Department of Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, Liverpool L69 3BX, UK.
Department of Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, Liverpool L69 3BX, UK Division of Biomedical Cell Biology, Warwick Medical School, Gibbet Hill Road, Coventry CV4 7AL, UK
Biol Open. 2014 Apr 4;3(5):326-31. doi: 10.1242/bio.20147955.
Small molecule inhibitors of clathrin-mediated endocytosis are highly desired for the dissection of membrane trafficking pathways in the lab and for potential use as anti-infectives in the clinic. One inhibition strategy is to prevent clathrin from contacting adaptor proteins so that clathrin-mediated endocytosis cannot occur. "Pitstop" compounds have been developed that block only one of the four functional interaction sites on the N-terminal domain of clathrin heavy chain. Despite this limitation, Pitstop 2 causes profound inhibition of clathrin-mediated endocytosis. In this study, we probed for non-specific activity of Pitstop 2 by examining its action in cells expressing clathrin heavy chain harbouring mutations in the N-terminal domain interaction sites. We conclude that the inhibition observed with this compound is due to non-specificity, i.e. it causes inhibition away from its proposed mode of action. We recommend that these compounds be used with caution in cells and that they should not be used to conclude anything of the function of clathrin's N-terminal domain.
小分子网格蛋白介导入胞抑制剂在实验室中用于剖析膜运输途径，在临床上有作为抗感染药物的潜力。一种抑制策略是防止网格蛋白与衔接蛋白结合，从而阻止网格蛋白介导入胞。已经开发出了“Pitstop”化合物，它只能阻断网格蛋白重链 N 端结构域上四个功能相互作用位点之一。尽管存在这种局限性，但 Pitstop 2 仍会导致网格蛋白介导入胞的显著抑制。在这项研究中，我们通过研究表达 N 端结构域相互作用位点突变的网格蛋白重链的细胞中 Pitstop 2 的作用，来探测其非特异性活性。我们的结论是，用这种化合物观察到的抑制作用是由于非特异性，即它导致了远离其作用模式的抑制。我们建议在细胞中谨慎使用这些化合物，并且不应该用它们来推断网格蛋白 N 端结构域的功能。
