Pathways and itinerary of clathrin-independent endocytosis

这个项目的重点是了解不依赖于笼蛋白的内吞作用形式。内吞作用在所有细胞中都会发生，但人们对此知之甚少。我们感兴趣的是研究通过这种机制进入细胞的货物蛋白质，一旦它们被内化，它们在细胞内的行程，以及它们是否包含允许在细胞内进行专门分选的氨基酸序列。我们一直在鉴定新的货运蛋白，并发现一旦这些蛋白进入细胞，它们的子集就会采取替代的运输路线。主要组织相容性复合体I类蛋白(MHCI)是一种典型的不依赖于笼蛋白的货运蛋白，内化后到达内体，内体中含有转铁蛋白受体等货物蛋白，通过胞吞作用进入细胞内。从那里，MHCI要么进入晚期的内小体和溶酶体，在那里被降解，要么进入将MHCI返回细胞表面的循环管。然而，CD44、CD98和CD147在许多细胞中显示了一种改变的行程，它们直接进入回收小管，避免运输到晚期的内体隔室。与这种改变的路线一致的是，CD44、CD98和CD147是长寿蛋白，不像MHCI那样被降解，后者被路由到较晚的内吞体内。 我们现在已经在CD98和CD147的细胞质结构域中发现了(Maldonado-Baez等，2013)序列，这些序列使这些货物蛋白能够避免运输到标记有早期内体抗原1(EEA1)的内小体和溶酶体，而是运输到回收小管。这些序列是可转移的，包括双酸性氨基酸残基。我们鉴定了微管和货物拴系蛋白Hook1与CD98和CD147的细胞质尾巴相互作用，并负责这种分选功能。此外，我们发现Hook1与Rab22a和微管一起作用，将CD98和CD147分类成回收管。我们目前正在研究这种分拣机器的其他方面，例如识别将货物分拣与管道形成联系起来的其他组件，以及促进回收。

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.