Miro1-mediated mitochondrial positioning supports subcellular redox status

Mitochondria are strategically trafficked throughout the cell by the action of microtubule motors, the actin cytoskeleton and adapter proteins. The intracellular positioning of mitochondria supports subcellular levels of ATP, Ca(2+) and reactive oxygen species (ROS, i.e. hydrogen peroxide, H(2)O(2)). Previous work from our group showed that deletion of the mitochondrial adapter protein Miro1 leads to perinuclear clustering of mitochondria, leaving the cell periphery devoid of mitochondria which compromises peripheral energy status. Herein, we report that deletion of Miro1 significantly restricts subcellular H(2)O(2) levels to the perinuclear space which directly affects intracellular responses to elevated mitochondrial ROS. Using the genetically encoded H(2)O(2)-responsive fluorescent biosensor HyPer7, we show that the highest levels of subcellular H(2)O(2) map to sites of increased mitochondrial density. Deletion of Miro1 or disruption of microtubule dynamics with Taxol significantly reduces peripheral H(2)O(2) levels. Following inhibition of mitochondrial complex 1 with rotenone we observe elevated spikes of H(2)O(2) in the cell periphery and complementary oxidation of mitochondrial peroxiredoxin 3 (PRX3) and cytosolic peroxiredoxin 2 (PRX2). Conversely, in cells lacking Miro1, rotenone did not increase peripheral H(2)O(2) or PRX2 oxidation but rather lead to increased nuclear H(2)O(2) and an elevated DNA-damage response. Lastly, local levels of HyPer7 oxidation correlate with the size and abundance of focal adhesions (FAs) in MEFs and cells lacking Miro1 have significantly smaller focal adhesions and reduced phosphorylation levels of vinculin and p130Cas compared to Miro1(+/+) MEFs. Together, we present evidence that the intracellular distribution of mitochondria influences subcellular H(2)O(2) levels and local cellular responses dependent on mitochondrial ROS.