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Clathrin-independent endocytosis of ErbB2 in geldanamycin-treated human breast cancer cells.

ABSTRACT: The epidermal growth factor (EGF)-receptor family member ErbB2 is commonly overexpressed in human breast cancer cells and correlates with poor prognosis. Geldanamycin (GA) induces the ubiquitylation, intracellular accumulation and degradation of ErbB2. Whether GA stimulates ErbB2 internalization is controversial. We found that ErbB2 was internalized constitutively at a rate that was not affected by GA in SK-BR-3 breast cancer cells. Instead, GA treatment altered endosomal sorting, causing the transport of ErbB2 to lysosomes for degradation. In contrast to earlier work, we found that ErbB2 internalization occurred by a clathrin- and tyrosine-kinase-independent pathway that was not caveolar, because SK-BR-3 cells lack caveolae. Similar to cargo of the glycosylphosphatidylinositol (GPI)-anchored protein-enriched early endosomal compartment (GEEC) pathway, internalized ErbB2 colocalized with cholera toxin B subunit, GPI-anchored proteins and fluid, and was often seen in short tubules or large vesicles. However, in contrast to the GEEC pathway in other cells, internalization of ErbB2 and fluid in SK-BR-3 cells did not require Rho-family GTPase activity. Accumulation of ErbB2 in vesicles containing constitutively active Arf6-Q67L occurred only without GA treatment; Arf6-Q67L did not slow transport to lysosomes in GA-treated cells. Further characterization of this novel clathrin-, caveolae- and Rho-family-independent endocytic pathway might reveal new strategies for the downregulation of ErbB2 in breast cancer.

SUBMITTER: Barr DJ

PROVIDER: S-EPMC2707784 | biostudies-literature | 2008 Oct

REPOSITORIES: biostudies-literature

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Clathrin-independent endocytosis of ErbB2 in geldanamycin-treated human breast cancer cells.

Barr Daniel J DJ Ostermeyer-Fay Anne G AG Matundan Rachel A RA Brown Deborah A DA

Journal of cell science 20080902 Pt 19

The epidermal growth factor (EGF)-receptor family member ErbB2 is commonly overexpressed in human breast cancer cells and correlates with poor prognosis. Geldanamycin (GA) induces the ubiquitylation, intracellular accumulation and degradation of ErbB2. Whether GA stimulates ErbB2 internalization is controversial. We found that ErbB2 was internalized constitutively at a rate that was not affected by GA in SK-BR-3 breast cancer cells. Instead, GA treatment altered endosomal sorting, causing the tr ...[more]

PMID: 18765569

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Project description:Breast Cancer Stem (BCS) cells play critical roles in self-renewal, Multi Drug Resistance (MDR), differentiation and generation of secondary tumors. Conventional chemotherapy may efficiently kill the bulk of differentiated drug sensitive breast cancer cells, but not the MDR self-renewable BCS cells, leading to enrichment of the MDR BCS cells. In order to target the MDR BCS cells, we have isolated: 1) BCS cells from either breast cancer cell lines or fresh breast cancer specimens; 2) ATP binding cassette (ABC) transporter group G number 2 (ABCG2)-specific aptamers; and 3) BCS cell-binding aptamers. Interestingly, ABCG2-specific aptamers labeled the membrane surface of the ABCG2-expressing baby hamster kidney (BHK) cells, but stained whole cells of the BCS cells derived from mammospheres, implying that BCS cells might have much higher rate of endocytosis than the ABCG2-expressing BHK cells. In addition, 5D3, a monoclonal antibody that recognizes the extracellular loops of ABCG2 protein, also stained whole BCS cells. Furthermore, BCS cell-binding aptamers stained whole BCS cells, but not the differentiated breast cancer MCF-7 cells. All these results support above conclusion that BCS cells might have high rate of endocytosis. Further experiments performed with aptamers and human transferrin or lactosylceramide showed that BCS cells do have much higher endocytosis rate than the differentiated breast cancer cells. Interestingly, clathrin dependent endocytosis inhibitors, such as monodansylcadaverine or sucrose, or caveolin-dependent endocytosis inhibitors, such as methyl-?-cyclodextrin or genistein, can inhibit the internalization of transferrin or lactosylceramide into the differentiated breast cancer cells, but cannot block the internalization of these compounds into the BCS cells, suggesting that BCS cells undergo clathrin-independent and caveolin-independent endocytosis. Taken together, our data suggest that BCS cells have high rate of endocytosis and open the possibilities for delivering therapeutic agents directly into the MDR BCS cells with aptamer-coated liposomes.

Dataset Information

Clathrin-independent endocytosis of ErbB2 in geldanamycin-treated human breast cancer cells.

Publications

Clathrin-independent endocytosis of ErbB2 in geldanamycin-treated human breast cancer cells.

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