Mitochondrial Calcium and Neuronal Health

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ABSTRACT Propagation of calcium (Ca2+) signals to the mitochondria through the Ca2+ uniporter has long been considered central to neuronal function. The discovery of the molecular components of the uniporter including the Ca2+ sensing regulator MICU1, was followed by the identification of many MICU1 loss-of-function patients who display progressive neurological disorder, dominated by motor and learning impairments, which were recapitulated in mouse models created by us and others. Emerging evidence indicates that acute MICU1 proteolytic degradation by YME1L also occurs during hypoxia, suggesting that dysregulation of mitochondrial Ca2+ homeostasis may also play a role in ischemia/stroke. However, the neuronal pathogenesis associated with MICU1 deficiency and, more broadly, the role of uniporter regulation in healthy neurons, remain poorly understood. MICU1 forms dimers with its paralogs, MICU2 and MICU3, and with itself. MICU2 has also been linked to human neurological impairments but is scarce in adult neurons, whereas MICU3 is abundant in adult brain. Our central hypothesis is that control of mitochondrial Ca2+ uptake by MICU1/2/3 is essential for coordinating mitochondrial function with synaptic activity. This control also prevents mitochondrial Ca2+ overload and oxidant dysregulation that lead to neuronal stress. Further, we postulate dynamic tuning of neuronal MICU-dependent gatekeeping: during development—by a switch from primarily MICU1-MICU2 to MICU1-MICU3 dimers during early life—and under hypoxic stress, by specific proteolytic degradation of MICU1, with potentially pathogenic consequences. To test these ideas, we will use a neuron-specific MICU1 knockout mouse which we found to display neurodegeneration characterized by mitochondrial Ca2+ overload, altered mitochondrial and neuronal ultrastructure, and motor and cognitive impairments. We have also generated mice with MICU2 and MICU3 loss and have obtained MICU1 and MICU2 patient-derived cells. We have set up advanced functional imaging and large volume 3D ultrastructure capacities. Thus, we are well positioned to study the neuronal pathogenesis associated with the loss of each or multiple MICU isoforms. In Aim#1 we will test the hypothesis that neurons derived from MICU1, MICU2 or MICU3-deficient mice and MICU1 and MICU2-deficient patient fibroblast-derived neurons have distinct impairments in Ca2+ signaling, mitochondrial dynamics and synaptic vesicle release. In Aim#2 we will test if neuronal MICU1/2/3 loss promotes mitochondrial oxidant production and oxidant-mediated cell injury. The results will help to decide if oxidants are relevant to MICU-linked neuronal/brain injury. In Aim#3 we will test the hypothesis that the relative abundance of the different MICU isoforms is dynamic in neurons during development, and the loss of each isoform specifically affects neuroanatomy and brain function primarily in the motor areas and the hippocampus. In Aim#4 we will test if ischemia-reperfusion induces YME1L-mediated turnover of MICU1 to promote mitochondrial Ca2+ overload injury and to sensitize the uniporter to inhibitors. These studies are expected to decide if MICU1 loss is a pathogenic component and therapy-modifying factor in brain hypoxia.