RAG Proteins

Kyle Cubin
3-26-07
Paper #2
Final Draft

The RAG Proteins: Ensuring Antibody Diversity

The ability of human immune cells to identify infected cells as harmful is critical to the immune system. Each different kind of pathogen, or infected cell, has its own unique signature that defines it as a danger to the human body and if it goes unrecognized by the lymphocytes it can kill the body. Because of the relative inability of the immune system to generally recognize harm and kill it efficiently, there must be an individual mechanism for identifying the specific threat and eliminating it effectively. It is the job of the antigen receptors on specific lymphocytes to recognize the antigen, or the distinguishing surface protein of pathogens, the only problem is the need for a specific antigen receptor for each antigen. The antigen receptor of the lymphocyte is guaranteed to be able to recognize nearly all antigens because of a process that occurs during DNA replication called V(D)J recombination. This process would not be possible without the RAG, Recombination Activating Genes, proteins that make sure the proper parts of the DNA are replicated. The RAG proteins are the essential machines of V(D)J recombination and make the human immune system as effective as it is.
The antigen receptor of immune cells is the critical part of the immune system that allows it to recognize nearly all threats that enter the body. Once a pathogen enters the body, it should hopefully be recognized by a lymphocyte so it can be destroyed before proliferation. The issue with this is that there is a specific lymphocyte that has the antigen receptor for the unique antigen and an immune response will ensue only if it comes into contact with that specific lymphocyte. Fortunately the immune system has an efficient way of presenting potential pathogens to all lymphocytes so the pathogen can be recognized. However, if the lymphocytes didn't undergo the process of V(D)J recombination during the cells DNA replication the diversity of the antigen receptors would not be great enough to recognize all viable threats.
The antigen receptors of lymphocytes consist of heavy and light chains with both constant and variable regions. These regions are encoded for in the DNA by three different genes; the heavy gene, plus the light chains kappa and lambda. The heavy chain gene has segments that fall into three categories, V, D, and J, while the light chain only uses V and J. In the light chain there are 200 possible kappa gene segments and 124 lambda gene segments; one of these light chains will pair with a heavy change variation. The heavy chain variation comes from there being 51 different possible V gene segments, 25 possible D segments, and 6 possible J segments. There are then nine constant regions that bind with the variable complex composed of one of each V, D, and J segments. All of the different gene segments that could be used to produce the antigen receptor create a diversity of almost 2.5 × 107 possible combinations. The mechanisms involved in making sure V(D)J recombination occurs properly by ensuring that only one of each gene segment is presented includes the RAG proteins.
The most important components of V(D)J recombination are the Recombination Signal Sequences (RSS) and the Recombination Activating Genes (RAG). The RSS appear in the ends of the DNA segments that encode for the various regions of the antigen receptor. These regions of the DNA are recognized by the RAG proteins that then cut the DNA at these points forming a double-stranded break. This double-stranded break is repaired through normal processes, and the cut ends come together to form the DNA that will code for the variable region in the antigen receptor. In the heavy chain, the DJ segments combine and then the complex combines with a V segment, while in light chains the V segment just combines with a J segment.
The RAG proteins, which are RAG-1 and RAG-2, are the mechanisms of V(D)J recombinase that are specific to lymphocyte development. Every species that undergoes V(D)J recombination has the RAG proteins. The RAG proteins function in the development of antigen receptors as follows. First they recognize and align the Recombination Signal Sequences, and it is RAG-1 job specifically to do so. The RAG complex then makes two double-stranded breaks at the 5’ ends of the RSS. The free 3’ group will then create a hairpin structure out of the DNA by attaching itself to the phosphodiester bond on the other strand. It is the job of the RAG complex to hold this new DNA structure together. RAG-1 and RAG-2 then create a single stranded break in the hairpin structure allowing the final result of the recombination of the V, D, and J segments of DNA.
The way RAG proteins work is directly related to their structure. It is known that RAG-1 is responsible for recognizing and binding to the RSS, and it is able to “recruit” the site to bind to using the nonamer-binding-domain (NBD) the RAG protein has. The nonamer is the anchor point for the anchor point for the binding of RAG and RSS complexes. After the RAG protein is anchored to the RSS, the second step of “stabilization” occurs in the presence of RAG-2 which makes the heptamer of the now combined complex available for adding an additional stabilizing site for the interaction. This machine can be compared to a portable saw because it goes right to where it needs to be to perform its function and then cuts through the strands of the DNA. The RAG proteins could also be compared to a vice because after they split the DNA they hold the newly freed end in place until it reconnects with the DNA at a different location, which the proteins lead the end of the DNA to where it is supposed to be like a shepherd.
The RAG proteins have been proven to be essential to a healthy immune system because of the detrimental effects of having a lack of them. Severe Combined Immune Deficiency (SCID) has been proven to be an effect of a lack of RAG proteins. SCID is characterized by the absence of mature B and T lymphocytes necessary for the immune system to function. Mutations in the RAG-1/RAG-2 proteins also result in a condition known as Omenn Syndrome. This is an autosomal recessive form of SCID that has the same effect of being unable to fight infections because of no functioning white blood cells being able to do the job.
In the end it is apparent that the RAG proteins are crucial to the well being of any human. Without the RAG proteins regulating the V(D)J recombination process the immune system would not have mature, diverse lymphocytes available to fight off infection. Without the “portable saw” and “vice-grip” of the RAG proteins the resulting effect would be devastating diseases such as SCID and Omenn Syndrome which significantly lower a person’s quality of life. Without the RAG proteins, human life would not be possible.



Works Cited
“Antigen Receptor Diversity”. http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/A/AgReceptorDiversity.html#V(D)J_Joining. 9 March 2006.

Posey, Jennifer E, Vicky L Brandt, David B Roth. “Paradigm switching in the germinal center”. Nature Immunology. 2004. 476-477.

Swanson, Patrick C. “Fine Structure and Activity of Discrete RAG-HMG Complexes on V(D)J Recombination Signals”. Molecular Cell Biology. March 2002. 1340-1351.

“RAG-1 and RAG-2”.
http://www.bio.davidson.edu/Courses/Immunology/Students/Spring2003/Beaghan/mfip.html. Davidson College. 2003.