Calpain inhibitor causes accumulation of ubiquitinated P-glycoprotein at the cell surface: possible role of calpain in P-glycoprotein turnover
| Autor: | Koji Takada, Tadashi Asakura, Nozomu Yanaihara, Kiyoshi Ohkawa, T Sawai, Yukio Hashizume, Yutaka Okawa |
|---|---|
| Rok vydání: | 1999 |
| Předmět: |
Cancer Research
ATP Binding Cassette Transporter Subfamily B Time Factors endocrine system diseases Leupeptins Blotting Western Cell Lactacystin Enzyme-Linked Immunosorbent Assay Cysteine Proteinase Inhibitors Ammonium Chloride chemistry.chemical_compound Leucine polycyclic compounds medicine Humans Ubiquitins Glycoproteins P-glycoprotein Protein Synthesis Inhibitors Brefeldin A integumentary system biology Calpain Cell Membrane Membrane Proteins Dipeptides Cell cycle female genital diseases and pregnancy complications Acetylcysteine Cell biology carbohydrates (lipids) medicine.anatomical_structure Oncology Biochemistry chemistry Doxorubicin Apoptosis Cytoplasm biology.protein K562 Cells |
| Zdroj: | International Journal of Oncology. |
| ISSN: | 1791-2423 1019-6439 |
| DOI: | 10.3892/ijo.15.4.677 |
| Popis: | P-glycoprotein (Pgp) is a plasma-membrane glycoprotein that confers multi-drug resistance (MDR) on cells and displays ATP-driven drug pumping. The possible contribution of calpain-mediated proteolytic pathways to the functional regulation of the Pgp molecule was evaluated using K562/DXR, MDR cells. N-Acetyl-L-leucyl-L-leucyl-norleucinal was effluxed by Pgp, but N-benzyloxycarbonyl-L-leucyl-L-leucinal (zLLal), an inhibitor of calpain, retarded the degradation of Pgp leading to accumulation of the molecule largely at the cell surface membrane. Treatment with brefeldin A did not obstruct the zLLal-induced Pgp accumulation. NH4Cl increased the cytoplasmic Pgp level, with a slight to significant decrease at the cell surface membrane. Ubiquitin-ELISA and western blot analysis confirmed that the Pgp molecule, which accumulated mainly at the cell surface, was ubiquitinated. However, lactacystin did not show any accumulation of Pgp in either the cytoplasm or the cell surface membrane, suggesting that the proteasome did not participate in the phenomenon. Additionally, the Pgp was limitedly proteolyzed by calpain into two 98 kDa and 69 kDa, fragments within one minute. Despite the increased accumulation of Pgp at the cell surface after treatment with calpain inhibitor, the cytoplasmic doxorubicin level of the cells treated with a calpain inhibitor was higher than that of non-treated cells and approached that of parental cells. These results indicated that calpain involved Pgp turnover and that calpain inhibition induced ubiquitinated Pgp-accumulation mainly at the cell surface membrane with a reduction in its own functions suggesting that the modulation of Pgp-turnover involves MDR-reversal by another approach. |
| Databáze: | OpenAIRE |
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