Sleckman B P, Bassing C H, Hughes M M, Okada A, D'Auteuil M, Wehrly T D, Woodman B B, Davidson L, Chen J, Alt F W
Howard Hughes Medical Institute, Children's Hospital, Harvard Medical School and Center for Blood Research, Boston, MA 02115, USA.
Proc Natl Acad Sci U S A. 2000 Jul 5;97(14):7975-80. doi: 10.1073/pnas.130190597.
T cell receptor (TCR) beta variable region genes are assembled in progenitor T cells from germ-line Vbeta, Dbeta, and Jbeta segments via an ordered two-step process in which Dbeta to Jbeta rearrangements occur on both alleles before appendage of a Vbeta to a preexisting DJbeta complex. Direct joining of Vbeta segments to nonrearranged Dbeta or Jbeta segments, while compatible with known restrictions on the V(D)J recombination mechanism, are infrequent within the endogenous TCRbeta locus. We have analyzed mechanisms that mediate ordered Vbeta, Dbeta, and Jbeta assembly via an approach in which TCRbeta minilocus recombination substrates were introduced into embryonic stem cells and then analyzed for rearrangement in normal thymocytes by recombinase-activating gene 2-deficient blastocyst complementation. These analyses demonstrated that Vbeta segments are preferentially targeted for rearrangement to Dbeta as opposed to Jbeta segments. In addition, we further demonstrated that Vbeta segments can be appended to nonrearranged endogenous Dbeta segments in which we have eliminated the ability of Dbeta segments to join to Jbeta segments. Our findings are discussed in the context of the mechanisms that regulate the ordered assembly and utilization of V, D, and J segments.
T细胞受体（TCR）β可变区基因在祖T细胞中通过一个有序的两步过程，由种系Vβ、Dβ和Jβ片段组装而成，在此过程中，两个等位基因上的Dβ到Jβ重排在Vβ附加到预先存在的DJβ复合体之前发生。Vβ片段直接连接到未重排的Dβ或Jβ片段，虽然符合对V（D）J重组机制的已知限制，但在内源TCRβ基因座中并不常见。我们通过一种方法分析了介导有序Vβ、Dβ和Jβ组装的机制，即将TCRβ微基因座重组底物导入胚胎干细胞，然后通过重组酶激活基因2缺陷型囊胚互补分析正常胸腺细胞中的重排。这些分析表明，与Jβ片段相比，Vβ片段优先被靶向重排到Dβ。此外，我们进一步证明，Vβ片段可以附加到未重排的内源性Dβ片段上，在这些片段中我们已经消除了Dβ片段与Jβ片段连接的能力。我们的研究结果将在调节V、D和J片段有序组装和利用的机制背景下进行讨论。
