Rappoport Joshua Z, Taha Bushra W, Lemeer Simone, Benmerah Alexandre, Simon Sanford M
The Laboratory of Cellular Biophysics, The Rockefeller University, New York, New York 10021, USA.
J Biol Chem. 2003 Nov 28;278(48):47357-60. doi: 10.1074/jbc.C300390200. Epub 2003 Oct 6.
Numerous biologically relevant substrates are selectively internalized via clathrin-mediated endocytosis. At the plasma membrane the AP-2 complex plays a major role in clathrin coat formation, interacting with both cargo and clathrin. Utilizing simultaneous dual-channel total internal reflection fluorescence microscopy we have analyzed components of the AP-2 complex (alpha- and beta 2-adaptin) during clathrin-mediated endocytosis. Although in static images enhanced green fluorescent protein-tagged AP-2 markers significantly co-localized with clathrin and other components of clathrin-coated pits, AP-2 did not seem to be present in clathrin spots that appeared to undergo internalization or motility parallel to the plane of the plasma membrane. Two populations of clathrin at the plasma membrane seem to exist, the dynamic and the static, and AP-2 appears to be only found within the latter. These results suggest that colocalized clathrin/AP-2 puncta may represent loci for coated pit production and that previous models that assumed AP-2 was retained within clathrin coats during endocytosis may need to be re-evaluated.
许多具有生物学相关性的底物通过网格蛋白介导的内吞作用被选择性内化。在质膜上，AP-2复合物在网格蛋白包被的形成中起主要作用，它与货物和网格蛋白都相互作用。利用同步双通道全内反射荧光显微镜，我们分析了网格蛋白介导的内吞作用过程中AP-2复合物的成分（α-和β2-衔接蛋白）。尽管在静态图像中，增强型绿色荧光蛋白标记的AP-2标记物与网格蛋白和网格蛋白包被小窝的其他成分显著共定位，但AP-2似乎不存在于那些似乎正在内化或平行于质膜平面移动的网格蛋白斑点中。质膜上似乎存在两种网格蛋白群体，即动态群体和静态群体，而AP-2似乎仅存在于后者中。这些结果表明，共定位的网格蛋白/AP-2斑点可能代表包被小窝产生的位点，并且先前认为AP-2在胞吞作用期间保留在网格蛋白包被内的模型可能需要重新评估。
