155 | Mitochondrial dynamics and therapeutic insights in spinal muscular atrophy: effects of MFN2 boosting in motoneurons and human fibroblasts

Francesco Paolo Zummo¹, Serena Stanga¹, Letizia Paladino², Maria Beltran³, Francesco Fedeli⁴, Maria Rosa Soler³, Fabio Bucchieri⁵, Marina Boido¹ | ¹Department of Neuroscience “Rita Levi Montalcini”, Neuroscience Institute Cavalieri Ottolenghi, University of Turin, Italy; ²E-Campus University, Palermo, Italy; ³Neuronal Signaling Unit, Experimental Medicine Department, University of Lleida-IRBLleida, Lleida, Spain; ⁴University of Pavia, Italy; ⁵Department of Biomedicine, Neuroscience and Advanced Diagnostics, University of Palermo, Italy.

Mitochondria form a dynamic network whose spatial architecture is governed by the balance between fusion and fission, known as mitochondrial dynamics. These two opposing processes are essential for mitochondrial turnover: fusion, mediated by MFN2 and OPA1 regulators, promotes mitochondrial biogenesis, whereas fission, driven by DRP1, commits damaged mitochondria to their selective elimination. Spinal Muscular Atrophy (SMA) is a neuromuscular disorder in which a deficiency in the SMN (Survival Motor Neuron) protein leads to motor neuron (MN) degeneration. Since the SMN protein is involved in mitochondrial homeostasis, several SMA models have shown mitochondrial dysfunction, including an imbalance in mitochondrial dynamics. This study first explored the state of mitochondrial dynamics in soma and neurites of early-stage MNs differentiated from SMA patient-derived iPSCs and primary MNs from the SMNΔ7 mouse model. In both models, the mitochondrial mass was significantly reduced, specifically in the neuritic compartment. Notably, the analysis of fusion and fission regulators exhibited a moderate imbalance toward fission, detected exclusively in neurites through the in silico mitochondrial network analysis (MiNA). Interestingly, the treatment with an MFN2 activator, promoting fusion within the network, was able to enhance mitochondrial content and increase SMN protein levels in SMA human MNs. Although MNs seem to be the most sensitive cells to suffer from the low level of SMN, nowadays SMA is considered a multisystemic disease. Therefore, we also investigated mitochondrial dynamics at the peripheral level using human fibroblasts derived from SMA patients. As in SMA MNs, the analysis of fusion and fission regulators revealed a shift toward fission; in fibroblasts, however, this imbalance was associated with functional rather than content-related mitochondrial alterations. Surprisingly, also in SMA human fibroblasts, the fusion-promoting compound increased mitochondrial content and SMN protein levels, together with improvements in mitochondrial membrane potential and ATP synthase levels in a dose-dependent manner. Although the regulation of mitochondrial dynamics is a highly sensitive process, these findings highlight how increasing mitochondrial fusion in SMA can positively modulate the organelle turnover, enhancing mitochondrial content and function, and preserving SMN expression. Collectively, the beneficial effects of this treatment across two distinct SMA models support MFN2 boosting as a promising therapeutic strategy, particularly in combination with already approved SMA treatments.