RAG1: Viral or TE?

Today’s post is another article summary but about something a bit different. This one concerns the origins of adaptive immunity in jawed vertebrates, and more specifically the origin of the RAG1 protein. RAG1 is a gene expressed in T and B-lymphocytes in the immune system which along with RAG2 mediates the creation antibodies. It does this via a process known as V(D)J recombination, after the variable and joining sequence segments that are joined together to create a complete antibody gene. The RAG1/2 complex binds to recognition signal sequences (RSS) between segments and cuts out the intervening DNA. The subsequent double-strand breaks are joined back together via non-homologous end-joining proteins to join the previously separated sequence into a single coding region. This is an oversimplification of the process but basically this occurs several times in a somewhat random manner to create a novel antibody gene. Through a selection process that occurs in the immune system only cells with antibodies which recognize foreign antigens survive to prevent autoimmunity.

Researchers noticed some time ago that the process of V(D)J recombination was very similar to what occurs during the transposition of a Class II TE, also known as a DNA transposon. A transposase protein recognizes and binds to the inverted terminal repeats (ITR) on either end of the element and cuts it out of the sequence it is inserted into. In a similar manner RAG1/2 complexes bind to a pair of RSS’s, cut out the sequence in between and circularize it for degradation by the cell. In 2005 Kapitonov and Jurka published a paper showing that roughly 60 amino acids in the C-terminus of RAG1, within the so-called “RAG core”, were found to be significantly similar to the Transib transposases. As well, the ITRs of Transib elements are very similar to the RSS’s (with a probability of the similarity occurring by chance of 1.0 x 10^-3) and both RAG complexes and Transib elements generate 5 bp target site duplications (TSD). It seemed like the origin of RAG1 had been at least partially solved.

Several weeks ago a paper popped up in my iGoogle Reader that has been published in PLoS One contesting the transposon origin of RAG. This seemed odd to me because I thought the conclusions of Kapitonov and Jurka (2005) were pretty sound. The paper by Dreyfus (2009) claimed that there was more evidence that RAG1 was derived from the insertion of a herpes virus-like element long ago in a deuterostome ancestor. You can judge for yourself, the paper is available free to download here, but I’d like to tell you why this new hypothesis doesn’t hold up as well to me as the Transib origin one does.

Dreyfus states that the regulatory sequences of the Epstein-Barr Virus (EBV), a herpes virus, are very similar, identical in some cases, to the regulatory sequences possessed by RAG1. He also cites the fact that EBV infections activate the expression of RAG1/2 and that when the virus excises from host DNA it forms a circle, much in the same way intervening sequences form circular intermediates during V(D)J recombination. And both RAG1 and the ICP-8 protein of EBV have DDE metal ion binding motifs important for cleavage of target DNA despite the fact they do not possess any significant primary sequence identity. Dreyfus says that despite the sequence similarity between the RAG1 core and Transib transposases there is the problem of the RAG1 amino terminus which is not similar. He also states that during the evolution of the proto-RAG1 there would be no selective benefit to the expression of a Transib element in proto-immune cells and that the antigenic effects of some herpes-virus proteins would provide a reason for the selective maintenance of an inserted herpes-like element. Also, no autonomous “RAG transposon” has been found in nature yet and neither has an element been found encoding a RAG2 homolog which is required for V(D)J recombination to occur.

The regulatory similarities between EBV and RAG1 are interesting but I think the answer as to the probable origin of RAG is pretty clear. The fact that both ICP-8 and RAG1 possess the DDE catalytic motif is not evidence of common ancestry as this motif evolved multiple times independently in several DNA transposon superfamilies, including Transib, and in proteins like RNase H and the Argonaute component of RISC. DNA transposons forming circles as intermediates is not unknown and a good example I read of recently are the TEs in ciliates that seem to be involved in the drastic chromosomal rearrangement of those species. Dreyfus did not put any sequence alignment diagrams in his paper because ICP-8 and RAG1 could not be aligned while Kapitonov and Jurka (2005) were able to align the Transib transposase and RAG1 and show their significant similarity. The so-called “ amino terminus problem” really isn’t a problem at all because last year Panchin and Moroz (2008) published a paper where they found a Chapaev superfamily DNA transposon, N-RAG, from the mollusc Aplysia californica whose transposase had significant similarity to the “ problematic” N terminus of RAG1. This paper is not cited by Dreyfus. It turns out RAG1 might actually be derived from two separate DNA transposons, not just one. As for the maintenance problem because TEs are selfish and can proliferate even at a fitness cost to the host there really is no need to explain why the proto-RAG1 transposon could persist.

The story is not over for RAG1 though. I agree with Dreyfus that the issue of where the RAG2 gene came from needs to be addressed as well as trying to flesh exactly how the evolutionary process proceeded from an autonomous DNA transposon to modern day RAG.

Dreyfus, D.H. 2009. Paleo-immunology: evidence consistent with insertion of a primordial herpes virus-like element in the origins of acquired immunity. PLoS One 4: e5778.

Kapitonov, V.V. and J. Jurka. 2005. RAG1 core and V(D)J recombination signal sequences were derived from Transib transposons. PLoS Biology 3: e181.

Klobutcher, L.A. and G. Herrick. 1997. Developmental Genome Reorganization in Ciliated Protozoa: The Transposon Link. In Progress in Nucleic Acid Research and Molecular Biology (eds. E. Cohn and K. Moldave), pp. 1-62. Academic Press.

Panchin, Y. and L.L. Moroz. 2008. Molluscan mobile elements similar to the vertebrate Recombination-Activating Genes. Biochemical and Biophysical Research Communications 369: 818-823.