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Localization of MRP‐1 to the outer mitochondrial membrane by the chaperone protein HSP90β
Author(s) -
Roundhill Elizabeth,
Turnbull Doug,
Burchill Susan
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.15-283408
Subject(s) - mitochondrion , inner mitochondrial membrane , hsp90 , submitochondrial particle , atp–adp translocase , biology , heat shock protein , mitochondrial carrier , microbiology and biotechnology , bacterial outer membrane , mitochondrial apoptosis induced channel , biochemistry , escherichia coli , gene
Overexpression of plasma membrane multidrug resistance‐associated protein 1 (MRP‐1) in Ewing's sarcoma (ES) predicts poor outcome. MRP‐1 is also expressed in mitochondria, and we have examined the submitochondrial localization of MRP‐1 and investigated the mechanism of MRP‐1 transport and role of this organelle in the response to doxorubicin. The mitochondrial localization of MRP‐1 was examined in ES cell lines by differential centrifugation and membrane solubilization by digitonin. Whether MRP‐1 is chaperoned by heat shock proteins (HSPs) was investigated by immunoprecipitation, immunofluorescence microscopy, and HSP knockout using small hairpin RNA and inhibitors (apoptozole, 17‐AAG, and NVPAUY). The effect of disrupting mitochondrial MRP‐1‐dependent efflux activity on the cytotoxic effect of doxorubicin was investigated by counting viable cell number. Mitochondrial MRP‐1 is glycosylated and localized to the outer mitochondrial membrane, where it is coexpressed with HSP90. MRP‐1 binds to both HSP90 and HSP70, although only inhibition of HSP90β decreases expression of MRP‐1 in the mitochondria. Disruption of mitochondrial MRP‐1‐dependent efflux significantly increases the cytotoxic effect of doxorubicin (combination index, <0.9). For the first time, we have demonstrated that mitochondrial MRP‐1 is expressed in the outer mitochondrial membrane and is a client protein of HSP90β, where it may play a role in the doxorubicin‐induced resistance of ES.—Roundhill, E., Turnbull, D., Burchill, S. Localization of MRP‐1 to the outer mitochondrial membrane by the chaperone protein HSP90β. FASEB J. 30, 1712–1723 (2016). www.fasebj.org