The Gemin Associates of Survival Motor Neuron Are Required for Motor Function in Drosophila

Abstract

Membership of the survival motor neuron (SMN) complex extends to nine factors, including the SMN protein, the product of the spinal muscular atrophy (SMA) disease gene, Gemins 2–8 and Unrip. The best-characterised function of this macromolecular machine is the assembly of the Sm-class of uridine-rich small nuclear ribonucleoprotein (snRNP) particles and each SMN complex member has a key role during this process. So far, however, only little is known about the function of the individual Gemin components in vivo. Here, we make use of the Drosophila model organism to uncover loss-of-function phenotypes of Gemin2, Gemin3 and Gemin5, which together with SMN form the minimalistic fly SMN complex. We show that ectopic overexpression of the dead helicase Gem3ΔN mutant or knockdown of Gemin3 result in similar motor phenotypes, when restricted to muscle, and in combination cause lethality, hence suggesting that Gem3ΔN overexpression mimics a loss-of-function. Based on the localisation pattern of Gem3ΔN, we predict that the nucleus is the primary site of the antimorphic or dominant-negative mechanism of Gem3ΔN-mediated interference. Interestingly, phenotypes induced by human SMN overexpression in Drosophila exhibit similarities to those induced by overexpression of Gem3ΔN. Through enhanced knockdown we also uncover a requirement of Gemin2, Gemin3 and Gemin5 for viability and motor behaviour, including locomotion as well as flight, in muscle. Notably, in the case of Gemin3 and Gemin5, such function also depends on adequate levels of the respective protein in neurons. Overall, these findings lead us to speculate that absence of any one member is sufficient to arrest the SMN-Gemins complex function in a nucleocentric pathway, which is critical for motor function in vivo.

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

“…Overall, these findings show that viability and motor function are significantly disrupted on loss of Valette in the muscle compartment of the neuromuscular system. These phenotypes which overlap those described for Gaulos/Gemin4 (above), the other Gemins or SMN , reinforce our case favouring Valette as the bona fide Gemin8 orthologue in Drosophila .…”

“…We show a similar interaction profile in Drosophila . Notably, we reveal that Gaulos/Gemin4 and Valette/Gemin8 are essential for neuromuscular survival and function in line with reports describing a similar requirement for SMN and Gemin2/3/5 . Besides constituting evidence favouring these two proteins as the bona fide Drosophila orthologues of the respective Gemin counterparts in vertebrates, these findings also expose a novel in vivo function in the motor system for Gemin4 and Gemin8.…”

“…6C). Taken together, these findings reveal that similar to SMN and the known Gemin components of the Drosophila SMN complex [18,21,24], Gaulos is required for viability and optimal motor behaviour in the neuromuscular system. In this regard, we are very confident that Gaulos is the Drosophila orthologue of Gemin4.…”

“…We next asked whether loss of Valette induces phenotypes that are similar to those described for Gaulos/ Gemin4 (above) or the Gemin components of the SMN complex [18]. To this end, we employed an RNAi transgene (Vlet RNAi) targeting most of the coding sequence, hence causing a severe reduction in the expression level of Valette mRNA transcripts compared to controls (Fig.…”

Negative Cooperativity between Gemin2 and RNA provides Insights into RNA Selection and the SMN Complex’s Release in snRNP Assembly

“…Either smn or Gemin2 gene knockout causes early embryonic death in vertebrates, indicating the essential roles of the SMN complex in eukaryotic cells (17,18). Moreover, the deficiency of SMN causes human neurodegenerative disease spinal muscular atrophy (SMA) (19)(20)(21), and knockdown of Gemin2 causes motor neuron degeneration in zebrafish (22) and depresses motoric abilities in Drosophila (23), emphasizing the pathophysiological relevance of the Sm-core assembly pathway. Therefore, understanding the mechanism of Sm core assembly, especially at the second phase, is of great importance because of both its fundamental role in gene expression and its potential application in SMA therapy.…”