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Separate endocytic pathways regulate IL‐5 receptor internalization and signaling
Author(s) -
Lei Jonathan T.,
MartinezMoczygemba Margarita
Publication year - 2008
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1189/jlb.1207828
Subject(s) - endocytic cycle , biology , microbiology and biotechnology , clathrin , endocytosis , internalization , lipid raft , endosome , signal transduction , transferrin receptor , cholera toxin , receptor , intracellular , biochemistry
Eosinophils are critically dependent on IL‐5 for their activation, differentiation, survival, and augmentation of cytotoxic activity. We previously showed that the cytoplasmic domain of the hematopoietic receptor, βc, which is shared by IL‐5, IL‐3, and GM‐CSF, is directly ubiquitinated and degraded by the proteasomes in a JAK2‐dependent manner. However, studies describing the spatial distribution, endocytic regulation, and trafficking of βc‐sharing receptors in human eosinophils are currently lacking. Using deconvolution microscopy and biochemical methods, we clearly demonstrate that IL‐5Rs reside in and are internalized by clathrin‐ and lipid raft‐dependent endocytic pathways. Microscopy analyses in TF1 cells and human eosinophils revealed significant colocalization of βc, IL‐5Rα, and Cy3‐labeled IL‐5 with transferrin‐ (clathrin) and cholera toxin‐B‐ (lipid raft) positive vesicles. Moreover, whereas internalized IL‐5Rs were detected in both clathrin‐ and lipid raft‐positive vesicles, biochemical data revealed that tyrosine phosphorylated, ubiquitinated, and proteasome‐degraded IL‐5Rs partitioned to the soluble, nonraft fractions (clathrin‐containing). Lastly, we show that optimal IL‐5‐induced signaling requires entry of activated IL‐5Rs into the intracellular compartment, as coimmunoprecipitation of key signaling molecules with the IL‐5R was completely blocked when either endocytic pathway was inhibited. These data provide the first evidence that IL‐5Rs segregate and traffic into two distinct plasma membrane compartments, and they further establish that IL‐5R endocytosis regulates signaling both positively and negatively.