Abstract 161: ABCB10 Plays a Role in Heme Synthesis and Mitochondrial δ-Aminolevulinic Acid Export

Abstract

Introduction: Heme is essential for cardiovascular health, as it is a component of myoglobin, cytochrome P450, electron transport chain complexes, and antioxidant enzymes. The eight steps of heme synthesis are well characterized; however, it is not known how the synthetic intermediates are transported between mitochondria and cytosol. We hypothesized that the mitochondrial ATP-binding cassette protein-B10 (ABCB10) plays a role in heme synthesis by mediating the export of δ-aminolevulinic acid (ALA) out of the mitochondria. Results: Lentiviral shRNA-based knockdown of ABCB10 in H9c2 cardiac myoblasts significantly reduced cytosolic and mitochondrial heme levels and the activities of heme-containing proteins (catalase, peroxidase and complex IV), while their mRNA levels remained unchanged. The observed heme deficiency was not due to a defect in heme synthesis, iron incorporation, or enhanced heme degradation, as ABCB10 shRNA had no effect on the expression of enzymes mediating these steps. We then tested whether ABCB10 regulates mitochondrial export of ALA. Addition of exogenous ALA completely reversed the reduction in heme levels and normalized the activities of heme containing enzymes in cells with ABCB10 knockdown. However, overexpression of ALA synthase 1 (ALAS1), an enzyme catalyzing the production of ALA in the mitochondria, resulted in increased heme levels at baseline, but failed to restore heme content in ABCB10 shRNA-treated cells. To examine the role of ABCB10 in cardiac disease, we measured ABCB10 expression in the hearts of mice subjected to myocardial infarction and in explanted human hearts with ischemic cardiomyopathy. ABCB10 protein was significantly elevated in mouse and human hearts, and adenoviral overexpression of ABCB10 in H9c2 cardiac myoblasts significantly attenuated the production of reactive oxygen species (ROS), suggesting the protective role for ABCB10 in heart disease. Conclusions: Our studies place ABCB10 between the mitochondrial synthesis of ALA by ALAS1 and the cytosolic conversion of ALA into heme, suggesting that ABCB10 plays a role ALA export out of the mitochondria. Furthermore, its overexpression exerts protective effects against oxidant induced cell death in cardiac cells.