Giorda R, Chambers W H, Dahl C A, Trucco M
Pittsburgh Cancer Institute, Pa.
Nat Immun Cell Growth Regul. 1990;9(2):91-102.
In this report, we describe the transcription of messenger ribonucleic acid (mRNA) specific for the core protein of chondroitin sulfate proteoglycan (CSPG), the entire sequence of the message for CSPG core protein and the presence of CSPG in the granules of a highly purified population of recombinant interleukin-2 (rIL-2)-stimulated rat natural killer (NK), i.e. adherent lymphokine-activated Killer (A-LAK), cells. The presence of CSPG in A-LAK cell granules was demonstrated by a variety of biochemical and immunologic methods. Further, we have demonstrated the presence of a 1.1-kilobase (kb) transcript for the core protein of CSPG by Northern blot analysis using a specific probe (pPG6) derived from a rat yolk sac tumor cell line (L-2). Three cell types that contained CSPG in granules produced a transcript of 1.1 kb, whereas L-2 cells, which localize CSPG on the cell surface, produced a transcript of 1.3 kb. A complementary DNA (cDNA) library was prepared from rat A-LAK cells and the gene for the CSPG core protein was cloned. From approximately 1.2 X 10(5) recombinant phages, 5 positive clones were obtained. The longest clone, PG-NK-5, was sequenced in its entirety and it was found to encode the entire sequence of the CSPG core protein. The other 4 clones were partially sequenced and were identical to PG-NK-5. Comparison of the sequence of PG-NK-5 with pPG6 indicated that they were nearly identical. However, all NK-cell-derived cDNAs contained a poly(A) tail that started 20 basepairs upstream from other published sequences for CSPG core proteins. These data represent the first description of the sequence of the core protein of CSPG contained in NK cells.
在本报告中，我们描述了硫酸软骨素蛋白聚糖（CSPG）核心蛋白特异性信使核糖核酸（mRNA）的转录、CSPG核心蛋白信息的完整序列以及CSPG在高度纯化的重组白细胞介素-2（rIL-2）刺激的大鼠自然杀伤（NK）细胞（即黏附性淋巴因子激活的杀伤细胞（A-LAK））颗粒中的存在情况。通过多种生化和免疫方法证实了A-LAK细胞颗粒中存在CSPG。此外，我们使用源自大鼠卵黄囊肿瘤细胞系（L-2）的特异性探针（pPG6），通过Northern印迹分析证明了存在一条1.1千碱基（kb）的CSPG核心蛋白转录本。三种在颗粒中含有CSPG的细胞类型产生了1.1 kb的转录本，而将CSPG定位在细胞表面的L-2细胞产生了1.3 kb的转录本。从大鼠A-LAK细胞制备了互补DNA（cDNA）文库，并克隆了CSPG核心蛋白的基因。从大约1.2×10⁵个重组噬菌体中获得了5个阳性克隆。最长的克隆PG-NK-5进行了全序列测定，发现它编码CSPG核心蛋白的完整序列。其他4个克隆进行了部分测序，与PG-NK-5相同。将PG-NK-5的序列与pPG6进行比较，发现它们几乎相同。然而，所有NK细胞来源的士DNA都含有一个多聚腺苷酸（poly(A)）尾巴，该尾巴起始于比其他已发表的CSPG核心蛋白序列上游20个碱基对处。这些数据首次描述了NK细胞中所含CSPG核心蛋白的序列。
