SMN complex member Gemin3 self-interacts and has a functional relationship with ALS-linked proteins TDP-43, FUS and Sod1.

The predominant motor neuron disease in infants and adults is spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS), respectively. SMA is caused by insufficient levels of the Survival Motor Neuron (SMN) protein, which operates as part of the multiprotein SMN complex that includes the DEAD-box RNA helicase Gemin3/DDX20/DP103. C9orf72, SOD1, TDP-43 and FUS are ranked as the four major genes causing familial ALS. Accumulating evidence has revealed a surprising molecular overlap between SMA and ALS. Here, we ask the question of whether Drosophila can also be exploited to study shared pathogenic pathways. Focusing on motor behaviour, muscle mass and survival, we show that disruption of either TBPH/TDP-43 or Caz/FUS enhance defects associated with Gemin3 loss-of-function. Gemin3-associated neuromuscular junction overgrowth was however suppressed. Sod1 depletion had a modifying effect in late adulthood. We also show that Gemin3 self-interacts and Gem3ΔN, a helicase domain deletion mutant, retains the ability to interact with its wild-type counterpart. Importantly, mutant:wild-type dimers are favoured more than wild-type:wild-type dimers. In addition to reinforcing the link between SMA and ALS, further exploration of mechanistic overlaps is now possible in a genetically tractable model organism. Notably, Gemin3 can be elevated to a candidate for modifying motor neuron degeneration.

Novel interactors of the Drosophila Survival Motor Neuron (SMN) Complex suggest its full conservation.

The Spinal Muscular Atrophy disease protein Survival Motor Neuron (SMN) operates as part of a multiprotein complex whose components also include Gemins 2-8 and Unrip. The fruit fly Drosophila melanogaster is thought to have a slightly smaller SMN complex comprised of SMN, Gemin2/3/5 and, possibly, Unrip. Based upon in vivo interaction methods, we have identified novel interacting partners of the Drosophila SMN complex with homologies to Gemin4/6/7/8. The Gemin4 and Gemin8 orthologues are required for neuromuscular function and survival. The Gemin6/7/Unrip module can be recruited via the SMN-associated Gemin8, hence mirroring the human SMN complex architecture. Our findings lead us to propose that an elaborate SMN complex that is typical in metazoans is also present in Drosophila.

Disruption of snRNP biogenesis factors Tgs1 and pICln induces phenotypes that mirror aspects of SMN-Gemins complex perturbation in Drosophila, providing new insights into spinal muscular atrophy.

The neuromuscular disorder, spinal muscular atrophy (SMA), results from insufficient levels of the survival motor neuron (SMN) protein. Together with Gemins 2-8 and Unrip, SMN forms the large macromolecular SMN-Gemins complex, which is known to be indispensable for chaperoning the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). It remains unclear whether disruption of this function is responsible for the selective neuromuscular degeneration in SMA. In the present study, we first show that loss of wmd, the Drosophila Unrip orthologue, has a negative impact on the motor system. However, due to lack of a functional relationship between wmd/Unrip and Gemin3, it is likely that Unrip joined the SMN-Gemins complex only recently in evolution. Second, we uncover that disruption of either Tgs1 or pICln, two cardinal players in snRNP biogenesis, results in viability and motor phenotypes that closely resemble those previously uncovered on loss of the constituent members of the SMN-Gemins complex. Interestingly, overexpression of both factors leads to motor dysfunction in Drosophila, a situation analogous to that of Gemin2. Toxicity is conserved in the yeast S. pombe where pICln overexpression induces a surplus of Sm proteins in the cytoplasm, indicating that a block in snRNP biogenesis is partly responsible for this phenotype. Importantly, we show a strong functional relationship and a physical interaction between Gemin3 and either Tgs1 or pICln. We propose that snRNP biogenesis is the pathway connecting the SMN-Gemins complex to a functional neuromuscular system, and its disturbance most likely leads to the motor dysfunction that is typical in SMA.

Genetic Interactions between the Members of the SMN-Gemins Complex in Drosophila.

The SMN-Gemins complex is composed of Gemins 2-8, Unrip and the survival motor neuron (SMN) protein. Limiting levels of SMN result in the neuromuscular disorder, spinal muscular atrophy (SMA), which is presently untreatable. The most-documented function of the SMN-Gemins complex concerns the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). Despite multiple genetic studies, the Gemin proteins have not been identified as prominent modifiers of SMN-associated mutant phenotypes. In the present report, we make use of the Drosophila model organism to investigate whether viability and motor phenotypes associated with a hypomorphic Gemin3 mutant are enhanced by changes in the levels of SMN, Gemin2 and Gemin5 brought about by various genetic manipulations. We show a modifier effect by all three members of the minimalistic fly SMN-Gemins complex within the muscle compartment of the motor unit. Interestingly, muscle-specific overexpression of Gemin2 was by itself sufficient to depress normal motor function and its enhanced upregulation in all tissues leads to a decline in fly viability. The toxicity associated with increased Gemin2 levels is conserved in the yeast S. pombe in which we find that the cytoplasmic retention of Sm proteins, likely reflecting a block in the snRNP assembly pathway, is a contributing factor. We propose that a disruption in the normal stoichiometry of the SMN-Gemins complex depresses its function with consequences that are detrimental to the motor system.