ESTABLISHMENT OF MOUSE-CELL LINES WHICH CONSTITUTIVELY SECRETE LARGE QUANTITIES OF INTERLEUKIN-2, INTERLEUKIN-3, INTERLEUKIN-4 OR INTERLEUKIN-5, USING MODIFIED CDNA EXPRESSION VECTORS
ESTABLISHMENT OF MOUSE-CELL LINES WHICH CONSTITUTIVELY SECRETE LARGE QUANTITIES OF INTERLEUKIN-2, INTERLEUKIN-3, INTERLEUKIN-4 OR INTERLEUKIN-5, USING MODIFIED CDNA EXPRESSION VECTORS
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DOI：
10.1002/eji.1830180115
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发表时间：
1988-01-01
影响因子：
5.4
通讯作者：
MELCHERS, F
中科院分区：
文献类型：
--
作者：
KARASUYAMA, H;MELCHERS, F
已经建立了不同谱系的小鼠细胞系，它们能结构性地分泌大量重组小鼠白介素mIL-2、mIL-3、mIL-4或mIL-5。对现有的牛乳头瘤病毒表达载体PBV-1MTHA进行了修饰，使转化的X63Ag8-653骨髓瘤细胞、NIH3T3成纤维细胞和C127乳腺肿瘤细胞能够稳定携带多个拷贝的载体，并能稳定表达编码单个白介素亚单位的插入片段，并能大量分泌白介素1。转化mIL-2基因的细胞系稳定携带30-100个拷贝/细胞的血浆，并能分泌具有生物活性的mIL-2，其分泌数量与小鼠EL4胸腺瘤细胞或大鼠脾细胞在有丝分裂原刺激下产生的数量相似。从mIL-2基因的3‘端非翻译区中删除富含AT的序列，使转化子合成和分泌mIL-2的能力提高了100倍。添加重金属可使产量进一步提高2~6倍。转化3‘’缺失mIL-3基因的细胞分泌mIL-3活性是粒单核细胞白血病细胞系WEHI3的300-1000倍。类似转化子产生的mIL4可诱导T细胞系、肥大细胞系和B白血病细胞摄取(~3H)胸腺嘧啶核苷，并促进B细胞产生IgG1。IL-4滴度是刀豆蛋白A刺激的T细胞系的150倍，类似的转化子分泌的IL-5的滴度是刀豆蛋白A刺激的2.19 T细胞的10倍，以B细胞白血病BCL1的增殖和成熟来判断2.19 T细胞的增殖和成熟。该表达载体可用于建立真核细胞系，以较高的速率从Lentgth全长克隆中产生蛋白质。已建立的高水平分泌IL-2、IL-3、IL-4、IL-5的细胞系有望成为研究IL-2、IL-3、IL-4、IL-5在免疫系统中功能的有用来源。
Mouse cells lines of different lineages have been established which constitutively secrete large quantities of recombinant mouse interleukins (mIL 2, mIL 3, mIL4 or mIL 5). An existing bovine papilloma virus-based expression vector, pBV-1MTHA, was modified to allow transformed X63Ag8-653 myeloma cells, NIH 3T3 fibroblasts and C127 mammary tumor cells to stably carry multiple copies of the vector, to express the inserted cDNA encoding a single interleukin constitiutively, and to secrete the interleukin in high quantities. Cell lines transformed with mIL 2 cDNA stably carried 30-100 copies of the plasmind per cell and constitutively secreted biologically active mIL 2 in qunatities similar to those produced by murine EL4 thymoma cells or rat spleen cells stimulated with mitogens. Delection of the 3'' untranslated region containing AT-rich sequences from the mIL 2 cDNA resulted in a 100-fold increase in the constitutive production and secretion of mIL 2 by the transformants. Addition of a heavy metal further increased the production 2 to 6-fold. Cells transformed with 3''deleted mIL3 cDNA constitutively secreted 300-1000 times higher activites of mIL 3 than the myelomonocytic leukemia line WEHI3. mIL4 produced by the similar transformants induced (3H)thymidine uptake of a T cell line, a mast cell line and B leukemia cells, and enhanced the production of IgG1 by B cells. IL 4 titers were 150 times higher than those produced by the concanavalin A- stimulated T cell line 2.19 mIL 5 was secreted by similar transformants at 10-fold higher titers than those produced by concanavalin A-stimulated 2.19 T cells, as judged by the proliferation and maturation of B cell leukemia lated 2.19 T cells, as judged by the proliferation and maturation of B cell leukemia BCL1. The expression vectors should be useful in establishing eukaryotic cell lines producing proteins from full lentgth cDNA clones at higher rates. The established cell lines secreting IL 2, 3, 4 or 5 at high rate should be useful sources of herse interleukins in the investigation of their function in the immune system.