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CPP32/Yama/Apopain/Caspase-3在不同形式的细胞凋亡途径中起核心作用。目前发现,CPP32可切割多种底物,但以CPP32为引诱蛋白用酵母双杂交系统分离与其相互作用的分子方面的研究,在国内外尚未见报道。为进一步了解肿瘤细胞信号转导途径,本文用该系统筛选与CPP32(作为引诱蛋白)相互作用的蛋白质。将CPP32引诱蛋白表达载体和肿瘤细胞文库质粒导入到酵母细胞HF7C中,用Trp-Leu-His-营养缺陷培养基初步筛选文库中插入cDNA所编码的序列与CPP32相互作用的菌落,再进一步检测β-半乳糖苷酶活性,结果从200多个Trp~+Leu~+His~+酵母菌落中获得7个LacZ~+阳性的酵母菌落。经分离阳性菌中的文库质粒和序列分析发现,在人白血病细胞株Jurkat中,新型G_sα缺失体(其正常蛋白与腺苷酸环化酶信号转导途径的激活相关)可与CPP32相互作用。纯化大肠杆菌中表达的G_sα缺失体及CPP32,用ELISA法进一步证实了这种相互作用。结论推测白血病细胞(至少某种白血病细胞)中CPP32参与了腺苷酸环化酶信号转导途径。
CPP32/Yama/Apopain/Caspase-3 plays a central role in different forms of apoptosis pathways. At present, it has been found that CPP32 can cleave a variety of substrates, but studies using CPP32 as a lure protein to separate molecules interacting with yeast two-hybrid system have not been reported at home and abroad. To further understand tumor cell signal transduction pathways, this system was used to screen for proteins that interact with CPP32 (as an attractive protein). The CPP32 induced protein expression vector and the tumor cell library plasmid were introduced into the yeast cell HF7C, and the Trp-Leu-His-auxotrophic medium was used to preliminarily screen colonies inserted in the cDNA library and the CPP32-interacting colonies, and the β was further detected. - Galactosidase activity. Results Seven LacZ+ positive yeast colonies were obtained from more than 200 Trp~+Leu~+His~+ yeast colonies. The library plasmid and sequence analysis in isolated positive bacteria revealed that in the human leukemia cell line Jurkat, a novel Gs alpha deletion body whose normal protein is associated with the activation of the adenylate cyclase signal transduction pathway can interact with CPP32. Deletion of G_sα expressed in E. coli and CPP32 were purified and further confirmed by ELISA. Conclusion It is speculated that CPP32 participates in adenylyl cyclase signal transduction pathway in leukemic cells (at least some kind of leukemia cells).