title: Molecular determinants of survival motor neuron (SMN) protein cleavage by the calcium-activated protease, calpain.
abstract: Spinal muscular atrophy (SMA) is a leading genetic cause of childhood mortality, caused by reduced levels of survival motor neuron (SMN) protein. SMN functions as part of a large complex in the biogenesis of small nuclear ribonucleoproteins (snRNPs). It is not clear if defects in snRNP biogenesis cause SMA or if loss of some tissue-specific function causes disease. We recently demonstrated that the SMN complex localizes to the Z-discs of skeletal and cardiac muscle sarcomeres, and that SMN is a proteolytic target of calpain. Calpains are implicated in muscle and neurodegenerative disorders, although their relationship to SMA is unclear. Using mass spectrometry, we identified two adjacent calpain cleavage sites in SMN, S192 and F193. Deletion of small motifs in the region surrounding these sites inhibited cleavage. Patient-derived SMA mutations within SMN reduced calpain cleavage. SMN(D44V), reported to impair Gemin2 binding and amino-terminal SMN association, drastically inhibited cleavage, suggesting a role for these interactions in regulating calpain cleavage. Deletion of A188, a residue mutated in SMA type I (A188S), abrogated calpain cleavage, highlighting the importance of this region. Conversely, SMA mutations that interfere with self-oligomerization of SMN, Y272C and SMNΔ7, had no effect on cleavage. Removal of the recently-identified SMN degron (Δ268-294) resulted in increased calpain sensitivity, suggesting that the C-terminus of SMN is important in dictating availability of the cleavage site. Investigation into the spatial determinants of SMN cleavage revealed that endogenous calpains can cleave cytosolic, but not nuclear, SMN. Collectively, the results provide insight into a novel aspect of the post-translation regulation of SMN.
authors.affiliation: Program in Molecular Biology and Biotechnology, Department of Biology, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina, United States of America.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Fuentes
authors.forename: Molly S
authors.initials: MS
authors.lastname: Strayer
authors.forename: A Gregory
authors.initials: AG
authors.lastname: Matera
mesh_terms: D002118:Calcium / Q000737:chemistry*; D002154:Calpain / Q000737:chemistry*; D045744:Cell Line, Tumor; D002474:Cell-Free System; D004247:DNA / Q000378:metabolism; D004252:DNA Mutational Analysis; D019281:Dimerization; D006801:Humans; D018482:Muscle, Skeletal / Q000166:cytology; D009206:Myocardium / Q000166:cytology; D011392:Proline / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D017411:Ribonucleoproteins, Small Nuclear / Q000737:chemistry; D013347:Subcellular Fractions / Q000378:metabolism; D055533:Survival of Motor Neuron 1 Protein / Q000378:metabolism*; D014018:Tissue Distribution
chemical_list: D011994:Recombinant Proteins; D017411:Ribonucleoproteins, Small Nuclear; D055533:Survival of Motor Neuron 1 Protein; D004247:DNA; D011392:Proline; D002154:Calpain; D002118:Calcium