompatibility complex (MHC) molecules. The binding be ...
* Basel Institute for Immunology * Charles M. Steinberg *
NKT cell Natural killer T (NKT) cells are a heterogeneous group of T cells that share properties of both T cells and natural killer cells. Many of these cells recognize the Polymorphism (biology), non-polymorphic CD1d molecule, an antigen-presenting molecu ...
*
Recombination-activating gene The recombination-activating genes (RAGs) encode parts of a protein complex is a protein complex functioning as a molecular biological machine A molecular machine, nanite, or nanomachine is a molecular component that produces quasi-mechanical ...


References


Further reading

* * V(D)J Recombination. Series: Advances in Experimental Medicine and Biology, Vol. 650 Ferrier, Pierre (Ed.) Landes Bioscience 2009, XII, 199 p. {{Immune system Immune system Lymphocytes Immunology