SPLICING OF EFFECTOR CELL PROTEASE RECEPTOR-1 MESSENGER-RNA IS MODULATED BY AN UNUSUAL RETAINED INTRON
SPLICING OF EFFECTOR CELL PROTEASE RECEPTOR-1 MESSENGER-RNA IS MODULATED BY AN UNUSUAL RETAINED INTRON
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DOI：
10.1021/bi00250a039
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发表时间：
1994-11-22
期刊：
影响因子：
2.9
通讯作者：
ALTIERI, DC
中科院分区：
文献类型：
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作者：
ALTIERI, DC
效应细胞蛋白酶受体-1 （EPR-1）是Xa因子的跨膜糖蛋白受体，参与蛋白水解活性和白细胞有丝分裂的细胞表面组装。现在表明EPR-1的膜表达是由mRNA剪接动态调节的。通过聚合酶链反应扩增和核糖核酸酶保护研究，对表达epr -1的细胞和基因工程转染物的北杂交分析表明，这种机制涉及去除70-90%成熟mRNA中保留的451 bp的中间序列。通过在某些白血病B淋巴细胞和单核细胞中EPR-1的组成膜过表达来判断，中间序列的剪接以细胞类型特异性的方式发生。此外，对功能剪接或未剪接的EPR-1构建物稳定转染的细胞系的表型分析表明，内含子顺式作用序列可能在剪接调节中发挥作用。代替因子Xa的跨膜受体（EPR-1a），最普遍的未剪接EPR-1转录物产生一个新的110个氨基酸的截断蛋白（EPR-1b），其中一个独特的内含子编码的-COOH末端携带潜在的核靶向信号PPQHRAKS。一种针对EPR-1b内含子编码序列产生的抗体在渗透细胞的间接免疫荧光染色中显示出这种变异亚型的显著核定位。这些发现为基于高效率内含子保留调节因子依赖的细胞效应功能的新机制提供了证据。
Effector cell protease receptor-1 (EPR-1) is a transmembrane glycoprotein receptor for factor Xa that contributes to cell surface assembly of proteolytic activities and leukocyte mitogenesis. It is now shown that membrane expression of EPR-1 is dynamically modulated by mRNA splicing. Northern hybridization analysis of EPR-1-expressing cells and genetically engineered transfectants demonstrates that this mechanism involves removal of a 451 bp intervening sequence retained in 70-90% of mature mRNA, as quantitated by polymerase chain reaction amplification and ribonuclease protection studies. Splicing of the intervening sequence occurs in a cell type-specific fashion, as judged by the constitutive membrane overexpression of EPR-1 in certain leukemic B lymphocytes and monocytic cells. Furthermore, phenotypic analysis of cell lines stably transfected with functionally spliced or unspliced EPR-1 constructs suggests a potential role of intron cis-acting sequence(s) in splicing regulation. Instead of a transmembrane receptor for factor Xa (EPR-1a), the most prevalent unspliced EPR-1 transcript generates a novel truncated protein of 110 amino acids (EPR-1b), in which a unique intron-encoded -COOH terminus carries a potential nuclear targeting signal PPQHRAKS. An antibody generated against the intron-encoded sequence of EPR-1b demonstrates prominent nuclear localization of this variant isoform in indirect immunofluorescence staining of permeabilized cells. These findings provide evidence for a novel mechanism based on high efficiency intron retention modulating factor Xa-dependent cellular effector functions.