用酵母双杂交系统筛选与PEN-2蛋白胞内端相互作用的蛋白

Identifying PEN-2-interacting Proteins by Yeast Two-hybrid

γ-分泌酶复合体对β-淀粉蛋白前体蛋白(Beta-amyloid precursor protein,βAPP)的水解是生成β-淀粉蛋白(Aβ)的关键步骤,在阿尔茨海默氏症(Alzhei mer′s disease,AD)的病理发生中起着重要作用.PEN-2蛋白(Presenilin enhancerprotein 2)是γ-分泌酶的一个重要组分,在γ-分泌酶的组装和成熟中扮演重要的角色.此外,PEN-2还具有促进神经细胞存活和防止细胞凋亡的作用.为了进一步研究PEN-2的生理功能,以PEN-2为诱饵蛋白用酵母双杂交系统筛选与之有相互作用的蛋白,发现PCBP4和GRK5肽段可结合PEN-2,并在酵母体系用β-半乳糖苷酶报告基因验证了它们的相互作用.这些结果表明PCBP4和GRK5可能影响PEN-2的正常生理功能,并且有可能在阿尔茨海默氏症的形成中起着一定的作用.

Presenilin enhancer protein 2 （PEN-2） is an essential component of the γ-secretase complex which plays an important role in the pathogenesis of Alzheimer＇s disease （AD）. In addition to PEN-2 ,the γ-secretase complex consists of other components in- cluding presenilin （PS）,nicastrin（NCT） and Anterior pharynx defectivel（APH-1）. PEN-2 has been demonstrated to be crucial for the endoproteolysis of the full-length PS to generate its N-terminal fragments and C-terminal fragments, which keep associated and interact with PEN-2 and other γ-secretase components. Moreover,PEN-2 is involved in promoting neuronal cell survival and protecting neurons from apoptosis in vivo. To further understand the function of PEN-2 in the pathology of AD,we utilized PEN-2 as a bait to identify PEN-2 interacting proteins in a yeast two-hybrid system. Two candidate proteins were identified： Homo sapiens poly （rC） binding protein 4 （PCBP4） and Homo sapiens G protein-coupled receptor kinase 5 （GRK5）. The β-galactosidase activity assay demonstrated that the two proteins can interact with PEN-2 in yeast strain AH109. These results suggest that PCBP4 and GRK5 may play potential roles in facilitating the physiological functions of PEN-2.