Loss of neuronal Miro1 disrupts mitophagy and induces hyperactivation of the integrated stress response.

Clearance of mitochondria following damage is critical for neuronal homeostasis. Here, we investigate the role of Miro proteins in mitochondrial turnover by the PINK1/Parkin mitochondrial quality control system in vitro and in vivo. We find that upon mitochondrial damage, Miro is promiscuously ubiquitinated on multiple lysine residues. Genetic deletion of Miro or block of Miro1 ubiquitination and subsequent degradation lead to delayed translocation of the E3 ubiquitin ligase Parkin onto damaged mitochondria and reduced mitochondrial clearance in both fibroblasts and cultured neurons. Disrupted mitophagy in vivo, upon post-natal knockout of Miro1 in hippocampus and cortex, leads to a dramatic increase in mitofusin levels, the appearance of enlarged and hyperfused mitochondria and hyperactivation of the integrated stress response (ISR). Altogether, our results provide new insights into the central role of Miro1 in the regulation of mitochondrial homeostasis and further implicate Miro1 dysfunction in the pathogenesis of human neurodegenerative disease.

Infrared nanoimaging of neuronal ultrastructure and nanoparticle interaction with cells.

Here we introduce scattering-type scanning near-field optical microscopy (s-SNOM) as a novel tool for nanoscale chemical-imaging of sub-cellular organelles, nanomaterials and of the interactions between them. Our setup uses a tuneable mid-infrared laser and a sharp scanning probe to image at a resolution substantially surpassing the diffraction limit. The laser can be tuned to excite vibrational modes of functional groups in biomolecules, (e.g. amide moieties), in a way that enables direct chemical mapping without the need for labelling. We, for the first time, chemically image neuronal ultrastructure, identify neuronal organelles and sub-organelle structures as small as 10 nm and validate our findings using transmission electron microscopy (TEM). We produce chemical and morphological maps of neurons treated with gold nanospheres and characterize nanoparticle size and intracellular location, and their interaction with the plasma membrane. Our results show that the label-free nature of s-SNOM means it has a 'true' chemical resolution of up to 20 nm which can be further improved. We argue that it offers significant potential in nanomedicine for nanoscale chemical imaging of cell ultrastructure and the subcellular distribution of nanomaterials within tissues.

Nanostars Carrying Multifunctional Neurotrophic Dendrimers Protect Neurons in Preclinical In Vitro Models of Neurodegenerative Disorders.

A challenge in neurology is the lack of efficient brain-penetrable neuroprotectants targeting multiple disease mechanisms. Plasmonic gold nanostars are promising candidates to deliver standard-of-care drugs inside the brain but have not been trialed as carriers for neuroprotectants. Here, we conjugated custom-made peptide dendrimers (termed H3/H6), encompassing motifs of the neurotrophic S100A4-protein, onto star-shaped and spherical gold nanostructures (H3/H6-AuNS/AuNP) and evaluated their potential as neuroprotectants and interaction with neurons. The H3/H6 nanostructures crossed a model blood-brain barrier, bound to plasma membranes, and induced neuritogenesis with the AuNS, showing higher potency/efficacy than the AuNP. The H3-AuNS/NP protected neurons against oxidative stress, the H3-AuNS being more potent, and against Parkinson's or Alzheimer's disease (PD/AD)-related cytotoxicity. Unconjugated S100A4 motifs also decreased amyloid beta-induced neurodegeneration, introducing S100A4 as a player in AD. Using custom-made dendrimers coupled to star-shaped nanoparticles is a promising route to activate multiple neuroprotective pathways and increase drug potency to treat neurodegenerative disorders.