Abstract 825: Translocation of p27 to the Mitochondria Links Mitochondrial Energy Metabolism with Endothelial Cell Migration
The migratory capacity of endothelial cells (EC) is a key prerequisite for the growth of new blood vessels during neovascularisation of ischemic and/or tumor tissues. Recently, mitochondria have been identified as signalling organelles in EC, which may contribute to migratory capacity. Therefore, the aim of this study was to investigate whether a molecular link exists between mitochondrial energy metabolism and migratory capacity of EC. To determine whether mitochondria are required for migratory capacity, we incubated EC with 50 ng/ml ethidiume bromide (EtBr) in the presence of uridine to deplete mitochondrial DNA (mtDNA) and, thereby, destroy active mitochondria. EC, which were depleted of mtDNA, did not migrate in a scratch wound assay (− EtBr: 22 +/− 3 migrated cells; + EtBr: 0.15 +/− 0.1 migrated cells). Since protein translocation is a major mechanism for modulating the functional capacity of the mitochondria, we hypothesized that induction of migration by the promigratory stimulus vascular endothelial growth factor (VEGF) leads to translocation of proteins to mitochondria. Recent studies demonstrated that a non-nuclear pool of p27 exerts biological activities including increase of migratory capacity independent of its cell cycle inhibitory activity. Stimulation of EC with VEGF induced a 2.5 +/− 0.4-fold increase in mitochondrially localized p27 protein. Reducing p27 expression by two different siRNAs completely abrogated VEGF-induced migration of EC (98 +/− 2 % inhibition). Next, we cloned p27 into a mitochondrial targeted vector (p27mito) to exclusively overexpress p27 in the mitochondria. Overexpression of p27mito stimulated mitochondrial ATP biosynthesis (3.24 +/− 1.06-fold empty vector), increased mitochondrial membrane potential and enhanced migratory capacity of EC to a similar extent as compared to VEGF. Cells overexpressing p27mito did not demonstrate further increased migratory capacity in response to VEGF. Finally, only p27mito but not nuclear-targeted p27 overexpression rescued the impaired migratory capacity after siRNA-mediated genetic ablation of endogenous p27 expression. These data demonstrate a novel role of mitochondrial p27 as a molecular link between cellular energy metabolism and migratory capacity of EC.