P. Greenhalgh et al., CHARACTERIZATION AND EXPRESSION OF RECOMBINATION ACTIVATING GENES (RAG-1 AND RAG-2) IN XENOPUS-LAEVIS, The Journal of immunology, 151(6), 1993, pp. 3100-3110
The primary repertoire of B and T cells is established by V(D)J recomb
ination. Two closely linked genes, RAG-1 and RAG-2, are essential for
this process, and have been identified in mice, humans, and chickens.
To study lymphocyte development in Xenopus laevis, we have characteriz
ed RAG-1 and RAG-2 in this species and examined their patterns of expr
ession. Degenerate oligonucleotides, based on the known highly conserv
ed RAG-1 sequences, were used to amplify, by the polymerase chain reac
tion, a segment of Xenopus RAG-1 from genomic DNA. A product of expect
ed size was obtained and used to identify a genomic clone that contain
ed the complete coding region of RAG-1 (1045 codons), and approximatel
y the 3'- half of the coding region of RAG-2. The coding regions of RA
G-1 and RAG-2 each lie on a single exon, are in opposite transcription
al orientation, and are separated by approximately 6 kb. The sequence
of the remainder of RAG-2 was determined by PCR amplification of genom
ic DNA, with primers based on sequence analysis of RAG-2 cDNA clones.
The predicted Xenopus RAG-1 protein is 71% identical in amino acid seq
uence to the sequences of each of the mouse, human, and chicken protei
ns; from position 392 to 1012 the identity is 88%. The coding region o
f Xenopus RAG-2 (520 codons) is somewhat less conserved among the diff
erent species. Tissue-specific expression of Xenopus RAG-1 and RAG-2 w
as examined both by Northern blotting and by a reverse transcription-p
olymerase chain reaction assay. In juvenile frogs, the highest levels
of RAG-1 and RAG-2 expression were observed in the thymus, with lower
levels in liver and spleen, and even lower levels in the kidneys. In a
dults, the thymus and bone marrow were found to be the principal sites
of expression of both genes. RAG-2, but not RAG-1, was expressed in o
ocytes.