Motley Alison, Bright Nicholas A, Seaman Matthew N J, Robinson Margaret S
University of Cambridge, Department of Clinical Biochemistry, Cambridge Institute for Medical Research, Cambridge CB2 2XY, UK.
J Cell Biol. 2003 Sep 1;162(5):909-18. doi: 10.1083/jcb.200305145.
We have used RNA interference to knock down the AP-2 mu2 subunit and clathrin heavy chain to undetectable levels in HeLaM cells. Clathrin-coated pits associated with the plasma membrane were still present in the AP-2-depleted cells, but they were 12-fold less abundant than in control cells. No clathrin-coated pits or vesicles could be detected in the clathrin-depleted cells, and post-Golgi membrane compartments were swollen. Receptor-mediated endocytosis of transferrin was severely inhibited in both clathrin- and AP-2-depleted cells. Endocytosis of EGF, and of an LDL receptor chimera, were also inhibited in the clathrin-depleted cells; however, both were internalized as efficiently in the AP-2-depleted cells as in control cells. These results indicate that AP-2 is not essential for clathrin-coated vesicle formation at the plasma membrane, but that it is one of several endocytic adaptors required for the uptake of certain cargo proteins including the transferrin receptor. Uptake of the EGF and LDL receptors may be facilitated by alternative adaptors.
我们利用RNA干扰技术将HeLaM细胞中的AP-2 μ2亚基和网格蛋白重链敲低至无法检测的水平。与质膜相关的网格蛋白包被小窝在AP-2缺失的细胞中仍然存在，但它们的丰度比对照细胞低12倍。在网格蛋白缺失的细胞中未检测到网格蛋白包被小窝或囊泡，高尔基体后膜区室肿胀。转铁蛋白的受体介导内吞作用在网格蛋白和AP-2缺失的细胞中均受到严重抑制。表皮生长因子（EGF）和低密度脂蛋白受体嵌合体的内吞作用在网格蛋白缺失的细胞中也受到抑制；然而，在AP-2缺失的细胞中，它们的内化效率与对照细胞一样高。这些结果表明，AP-2对于质膜上网格蛋白包被囊泡的形成并非必不可少，但它是摄取某些货物蛋白（包括转铁蛋白受体）所需的几种内吞衔接蛋白之一。EGF和低密度脂蛋白受体的摄取可能由其他衔接蛋白促进。
