Rusk Nicole, Le Phuong U, Mariggio Stefania, Guay Ginette, Lurisci Cristiano, Nabi Ivan R, Corda Daniela, Symons Marc
Center for Oncology and Cell Biology, North Shore-LIJ Research Institute, 350 Community Drive, Manhasset, NY 11030, USA.
Curr Biol. 2003 Apr 15;13(8):659-63. doi: 10.1016/s0960-9822(03)00241-0.
Synaptojanin 2 is a ubiquitously expressed polyphosphoinositide phosphatase that displays a high degree of homology in its catalytic domains with synaptojanin 1 [1,2]. Neurons of synaptojanin 1-deficient mice display an increase in clathrin-coated vesicles and delayed reentry of recycling vesicles into the fusion-competent vesicle pool, but no defects in early steps of endocytosis [3,4]. Here we show that inhibition of synaptojanin 2 expression via small interfering (si) RNA causes a strong defect in clathrin-mediated receptor internalization in a lung carcinoma cell line. This inhibitory phenotype is rescued by overexpression of wild-type synaptojanin 2, but not of wild-type synaptojanin 1 or mutant synaptojanin 2 that is deficient in 5'-phosphatase activity. In addition, electron-microscopic analysis shows that synaptojanin 2 depletion causes a decrease in clathrin-coated pits and vesicles. These results suggest a role for synaptojanin 2 in clathrin-coated pit formation and imply that lipid hydrolysis is required at an early stage of clathrin-mediated endocytosis. Taken together, our results also indicate that synaptojanin 2 is functionally distinct from synaptojanin 1.
突触结合蛋白2是一种广泛表达的多磷酸肌醇磷酸酶，其催化结构域与突触结合蛋白1具有高度同源性[1,2]。突触结合蛋白1缺陷型小鼠的神经元中，网格蛋白包被小泡增多，回收小泡重新进入有融合能力的小泡池的过程延迟，但内吞作用的早期步骤没有缺陷[3,4]。在此我们表明，通过小干扰（si）RNA抑制突触结合蛋白2的表达会在肺癌细胞系中导致网格蛋白介导的受体内化出现严重缺陷。野生型突触结合蛋白2的过表达可挽救这种抑制表型，但野生型突触结合蛋白1或缺乏5'-磷酸酶活性的突变型突触结合蛋白2的过表达则不能。此外，电子显微镜分析表明，突触结合蛋白2的缺失会导致网格蛋白包被小窝和小泡减少。这些结果表明突触结合蛋白2在网格蛋白包被小窝形成中发挥作用，并暗示在网格蛋白介导的内吞作用早期需要脂质水解。综上所述，我们的结果还表明突触结合蛋白2在功能上与突触结合蛋白1不同。
