Program in Molecular Medicine, University of Massachusetts Medical School, Worcester, MA 01605, USA.
Proc Natl Acad Sci U S A. 2012 Feb 21;109(8):E471-80. doi: 10.1073/pnas.1115495109. Epub 2012 Jan 30.
Cell surface receptors and other proteins internalize through diverse mechanisms at the plasma membrane and are sorted to different destinations. Different subpopulations of early endosomes have been described, raising the question of whether different internalization mechanisms deliver cargo into different subsets of early endosomes. To address this fundamental question, we developed a microscopy platform to detect the precise position of endosomes relative to the plasma membrane during the uptake of ligands. Axial resolution is maximized by concurrently applied total internal reflection fluorescence and epifluorescence-structured light. We found that transferrin receptors are delivered selectively from clathrin-coated pits on the plasma membrane into a specific subpopulation of endosomes enriched in the multivalent Rab GTPase and phosphoinositide-binding protein Rabenosyn-5. Depletion of Rabenosyn-5, but not of other early endosomal proteins such as early endosome antigen 1, resulted in impaired transferrin uptake and lysosomal degradation of transferrin receptors. These studies reveal a critical role for Rabenosyn-5 in determining the fate of transferrin receptors internalized by clathrin-mediated endocytosis and, more broadly, a mechanism whereby the delivery of cargo from the plasma membrane into specific early endosome subpopulations is required for its appropriate intracellular traffic.
细胞表面受体和其他蛋白质通过质膜中的多种机制内化，并被分类到不同的目的地。已经描述了不同的早期内涵体亚群，这引发了一个问题，即不同的内吞作用机制是否将货物输送到不同的早期内涵体亚群中。为了解决这个基本问题，我们开发了一种显微镜平台，用于在配体摄取过程中检测相对于质膜的内涵体的精确位置。通过同时应用全内反射荧光和荧光结构光，最大限度地提高了轴向分辨率。我们发现，转铁蛋白受体从质膜上的网格蛋白包被小窝中被选择性地递送到富含多价 Rab GTPase 和磷酸肌醇结合蛋白 Rabenosyn-5 的特定内涵体亚群中。Rabenosyn-5 的耗竭，但不是其他早期内涵体蛋白，如早期内涵体抗原 1，导致转铁蛋白摄取受损和转铁蛋白受体的溶酶体降解。这些研究揭示了 Rabenosyn-5 在决定网格蛋白介导的内吞作用内化的转铁蛋白受体命运中的关键作用，更广泛地说，是一种将货物从质膜递送到特定早期内涵体亚群中以进行适当的细胞内运输的机制。
