Sackstein R, Meng L, Xu X M, Chin Y H
Department of Internal Medicine, H. Lee Moffitt Cancer Center, University of South Florida College of Medicine, Tampa, USA.
Immunology. 1995 Jun;85(2):198-204.
Early investigations of lymphocyte migration in the rat operationally identified a lymphocyte membrane protein, designated 'A.11', which mediates lymphocyte adherence to lymph node (LN) high endothelial venules (HEV). To determine the primary structure of A.11 and examine its expression in lymphoid cells, we constructed an expression phage cDNA library of rat thoracic duct lymphocytes (TDL) and performed screening by immunoselection (utilizing an anti-A.11 polyclonal antiserum) as well as by hybridization selection. We have isolated a approximately 1.6 kb clone, RS-2, and sequencing revealed that it encodes rat L-selectin. The clone contains the complete coding sequence, a 105-bp 5' untranslated region and a 359-bp 3' untranslated region. Transfection of RS-2 cDNA into 70Z/3 cells conferred binding to HEV concomitant with expression of A.11, providing direct evidence that A.11 is rat L-selectin. Metabolic radiolabelling studies revealed that thymocytes synthesize markedly less L-selectin than do TDL or LN lymphocytes. However, Northern blot studies using RS-2 as a probe indicate that thymocytes possess more L-selectin RNA than does TDL. Together, these data provide evidence that post-transcriptional events contribute to regulation of L-selectin expression in thymocytes.
早期对大鼠淋巴细胞迁移的研究通过实验鉴定出一种淋巴细胞膜蛋白，命名为“A.11”，它介导淋巴细胞与淋巴结（LN）高内皮微静脉（HEV）的黏附。为了确定A.11的一级结构并检测其在淋巴细胞中的表达，我们构建了大鼠胸导管淋巴细胞（TDL）的表达噬菌体cDNA文库，并通过免疫选择（利用抗A.11多克隆抗血清）以及杂交选择进行筛选。我们分离出一个约1.6 kb的克隆RS-2，测序显示它编码大鼠L-选择素。该克隆包含完整的编码序列、一个105 bp的5'非翻译区和一个359 bp的3'非翻译区。将RS-2 cDNA转染到70Z/3细胞中，使其与HEV结合并伴随A.11的表达，这提供了直接证据表明A.11就是大鼠L-选择素。代谢性放射性标记研究表明，胸腺细胞合成的L-选择素明显少于TDL或LN淋巴细胞。然而，使用RS-2作为探针的Northern印迹研究表明，胸腺细胞比TDL拥有更多的L-选择素RNA。这些数据共同提供了证据，表明转录后事件参与了胸腺细胞中L-选择素表达的调控。
