Identification of a chromosomal region that is responsible for memory T cell homeostasis in a mutant mouse.

众所周知，与幼稚T细胞相比，记忆T细胞增殖更快，对病原体的反应更强。我在科罗拉多州丹佛市国家犹太医学中心的SPF群体中发现了一只雌性突变小鼠，携带着过量的CD4+和CD8+记忆T细胞。表型遗传为常染色体隐性遗传。胸腺体积较小(约为对照组的10%)，双阳性细胞数量急剧减少。T细胞分化在一个称为DN3的步骤中被部分但主要地阻断，其中前TCR信号是必要的。血清中的血小板显着升高，这一结果与脾中巨核细胞的过量相一致。此外，在大的Preb期，未成熟的B细胞数量显著减少，B细胞的分化受阻，这是前BCR信号的重要组成部分。我进行了骨髓移植实验，以分析淋巴祖细胞的突变是否存在，或者微环境中的突变是否对突变小鼠的T和B细胞表型是必需的。淋巴祖细胞本身的突变是观察表型所必需的。为了确定携带诱导突变小鼠表型的负责基因的DNA片段，我们采用了SSPL方法。我们将B10#4小鼠与NZB小鼠杂交，得到F2小鼠，获得了100多只表型为+和-的后代。根据NCBI数据库，我们在2号染色体上发现了与该突变体有关的1 cM DNA片段，该片段含有约40个基因。该片段中的一个基因的RT-PCR实验结果令人着迷，因为该基因的剪接模式在B10#4和对照组小鼠中是不同的。我分析了该基因的序列，发现该基因的内含子有两个突变。

It is well known that memory T cells proliferate faster and react to the pathogens stronger compared to naive T cells. I found a female mutant mouse carrying excess amount of memory T cells of both CD4+ and CD8+ in a SPF colony in National Jewish Medical Center, Denver, CO. The phenotype is inherited as an autosomal recessive manor. Thymus size is small (about 10% of controls) and the numbers of double positive cells reduces dramatically. T cell differentiation is partially but dominantly blocked at a step where preTCR signaling is necessary named DN3. Platelets significantly increased in serum and this result is consistent with excess amount of megakaryocytes in spleen. In addition, the number of immature B cells decreased significantly and differentiation of B cells is blocked at large preB stage where preBCR signaling is important. I performed bone marrow transplantation experiment to analyze if the mutation in lymphoid progenitor is or if it in microenvironment is necessary for the phenotype of T and B cells in the mutant mice. The mutation in lymphoid progenitor cells themselves is necessary to see the phenotype. In order to identify a DNA fragment carrying a responsible gene that induces the phenotype in the mutant mice, we employed SSPL method. We crossed B10#4 mice with NZB mice to get F2 mice and got over 100 phenotype + and -offspring. We identified 1cM DNA fragment in chromosome #2 that is responsible for the mutant and the fragment has about 40 genes according to NCBI database. Result of RT-PCR experiment for a gene in the fragment was fascinating, since splicing pattern of the gene is different between the B10#4 and control mice. I analyzed sequence of the gene and found that there were two mutations in the introns of the gene.