A signal sequence trap based on a constitutively active cytokine receptor
A signal sequence trap based on a constitutively active cytokine receptor
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DOI：
10.1038/8666
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发表时间：
1999-05-01
影响因子：
46.9
通讯作者：
Kitamura, T
中科院分区：
文献类型：
--
作者：
Kojima, T;Kitamura, T
将分泌的和细胞表面的蛋白质靶向细胞膜是由称为信号序列的氨基酸的短的疏水延伸介导的。我们已经开发了一种方法，检测cDNA片段中的信号序列的基础上，他们的能力，重定向的细胞因子受体的组成型活性突变体的细胞表面，从而允许白细胞介素-3（IL-3）的Ba/F3细胞的非依赖性生长。在IL-3依赖性细胞中逆转录病毒介导的融合表达之后，选择在不存在IL-3的情况下生长的克隆。在中试实验中，用5 × 106病毒颗粒感染细胞，分离出150个已知的和48个新的cDNA克隆，发现所有已知的cDNA克隆都编码分泌蛋白和细胞表面蛋白。此外，我们分离的II型膜蛋白，这还没有被检测到现有的信号序列陷阱策略。
Targeting of secreted and cell-surface proteins to the cell membrane is mediated by a short hydrophobic stretch of amino acids, termed the signal sequence. We have developed a method that detects signal sequences in cDNA fragments based on their ability to redirect a constitutively active mutant of a cytokine receptor to the cell surface, thereby permitting interleukin-3 (IL-3)-independent growth of Ba/F3 cells. Retrovirus-mediated expression of the fusions in IL-3-dependent cells was followed by selection of clones for growth in the absence of IL-3. Infection of cells with 5x10(6) viral particles in a pilot experiment led to the isolation of 150 known and 48 novel cDNA clones, and all the known cDNA clones were found to encode secreted and cell-surface proteins. In addition, we isolated type II membrane proteins, which have not been detected by existing signal sequence trap strategies.