Fabbrini M S, Carpani D, Soria M R, Ceriotti A
Department of Biological and Technological Research-Dibit, San Raffaele Scientific Institute, 20132 Milano, Italy.
FASEB J. 2000 Feb;14(2):391-8. doi: 10.1096/fasebj.14.2.391.
In this work, we have devised an intracellular immunization strategy for the expression in high amounts of ATF-saporin, a targeted chimeric toxin constituted by the ATF receptor binding domain of human urokinase and the plant ribosome-inactivating protein saporin, which has been shown to be highly cytotoxic to target cells. This strategy may allow the production of highly toxic secretory proteins in eukaryotic cells, avoiding cell suicide caused by autointoxication. The procedure consists of equipping host cells with cytosolic neutralizing antibodies directed toward the toxic domain of the heterologous polypeptide. We show that this intracellular immunization is essential for the synthesis of correctly folded, biologically active ATF-SAP in the high amounts needed to investigate its in vivo anti-metastatic potential. Such a strategy should be generally useful for the production of toxic molecules of therapeutic value whose folding and maturation require transit through the eukaryotic secretory pathway. Fabbrini, M. S., Carpani, D., Soria, M. R., Ceriotti, A. Cytosolic immunization allows the expression of preATF-saporin chimeric toxin in eukaryotic cells.
在这项工作中，我们设计了一种细胞内免疫策略，用于大量表达ATF-皂草素，这是一种靶向嵌合毒素，由人尿激酶的ATF受体结合域和植物核糖体失活蛋白皂草素组成，已证明对靶细胞具有高度细胞毒性。该策略可能允许在真核细胞中产生高毒性分泌蛋白，避免自中毒引起的细胞自杀。该程序包括为宿主细胞配备针对异源多肽毒性结构域的胞质中和抗体。我们表明，这种细胞内免疫对于合成正确折叠、具有生物活性的ATF-SAP至关重要，而这是研究其体内抗转移潜力所需的大量蛋白质。这种策略对于生产其折叠和成熟需要通过真核分泌途径的具有治疗价值的毒性分子通常应该是有用的。法布里尼，M.S.，卡尔帕尼，D.，索里亚，M.R.，切里奥蒂，A. 胞质免疫允许preATF-皂草素嵌合毒素在真核细胞中表达。
