Macia Eric, Ehrlich Marcelo, Massol Ramiro, Boucrot Emmanuel, Brunner Christian, Kirchhausen Tomas
Department of Cell Biology, Harvard Medical School and the CBR Institute for Biomedical Research, Inc., 200 Longwood Avenue, Boston, Massachusetts 02115, USA.
Dev Cell. 2006 Jun;10(6):839-50. doi: 10.1016/j.devcel.2006.04.002.
Dynamin is essential for clathrin-dependent coated vesicle formation. It is required for membrane budding at a late stage during the transition from a fully formed pit to a pinched-off vesicle. Dynamin may also fulfill other roles during earlier stages of vesicle formation. We have screened about 16,000 small molecules and have identified 1, named here dynasore, that interferes in vitro with the GTPase activity of dynamin1, dynamin2, and Drp1, the mitochondrial dynamin, but not of other small GTPases. Dynasore acts as a potent inhibitor of endocytic pathways known to depend on dynamin by rapidly blocking coated vesicle formation within seconds of dynasore addition. Two types of coated pit intermediates accumulate during dynasore treatment, U-shaped, half formed pits and O-shaped, fully formed pits, captured while pinching off. Thus, dynamin acts at two steps during clathrin coat formation; GTP hydrolysis is probably needed at both steps.
发动蛋白对于网格蛋白依赖性被膜小泡的形成至关重要。在从完全形成的凹陷向缢断的小泡转变的后期阶段，膜出芽需要它。发动蛋白在小泡形成的早期阶段可能也发挥其他作用。我们筛选了约16000种小分子，并鉴定出1种，在此命名为发动蛋白抑制剂，它在体外干扰发动蛋白1、发动蛋白2和线粒体发动蛋白Drp1的GTP酶活性，但不影响其他小GTP酶。发动蛋白抑制剂是已知依赖发动蛋白的内吞途径的有效抑制剂，在添加发动蛋白抑制剂后数秒内即可迅速阻断被膜小泡的形成。在发动蛋白抑制剂处理期间，会积累两种类型的被膜凹陷中间体，即U形的、半形成的凹陷和O形的、完全形成且在缢断时捕获的凹陷。因此，发动蛋白在网格蛋白包被形成过程中作用于两个步骤；这两个步骤可能都需要GTP水解。
