Mitochondrial ABC transporters function: the role of ABCB10 (ABC-me) as a novel player in cellular handling of reactive oxygen species

Abstract

Mitochondria are one of the major sources of reactive oxygen species (ROS) in the cell. When exceeding the capacity of antioxidant mechanisms, ROS production may lead to different pathologies, such as ischemia-reperfusion injury, neurodegeneration, anemia and ageing. As a consequence of the endosymbiotic origin of mitochondria, eukaryotic cells have developed different transport mechanisms that coordinate mitochondrial function with other cellular compartments. Four mitochondrial ATP-binding cassette (ABC) transporters have been described to date in mammals: ABCB6, ABCB8, ABCB7 and ABCB10. ABCB10 is located in the inner mitochondrial membrane forming homodimers, with the ATP binding domain facing the mitochondrial matrix. ABCB10 expression is highly induced during erythroid differentiation and its overexpression increases hemoglobin synthesis in erythroid cells. However, ABCB10 is also expressed in nonerythroid tissues, suggesting a role not directly related to hemoglobin synthesis. Recent evidence points toward ABCB10 as an important player in the protection from oxidative stress in mammals. In this regard, ABCB10 is required for normal erythropoiesis and cardiac recovery after ischemia-reperfusion, processes intimately related to mitochondrial ROS generation. Here, we review the current knowledge on mitochondrial ABC transporters and ABCB10 and discuss the potential mechanisms by which ABCB10 and its transport activity may regulate oxidative stress. We discuss ABCB10 as a potential therapeutic target for diseases in which increased mitochondrial ROS production and oxidative stress play a major role.

Figure 1. ABCB10 topology in the inner mitochondrial membrane and its functional domains

A) Cartoon representing ABCB10 topology in the inner mitochondrial membrane. ABCB10 nucleotide binding domain NBD is facing the mitochondrial matrix and 6 transmembrane domains span the inner mitochondrial membrane. ABCB10 forms homodimers and not heterodimers. The location of the NBD, in the C-terminal portion of the protein, suggests that ABCB10 is exporting its substrate/s to the inner membrane space (IMS) (see arrow). The N-terminal part of the protein contains a long mitochondrial pretargeting sequence cleaved after insertion. Fluorescence microscope images of a mouse NIH-3T3 fibroblast expressing ABCB10-GFP (green) and stained with the mitochondrial membrane potential sensitive probe TMRE (red). Merged images show perfect colocalization of ABCB10-GFP with TMRE, demonstrating that ABCB10-GFP targets to mitochondria. B) ABCB10 expression in different mouse tissues measured by Northern blot. Liver, heart, brain and kidney show ABCB10 expression (highest levels in liver, kidney and heart). Differentiation of Mouse erythroleukemia cells (MEL) is associated with GATA-1 activation (master transcription factor regulating terminal eythroid differentiation and hemoglobin synthesis) and a large increase in ABCB10 expression measured by Northern blot. This figure was adapted from reference 76, Shirihai et al. (2000) EMBO Journal. C) Alignment of human ABCB10 (NP_036221) and human ABCB1/MDR1 (NP_000918.2) amino-acidic sequences using Blast P (37% identity covering a 78% of ABCB10 sequence). The transmembrane-TM- and nucleotide binding-NBD- domains are in light green and grey respectively. The Walker A, B and C-loop motifs are highlighted within the NBD in different colors. Residues conserved in ABCB10 that were mutated in functional studies of ABCB1 or other ABC proteins are marked with asterisks.

Figure 2. ABCB10 is essential for erythropoiesisin vivo

A) Light microscope image of day 10.5 embryos still attached to the placenta and inside the yolk sac. The 2 embryos in the upper part of the image harbor inactivated ABCB10 alleles (−/−) and showed beating hearts. The embryo in the bottom part of the image was a wild type. Note that the wild type embryo has hemoglobinized cells in the vasculature of the yolk sac (give red color to vessels), whereas the ABCB10 −/− embryos show vessels with no red color (lack of hemoglobinized cells). The yolk sac is the main site of primitive erythropoiesis at day 9.5– 10.5. See reference 38 for a more detailed description. B) Representative images of differentiated MEL cells (murine erythroleukemia cells) stably transfected with an empty vector (control) or containing ABCB10 cDNA (ABCB10 over-expression) and stained with benzidine for hemoglobin detection (orange / brown color). This figure panel was adapted from reference 76, Shirihai et al. (2000) EMBO Journal.

Figure 3. ABCB10 is required to protect from increased oxidative stress associated to cardiac ischemia-reperfusion

A) Diastolic and systolic pressure in wild type (WT) and ABCB10 +/− hearts (red trace) during ischemia-reperfusion. Increased diastolic and decreased systolic pressures are symptomatic of impaired mechanical function. Pretreatment for 20 minutes with the antioxidant EUK-207, a superoxide dismutase and catalase mimetic, prevented these defects in ABCB10 +/− hearts (dashed open square traces). B) Mitochondrial ATP synthesis rates after ischemia-reperfusion of wild type and ABCB10 +/− hearts. The functional defect in ABCB10 +/− cardiac mitochondria was prevented by EUK-207 treatment. See reference 53 for a more detailed description.

Figure 4. Mitochondrial ABC transporters cluster according to their different physiological roles reported

A) Phylogenic analysis: dendogram obtained using ClustalW2 software from the EMBL website and using the aminoacid sequences of ABC proteins. Hs: Homo sapiens; Sc: Saccharomyces cerevisae (fungi); Rp22: Rickettsia prowazekii strain 22 (prokaryote, ancestor of mitochondria); Pb: Paracoccidioides brasiliensis (fungi). Note that human ABCB7 clusters together with ATM1 from Saccharomyces cerevisae and from Rickettsia prowazekii, suggesting that its function permitting iron/sulfur cluster synthesis is likely to be conserved. ABCB6, despite its high levels of aminoacid identity by BLAST protein analysis to ABCB7 and ATM1Sc, clusters separately. This separation fits well with ABCB6 heterogeneous localization inside the cell and the likely possibility that it transports multiple substrates. ABCB10, Pfr1 and MDL1 have all been shown to be required for protection under conditions of increased oxidative stress related to heme and iron metabolism. Consistent with this, they cluster together. ABCB8, on the other hand, has been reported to be involved in different processes, such as multidrug resistance in cancer cell lines (in doxorubicin export), mitochondrial K+ ATP channel, iron and GSH export in the heart. These heterogeneous functions suggest a less conserved and more specialized function of ABCB8. In addition, MDR (multidrug resistance protein) of Rickettsia Prowazekii clusters separately from ABCB10 and ABCB8. Rickettsia Prowazekii MDR function is unknown, but we think that it will be closer to ABCB10, as the latter clusters together with lower eukaryotes orthologues, suggesting a more conserved function. B) Sequence alignment of human ABCB10 (ABCB10Hs) with Saccharomyces cerevisae MDL1 (Mdl1pSc) and Paracoccidioides brasiliensis Pfr-1 (Pfr1Pb) using Clustal W2 software. Identical residues are marked with asterisks. The transmembrane-TM- and nucleotide binding-NBD- domains, according to ABCB10 sequence, are in light green and grey respectively. The Walker A, B and C-loop motifs are highlighted within the NBD in different colors. ABCB10 mitochondrial target sequence (MTS) is also highlighted in orange. Note that MTS is the less conserved region of the protein.