The sequence NPFXD defines a new class of endocytosis signal in Saccharomyces cerevisiae.
The sequence NPFXD defines a new class of endocytosis signal in Saccharomyces cerevisiae.
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DOI：
10.1083/jcb.135.6.1789
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发表时间：
1996-12
期刊：
影响因子：
--
通讯作者：
Payne GS
中科院分区：
文献类型：
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作者：
Tan PK;Howard JP;Payne GS
酵母膜蛋白Kex2p在细胞质结构域中使用一个含有酪氨酸的基序来定位到高尔基体晚期隔室。由于酵母中错位到质膜上的高尔基体膜蛋白可以发生内吞作用，我们研究了高尔基体定位序列或Kex2p胞质结构域中的其他序列是否介导了内吞作用。为了评估内吞功能，Kex2p胞浆结构域被融合到内吞缺陷形式的α-因子受体上。第2步。与完整的Ste2p一样，嵌合蛋白Stex22p也经历了依赖于笼蛋白和End3p的快速内吞。Stex22p的摄取不需要Kex2p高尔基定位基序。相反，位于COOH末端37个氨基酸的NPFSD序列对Stex22p的内吞作用是必不可少的。当N、P或F残基被转化为丙氨酸时，内化作用被取消，而当D转化为A时，NPFSD被严重损害。当添加到野生型Ste2p中缺乏赖氨酸内吞信号的内吞缺陷的Ste2p嵌合体的COOH末端时，NPFSD恢复了摄取。NPF序列存在于a因子受体Ste3p的细胞质结构域中。该序列的突变阻止了信息素刺激的截短形式的Ste3p的内吞作用。我们的结果证实NPFSD是一种依赖于笼蛋白的内吞信号，不同于酵母中含有芳香族氨基酸的高尔基体定位基序和基于赖氨酸的泛素依赖的内吞信号。
The yeast membrane protein Kex2p uses a tyrosine-containing motif within the cytoplasmic domain for localization to a late Golgi compartment. Because Golgi membrane proteins mislocalized to the plasma membrane in yeast can undergo endocytosis, we examined whether the Golgi localization sequence or other sequences in the Kex2p cytoplasmic domain mediate endocytosis. To assess endocytic function, the Kex2p cytoplasmic domain was fused to an endocytosis-defective form of the alpha-factor receptor. Ste2p. Like intact Ste2p, the chimeric protein, Stex22p, undergoes rapid endocytosis that is dependent on clathrin and End3p. Uptake of Stex22p does not require the Kex2p Golgi localization motif. Instead, the sequence NPFSD, located 37 amino acids from the COOH terminus, is essential for Stex22p endocytosis. Internalization was abolished when the N, P, or F residues were converted to alanine and severely impaired upon conversion of D to A. NPFSD restored uptake when added to the COOH terminus of an endocytosis-defective Ste2p chimera lacking lysine-based endocytosis signals present in wild-type Ste2p. An NPF sequence is present in the cytoplasmic domain of the a- factor receptor, Ste3p. Mutation of this sequence prevented pheromone- stimulated endocytosis of a truncated form of Ste3p. Our results identify NPFSD as a clathrin-dependent endocytosis signal that is distinct from the aromatic amino acid-containing Golgi localization motif and lysine-based, ubiquitin-dependent endocytosis signals in yeast.