Analysis of the expression and function of T cell differentiation antigen-CD7

1.小鼠CD7基因的分子克隆：利用已报道的小鼠CD7基因，从BALB/c小鼠脾基因组文库中分离到4个阳性克隆(Yoshikawa，1993)。这些克隆全长24.4kb，包含4个外显子，内含子中含有编码小鼠CD7抗原的内含子。第一外显子对应于5‘非翻译区和信号肽，第二外显子对应于免疫球蛋白样区，第三外显子对应于胞外区和跨膜部分，第四外显子对应于细胞质尾部和3’UT。与人CD7启动子区域相比，发现了9条相同的核苷酸序列和一个GC盒。T细胞特异性表达也可能由小鼠TCR-V或Thy1基因5‘侧翼区的一串核苷酸序列决定。2.抗小鼠CD7抗体的制备：用两种合成肽(胞外区分别为21和19个氨基酸)和小鼠CD7基因和基因组DNA转基因细胞免疫兔和大鼠。3.转基因小鼠的制备：利用Yoshikawa，1991报道的人CD7基因编码基因的2个构建体，获得了7株人CD7基因转基因小鼠。除1株有CD7抗原表达外，其余6株均未见表达。

1.Molecular cloning of mouse Cd7 gene : Four positive clones from BALB/c mouse spleen genomic library were isolated using mouse CD7 cDNA previously reported (Yoshikawa, 1993). These clones covered 24.4kb containing four exons with intervening introns coding for the mouse CD7 antigen. The first exon correspondens to a 5'untranslated (UT) region and the signal peptide, the second exon to the immunoglobulin like region, the third exon to the part of extracellular domain with the transmembrane portion, and the fourth exon to the cytoplasmic tail and 3'UT.When compared to the human CD7 promoter region, nine streches of identical nucleotide sequences as well as a GC box were found. T cell specific expresson may be also defined by streches of nucleotide sequence in 5' flanking regions of either Tcr-V or Thy1 genes in the mouse. We are now inthe process of producing tarageted Cd7 gene disruption mouse to analyze the function of the antigen.2.Production of anti mouse CD7 antibody : Two synthetic peptides (21 and 19 amino acids in the extracellular domain), and mouse CD7 cDNA-and genomic DNA-transfected cells were used for immunization of rabbits and rats. No antibody reactive to mouse thymocytes was, however, obtained to date.3.Production of transgenic mouse : Using 2 constructs from the gene coding for human CD7 antigen reported previously (Yoshikawa, 1991), 7strains of human CD7 gene transgenic mouse were obtained. In only one strain, very week expression of CD7 antigen was observed, while in the rest of 6 strains, the expression was not observed.