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

1.小鼠CD 7基因的分子克隆：使用先前报道的小鼠CD 7 cDNA（Yoshikawa，1993）从BALB/c小鼠脾基因组文库中分离出4个阳性克隆。这些克隆覆盖24.4kb，含有编码小鼠CD 7抗原的四个外显子和插入的内含子。第一外显子对应于5 '非翻译（UT）区和信号肽，第二外显子对应于免疫球蛋白样区，第三外显子对应于具有跨膜部分的胞外结构域的部分，并且第四外显子对应于胞质尾区和3' UT。发现了9段相同的核苷酸序列和一个GC盒，T细胞特异性表达也可由Tcr-1或Tcr-2的5 ′侧翼区的核苷酸序列确定。V或Thy 1基因。2.抗小鼠CD 7抗体的制备：用两种人工合成的多肽（胞外区21和19个氨基酸）和小鼠CD 7 cDNA及基因组DNA转染的细胞免疫家兔和大鼠。3.转基因小鼠的制备：利用Yoshikawa，1991报道的编码人CD 7抗原基因的2个构建体，获得了7株人CD 7基因转基因小鼠。仅在1株中观察到CD 7抗原的极弱表达，而在其余6株中未观察到CD 7抗原的表达。

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.