SMA-Causing Missense Mutations in Survival motor neuron (Smn) Display a Wide Range of Phenotypes When Modeled in Drosophila

Abstract

Mutations in the human survival motor neuron 1 (SMN) gene are the primary cause of spinal muscular atrophy (SMA), a devastating neuromuscular disorder. SMN protein has a well-characterized role in the biogenesis of small nuclear ribonucleoproteins (snRNPs), core components of the spliceosome. Additional tissue-specific and global functions have been ascribed to SMN; however, their relevance to SMA pathology is poorly understood and controversial. Using Drosophila as a model system, we created an allelic series of twelve Smn missense mutations, originally identified in human SMA patients. We show that animals expressing these SMA-causing mutations display a broad range of phenotypic severities, similar to the human disease. Furthermore, specific interactions with other proteins known to be important for SMN's role in RNP assembly are conserved. Intragenic complementation analyses revealed that the three most severe mutations, all of which map to the YG box self-oligomerization domain of SMN, display a stronger phenotype than the null allele and behave in a dominant fashion. In support of this finding, the severe YG box mutants are defective in self-interaction assays, yet maintain their ability to heterodimerize with wild-type SMN. When expressed at high levels, wild-type SMN is able to suppress the activity of the mutant protein. These results suggest that certain SMN mutants can sequester the wild-type protein into inactive complexes. Molecular modeling of the SMN YG box dimer provides a structural basis for this dominant phenotype. These data demonstrate that important structural and functional features of the SMN YG box are conserved between vertebrates and invertebrates, emphasizing the importance of self-interaction to the proper functioning of SMN.

Transcriptomic comparison of Drosophila snRNP biogenesis mutants reveals mutant-specific changes in pre-mRNA processing: implications for spinal muscular atrophy

“…SMA is, however, a hypomorphic condition, as complete loss of SMN activity is lethal in all organisms tested. To better model the disease in the fly, we previously engineered an allelic series of transgenic animals that express Smn missense mutations known to cause SMA in humans (Praveen et al 2012(Praveen et al , 2014 (Praveen et al 2014). These lines cover a range of phenotypic outcomes: Smn T205I is semi-viable (∼30% eclose as adults), Smn Y107C is pharatelethal (very few animals eclose), and Smn V72G mutants all die as pupae (Praveen et al 2014).…”

“…To better model the disease in the fly, we previously engineered an allelic series of transgenic animals that express Smn missense mutations known to cause SMA in humans (Praveen et al 2012(Praveen et al , 2014 (Praveen et al 2014). These lines cover a range of phenotypic outcomes: Smn T205I is semi-viable (∼30% eclose as adults), Smn Y107C is pharatelethal (very few animals eclose), and Smn V72G mutants all die as pupae (Praveen et al 2014). Importantly, in this system, the wild-type and mutant transgenes are integrated at the identical chromosomal locus and are expressed from the native promoter in an otherwise Smn null background (Praveen et al 2012(Praveen et al , 2014.…”

“…These lines cover a range of phenotypic outcomes: Smn T205I is semi-viable (∼30% eclose as adults), Smn Y107C is pharatelethal (very few animals eclose), and Smn V72G mutants all die as pupae (Praveen et al 2014). Importantly, in this system, the wild-type and mutant transgenes are integrated at the identical chromosomal locus and are expressed from the native promoter in an otherwise Smn null background (Praveen et al 2012(Praveen et al , 2014. Using qRT-PCR, we analyzed the alternative splicing patterns of the four transcripts described above.…”

“…The wild-type and mutant Smn transgenes are expressed using the native Smn promoter. Transgenic constructs were integrated into the PhiC31 landing site located at 86F8, which was previously recombined into an Smn X7 background (Praveen et al 2012(Praveen et al , 2014. For RNA-seq analysis, Smn transgenes in the Smn X7 null background were crossed to Smn X7 animals, and trans-heterozygous animals were put into TRIzol <12 h after pupation, as brown prepupa.…”

SMN - A chaperone for nuclear RNP social occasions?

“…25,106,107 Furthermore, mutations that completely disrupt SMN's ability to self-oligomerize display severe phenotypes in human SMA patients as well as in animal models. 86,[108][109][110][111][112][113] Despite this strong correlation, the composition and stoichiometry of the various complexes formed by SMN are not well understood. In vitro, SMN-Gem2 exists as a stable heterodimer that, for purposes of discussing higher order oligomerization, can be considered as a single structural unit.…”