Composition of the Survival Motor Neuron (SMN) Complex in Drosophila melanogaster

AbstractSpinal Muscular Atrophy (SMA) is caused by homozygous mutations in the human survival motor neuron 1 (SMN1) gene. SMN protein has a well-characterized role in the biogenesis of small nuclear ribonucleoproteins (snRNPs), core components of the spliceosome. SMN is part of an oligomeric complex with core binding partners, collectively called Gemins. Biochemical and cell biological studies demonstrate that certain Gemins are required for proper snRNP assembly and transport. However, the precise functions of most Gemins are unknown. To gain a deeper understanding of the SMN complex in the context of metazoan evolution, we investigated the composition of the SMN complex in Drosophila melanogaster. Using a stable transgenic line that exclusively expresses Flag-tagged SMN from its native promoter, we previously found that Gemin2, Gemin3, Gemin5, and all nine classical Sm proteins, including Lsm10 and Lsm11, co-purify with SMN. Here, we show that CG2941 is also highly enriched in the pulldown. Reciprocal co-immunoprecipitation reveals that epitope-tagged CG2941 interacts with endogenous SMN in Schneider2 cells. Bioinformatic comparisons show that CG2941 shares sequence and structural similarity with metazoan Gemin4. Additional analysis shows that three other genes (CG14164, CG31950 and CG2371) are not orthologous to Gemins 6-7-8, respectively, as previously suggested. In D.melanogaster, CG2941 is located within an evolutionarily recent genomic triplication with two other nearly identical paralogous genes (CG32783 and CG32786). RNAi-mediated knockdown of CG2941 and its two close paralogs reveals that Gemin4 is essential for organismal viability.

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

Scientific Reports ◽

10.1038/s41598-019-53508-4 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 4

Author(s):

Rebecca Cacciottolo ◽

Joanna Ciantar ◽

Maia Lanfranco ◽

Rebecca M. Borg ◽

Neville Vassallo ◽

...

Keyword(s):

Motor Neuron ◽

Muscular Atrophy ◽

Model Organism ◽

Survival Motor Neuron ◽

Helicase Domain ◽

Loss Of Function ◽

Dead Box ◽

Smn Complex ◽

Wild Type Counterpart ◽

Smn Protein

AbstractThe 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.

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A Functional Interaction between the Survival Motor Neuron Complex and RNA Polymerase II

The Journal of Cell Biology ◽

10.1083/jcb.152.1.75 ◽

2001 ◽

Vol 152 (1) ◽

pp. 75-86 ◽

Cited By ~ 154

Author(s):

Livio Pellizzoni ◽

Bernard Charroux ◽

Juri Rappsilber ◽

Matthias Mann ◽

Gideon Dreyfuss

Keyword(s):

Rna Polymerase ◽

Motor Neuron ◽

Rna Polymerase Ii ◽

Rna Helicase ◽

Survival Motor Neuron ◽

Dominant Negative Mutant ◽

Pol Ii ◽

Smn Complex

The survival motor neuron (SMN) protein, the protein product of the spinal muscular atrophy (SMA) disease gene, plays a role in the assembly and regeneration of small nuclear ribonucleoproteins (snRNPs) and spliceosomes. By nanoelectrospray mass spectrometry, we identified RNA helicase A (RHA) as an SMN complex–associated protein. RHA is a DEAH box RNA helicase which binds RNA polymerase II (pol II) and reportedly functions in transcription. SMN interacts with RHA in vitro, and this interaction is impaired in mutant SMNs found in SMA patients. Coimmunoprecipitation demonstrated that the SMN complex is associated with pol II, snRNPs, and RHA in vivo. In vitro experiments suggest that RHA mediates the association of SMN with the COOH-terminal domain of pol II. Moreover, transfection of cells with a dominant negative mutant of SMN, SMNΔN27, causes accumulation of pol II, snRNPs, and RHA in nuclear structures that contain the known markers of gems and coiled bodies, and inhibits RNA pol I and pol II transcription in vivo. These findings indicate a functional as well as physical association of the SMN complex with pol II and suggest a role for the SMN complex in the assembly of the pol II transcription/processing machinery.

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