Spinal muscular atrophy: Analysis of pathomechanistic impact of protective genetic modifiers in mouse models

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Spinal muscular atrophy (SMA) is a common neuromuscular disorder leading to early childhood lethality in about 50% of patients. Mutations in the SMN1 gene cause functional loss of the ¿-motor neurons (MNs) in the spinal cord mainly affecting development and maturation of neuromuscular junctions. Impaired synaptic transmission causes muscle weakness and atrophy of proximal voluntary muscles. The disease severity is mainly influenced by a copy gene, SMN2, which is aberrantly spliced lacking exon 7 in 90% of transcripts and, rarely, by additional genetic modifiers. In the past years we identified and functionally characterized the human genetic modifier plastin 3 (PLS3), while during the last funding period we identified a second modifier, denominated here as MOD2. Most importantly, only the discovery of both modifiers pointed us recently towards the main pathocellular disturbance in SMA.We analyzed a large SMA family with five SMN1-deleted individuals, who were fully asymptomatic despite carrying only four SMN2 copies, usually causing type II or III SMA. Transcriptome and linkage analysis unraveled MOD2 as a novel SMA protective modifier. All five asymptomatic individuals showed low MOD2 expression in comparison to the affected persons in the family and other independent SMA patients with four SMN2 copies. MOD2 is a neuronal calcium sensor protein, strongly expressed in brain and at NMJ level. Importantly, suppression of Mod2 restores SMA caused phenotype in cell culture and MN function across various SMA models, including zebrafish, worm, and, according to preliminary data, in mice. Within the next funding period we aim to unravel the impact of knock-down of Mod2 using a novel mouse model and its modifying effect on two different SMA models: a severe SMA model resembling a type I SMA patient and a milder SMA model resembling a type II patient. Detailed morphological, histological and functional analysis will be carried out. To unravel the impact of Mod2 knock-down on neuronal development and synaptic transmission, we will extensively analyze cultured MNs and hippocampal neurons and investigate synaptic neurotransmission using pH-sensitive GFP-reporter constructs and live cell imaging. Providing the required scientific proof for MOD2 suppression may allow its use in future SMA clinical trials.