N. Yannoutsos et al., The role of recombination activating gene (RAG) reinduction in thymocyte development in vivo, J EXP MED, 194(4), 2001, pp. 471-480
Assembly of T cell receptor (TCR)alpha/beta genes by variable/diversity/joi
ning (V[D]J) rearrangement is an ordered process beginning with recombinati
on activating gene (RAG) expression and TCR beta recombination in CD4(-)CD8
(-)CD25(+) thymocytes. In these cells, TCR beta expression leads to clonal
expansion, RAG downregulation, and TCR beta allelic exclusion. At the subse
quent CD4(+)CD8(+) stage, RAG expression is reinduced and V(D)J recombinati
on is initiated at the TCR alpha locus. This second wave of RAG expression
is terminated upon expression of a positively selected alpha/beta TCR. To e
xamine the physiologic role of the second wave of RAG expression, we analyz
ed mice that cannot reinduce RAG expression in CD4(+)CD8(+) T cells because
the transgenic locus that directs RAG1 and RAG2 expression in these mice i
s missing a distal regulatory element essential for reinduction. In the abs
ence of RAG reinduction we find normal numbers of CD4(+)CD8(+) cells but a
50-70% reduction in the number of mature CD4(+)CD8(-) and CD4(-)CD8(+) thym
ocytes. TCR alpha rearrangement is restricted to the 5' end of the J alpha
cluster and there is little apparent secondary TCR alpha recombination. Com
parison of the TCR alpha genes expressed in wild-type or mutant mice shows
that 65% of all alpha/beta T cells carry receptors that are normally assemb
led by secondary TCR alpha rearrangement. We conclude that RAG reinduction
in CD4(+)CD8(+) thymocytes is not required for initial TCR alpha recombinat
ion but is essential for secondary TCR alpha recombination and that the maj
ority of TCR alpha chains expressed in mature T cells are products of secon
dary recombination.