Pathways and itinerary of clathrin-independent endocytosis

这个项目的重点是了解网格蛋白独立形式的内吞作用。 在没有网格蛋白外壳的情况下发生的内吞作用发生在所有细胞中，但了解甚少。 我们有兴趣研究通过这种机制进入细胞的货物蛋白，一旦它们被内化，它们的细胞内行程以及它们是否含有允许细胞内专门分选的氨基酸序列。 我们一直在识别新的货物蛋白，并发现这些蛋白质的一个子集一旦进入细胞就采取替代的交通路线。 主要组织相容性复合物I类蛋白（MHCI）是一种原型网格蛋白依赖性货物蛋白，内化后，它到达含有货物蛋白（如转铁蛋白受体）的内体，这些货物蛋白通过网格蛋白依赖性内吞作用进入。 从那里，MHCI要么行进到晚期内体和溶酶体，在那里它被降解，要么行进到再循环小管，将MHCI返回到细胞表面。然而，CD 44、CD 98和CD 147在许多细胞中显示出改变的行程，其中它们直接运输到再循环小管中并避免运输到晚期内体区室。 与这种改变的行程一致，CD 44、CD 98和CD 147是长寿命蛋白质，并且不像MHCI那样降解，其被路由到晚期内体。 我们现在已经确定了（Maldonado-Baez et al，2013）CD 98和CD 147胞质结构域中的序列，这些序列允许这些货物蛋白避免运输到用早期内体抗原1（EEA 1）标记的内体和溶酶体，而是运输到再循环小管。 这些序列是可转移的并且包括二酸氨基酸残基。 我们鉴定了微管和货物拴系蛋白Hook 1与CD 98和CD 147的胞质尾相互作用，并负责这种分选功能。 此外，我们发现Hook 1与Rab 22 a和微管一起将CD 98和CD 147分选到再循环小管中。 我们目前正在研究这种分拣机的其他方面，例如确定将货物分拣与管道形成和促进回收联系起来的其他组件。

This project is focused on understanding clathrin-independent forms of endocytosis. Endocytosis that occurs without clathrin coats occurs in all cells but is poorly understood. We are interested in studying the cargo proteins that enter cells by this mechanism, their intracellular itinerary once they have been internalized and whether they contain amino acid sequences that allow for specialized sorting within cells. We have been identifying new cargo proteins and found that a subset of these proteins take alternative traffic routes once they have entered cells. The major histocompatibility complex Class I protein (MHCI), is a prototypical clathrin-indepenent cargo protein and after internalization it reaches endosomes that contain cargo proteins such as the transferrin receptor that enter via clathrin-depenent endocytosis. From there, MHCI travels either to late endosomes and lysosomes where it is degraded or on to recycling tubules that return MHCI back to the cell surface. CD44, CD98, and CD147, however, show an altered itinerary in many cells where they traffic directly into the recycling tubules and avoid trafficking to late endosomal compartments. Consistent with this altered itinerary, CD44, CD98 and CD147 are long-lived proteins and are not degraded like MHCI, which is routed to late endosomes. We have now identified (Maldonado-Baez et al, 2013) sequences in the cytoplasmic domain of CD98 and CD147 that allow these cargo proteins to avoid trafficking to endosomes labeled with the early endosomal antigen 1 (EEA1) and lysosomes but instead traffic to recycling tubules. These sequences are transferrable and include di-acidic amino acid residues. We identified the microtubule and cargo tethering protein Hook1 as interacting with the cytoplasmic tails of CD98 and CD147 and responsible for this sorting function. In addition, we found that Hook1 works together with Rab22a and microtubules to sort CD98 and CD147 into recycling tubules. We are currently investigating other aspects of this sorting machine, such as identifying other components that link cargo sorting to tube formation and facilitation of recycling.