Detection and characterisation of multi-drug resistance protein 1 (MRP-1) in human mitochondria.
Br J Cancer. 2012 Feb 21. Epub 2012 Feb 21. PMID: 22353810
Children's Cancer Research Group, Leeds Institute of Molecular Medicine, St. James's University Hospital, Beckett Street, Leeds LS9 7TF, UK.
Background:Overexpression of plasma membrane multi-drug resistance protein 1 (MRP-1) can lead to multidrug resistance. In this study, we describe for the first time the expression of mitochondrial MRP-1 in untreated human normal and cancer cells and tissues.Methods:MRP-1 expression and subcellular localisation in normal and cancer cells and tissues was examined by differential centrifugation and western blotting, and immunofluorescence microscopy. Viable mitochondria were isolated and MRP-1 efflux activity measured using the calcein-AM functional assay. MRP-1 expression was increased using retroviral infection and specific overexpression confirmed by RNA array. Cell viability was determined by trypan blue exclusion and annexin V-propidium iodide labelling of cells.Results:MRP-1 was detected in the mitochondria of cancer and normal cells and tissues. The efflux activity of mitochondrial MRP-1 was more efficient (55-64%) than that of plasma membrane MRP-1 (11-22%; P<0.001). Induced MRP-1 expression resulted in a preferential increase in mitochondrial MRP-1, suggesting selective targeting to this organelle. Treatment with a non-lethal concentration of doxorubicin (0.85 nM, 8 h) increased mitochondrial and plasma membrane MRP-1, increasing resistance to MRP-1 substrates. For the first time, we have identified MRP-1 with efflux activity in human mitochondria.Conclusion:Mitochondrial MRP-1 may be an exciting new therapeutic target where historically MRP-1 inhibitor strategies have limited clinical success.British Journal of Cancer advance online publication, 21 February 2012; doi:10.1038/bjc.2012.40 www.bjcancer.com.