11 Jun 2011

The protein survival of motor neuron (SMN), whose absence causes spinal muscular atrophy, has a new charge: an mRNA needed to make neuromuscular junctions. Together with the RNA-binding protein HuD, SMN picks up the mRNA for cpg15 (candidate plasticity-related gene 15) and totes it from the cell body to the ends of axons, according to a study released online June 7 by the Proceedings of the National Academy of Sciences USA. Researchers used extra cpg15 to rescue pathology in SMN-deficient zebrafish, and suggest cpg15 protein receptors might be druggable targets for treating motor neuron diseases.

Mutations in the SMN gene reduce the protein’s levels, leading to progressive muscle weakness, frequently in babies, and ultimately death, often in childhood. SMN has long been known to play a role in mRNA splicing (see ARF related news story on Zhang et al., 2008), but also, it associates with axons. Two recent studies found that SMN interacts with HuD, an RNA-stabilizing protein (Hubers et al., 2011; see ARF related news story on Fallini et al., 2011). Since SMN does not bind mRNA itself, it may help assemble ribonuclear complexes, suggested Wilfried Rossoll of Emory University in Atlanta, Georgia. The current work, led by first author Bikem Akten and senior authors Judith Steen and Mustafa Sahin of Children’s Hospital in Boston, adds cpg15 mRNA into the mix.

The researchers immunoprecipitated SMN from mouse primary embryonic cortical neurons and used mass spectrometry to identify HuD as a binding partner, along with a set of known SMN interactors. They confirmed that the two proteins co-localize in primary neuron cultures.

HuD was an interesting hit, Akten said, because it is an RNA binding protein that promotes synaptic activity. The researchers wondered what mRNAs might also partner with SMN and HuD. They immunoprecipitated both SMN and HuD from cortical neurons and looked for the RNAs they suspected of binding the pair: mRNAs essential for neural growth and HuD binding partners. The mRNA cpg15 came down with both HuD and SMN.

Also known as neuritin, cpg15 is expressed in post-mitotic neurons, and participates in the branching of motor neuron axons and formation of neuromuscular junctions. In vivo, the scientists found that cpg15 mRNAs dropped below normal in cortical neurites depleted of SMN.

The authors reasoned that if lack of cpg15 is part of the problem in SMN-deficient cells, then restoring cpg15 should alleviate the pathology. To test the idea, they teamed with Christine Beattie of Ohio State University in Columbus. Beattie works with zebrafish, knocking down SMN expression to create an SMA model for spinal muscular atrophy. In the fish embryos, motor neurons have shortened axons and abnormal axon branching (McWhorter et al., 2003). When the researchers overexpressed cpg15 in the zebrafish embryos, the defects were partially rescued, with fewer severely altered axons. “The axon projections looked much more like the wild-type,” Sahin said.

SMA cases range in severity, perhaps because of modifier genes that worsen or alleviate the effects of SMN loss, said Marco Passini of the Genzyme Corporation Science Center in Framingham, Massachusetts. SMN2 (Lefebvre et al., 1997) and plastin 3 (Oprea et al., 2008) are two known genetic modifiers of SMN pathology. Cpg15 may be a third.

SMN acts as a “taxicab” for cpg15 mRNA, delivering it to the axon tips so it can be on hand when the protein is needed. “The question is, What other mRNAs does the complex bring down the axon?” Passini asked. The study authors wondered the same thing. For now, it is not certain if cpg15 is the key mRNA that needs SMN, or if it is one of many SMN-reliant mRNAs, Akten said, but she suspects the latter.

Treating SMA would require replacing either SMN itself or its functions. Sahin noted that cpg15 is secreted and presumably binds to a receptor. Either the protein or the receptor might be accessible to small molecules, he suggested, which could be simpler than trying to replace intracellular SMN.

Amyotrophic lateral sclerosis (ALS) is another disease of motor neurons, although it begins much later in life. The RNA-binding proteins TDP-43 and FUS have already been implicated in ALS (see ARF related news story on Kwiatkowski et al., 2009 and Vance et al., 2009), and TDP-43 affects SMN localization (see ARF related news story on Shan et al., 2010). Could cpg15 be involved in ALS as well? It is hard to guess, because the cellular processes of adult motor neurons differ greatly from developing ones, said Claudia Fallini of Emory University. Cpg15 mRNA has not yet made the list of published TDP-43 targets (see ARF related news story on Sephton et al., 2011 and ARF related news story on Tollervey et al., 2011 and Polymenidou et al., 2011).—Amber Dance

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References

News Citations

1. Infant Killer SMA a Splicing Disease? 16 May 2008
2. All Aboard? SMA Protein Predicted to Organize Axonal RNA Transport 11 Mar 2011
3. New Gene for ALS: RNA Regulation May Be Common Culprit 27 Feb 2009
4. Latest TDP-43 Mouse Unites ALS and SMA Pathways 8 Sep 2010
5. San Diego: TDP-43 Targets Loom Large—But Where’s the Bull’s Eye? 25 Nov 2010
6. CLIPs of TDP-43 Provide a Glimpse Into Pathology 9 Mar 2011

Paper Citations

1. Zhang Z, Lotti F, Dittmar K, Younis I, Wan L, Kasim M, Dreyfuss G. SMN deficiency causes tissue-specific perturbations in the repertoire of snRNAs and widespread defects in splicing. Cell. 2008 May 16;133(4):585-600. PubMed.
2. Hubers L, Valderrama-Carvajal H, Laframboise J, Timbers J, Sanchez G, Côté J. HuD interacts with survival motor neuron protein and can rescue spinal muscular atrophy-like neuronal defects. Hum Mol Genet. 2011 Feb 1;20(3):553-79. PubMed.
3. Fallini C, Zhang H, Su Y, Silani V, Singer RH, Rossoll W, Bassell GJ. The survival of motor neuron (SMN) protein interacts with the mRNA-binding protein HuD and regulates localization of poly(A) mRNA in primary motor neuron axons. J Neurosci. 2011 Mar 9;31(10):3914-25. PubMed.
4. McWhorter ML, Monani UR, Burghes AH, Beattie CE. Knockdown of the survival motor neuron (Smn) protein in zebrafish causes defects in motor axon outgrowth and pathfinding. J Cell Biol. 2003 Sep 1;162(5):919-31. PubMed.
5. Lefebvre S, Burlet P, Liu Q, Bertrandy S, Clermont O, Munnich A, Dreyfuss G, Melki J. Correlation between severity and SMN protein level in spinal muscular atrophy. Nat Genet. 1997 Jul;16(3):265-9. PubMed.
6. Oprea GE, Kröber S, McWhorter ML, Rossoll W, Müller S, Krawczak M, Bassell GJ, Beattie CE, Wirth B. Plastin 3 is a protective modifier of autosomal recessive spinal muscular atrophy. Science. 2008 Apr 25;320(5875):524-7. PubMed.
7. Kwiatkowski TJ Jr, Bosco DA, Leclerc AL, Tamrazian E, Vanderburg CR, Russ C, Davis A, Gilchrist J, Kasarskis EJ, Munsat T, Valdmanis P, Rouleau GA, Hosler BA, Cortelli P, de Jong PJ, Yoshinaga Y, Haines JL, Pericak-Vance MA, Yan J, Ticozzi N, Siddique T, McKenna-Yasek D, Sapp PC, Horvitz HR, Landers JE, Brown RH Jr. Mutations in the FUS/TLS gene on chromosome 16 cause familial amyotrophic lateral sclerosis. Science. 2009 Feb 27;323(5918):1205-8. PubMed.
8. Vance C, Rogelj B, Hortobágyi T, De Vos KJ, Nishimura AL, Sreedharan J, Hu X, Smith B, Ruddy D, Wright P, Ganesalingam J, Williams KL, Tripathi V, Al-Saraj S, Al-Chalabi A, Leigh PN, Blair IP, Nicholson G, de Belleroche J, Gallo JM, Miller CC, Shaw CE. Mutations in FUS, an RNA processing protein, cause familial amyotrophic lateral sclerosis type 6. Science. 2009 Feb 27;323(5918):1208-11. PubMed.
9. Shan X, Chiang PM, Price DL, Wong PC. Altered distributions of Gemini of coiled bodies and mitochondria in motor neurons of TDP-43 transgenic mice. Proc Natl Acad Sci U S A. 2010 Sep 14;107(37):16325-30. Epub 2010 Aug 24 PubMed.
10. Sephton CF, Cenik C, Kucukural A, Dammer EB, Cenik B, Han Y, Dewey CM, Roth FP, Herz J, Peng J, Moore MJ, Yu G. Identification of neuronal RNA targets of TDP-43-containing ribonucleoprotein complexes. J Biol Chem. 2011 Jan 14;286(2):1204-15. PubMed.
11. Tollervey JR, Curk T, Rogelj B, Briese M, Cereda M, Kayikci M, König J, Hortobágyi T, Nishimura AL, Zupunski V, Patani R, Chandran S, Rot G, Zupan B, Shaw CE, Ule J. Characterizing the RNA targets and position-dependent splicing regulation by TDP-43. Nat Neurosci. 2011 Apr;14(4):452-8. PubMed.
12. Polymenidou M, Lagier-Tourenne C, Hutt KR, Huelga SC, Moran J, Liang TY, Ling SC, Sun E, Wancewicz E, Mazur C, Kordasiewicz H, Sedaghat Y, Donohue JP, Shiue L, Bennett CF, Yeo GW, Cleveland DW. Long pre-mRNA depletion and RNA missplicing contribute to neuronal vulnerability from loss of TDP-43. Nat Neurosci. 2011 Apr;14(4):459-68. PubMed.

Further Reading

Papers

1. Ishihara T, Hong M, Zhang B, Nakagawa Y, Lee MK, Trojanowski JQ, Lee VM. Age-dependent emergence and progression of a tauopathy in transgenic mice overexpressing the shortest human tau isoform. Neuron. 1999 Nov;24(3):751-62. PubMed.
2. Murray LM, Lee S, Bäumer D, Parson SH, Talbot K, Gillingwater TH. Pre-symptomatic development of lower motor neuron connectivity in a mouse model of severe spinal muscular atrophy. Hum Mol Genet. 2010 Feb 1;19(3):420-33. PubMed.
3. Rossoll W, Jablonka S, Andreassi C, Kröning AK, Karle K, Monani UR, Sendtner M. Smn, the spinal muscular atrophy-determining gene product, modulates axon growth and localization of beta-actin mRNA in growth cones of motoneurons. J Cell Biol. 2003 Nov 24;163(4):801-12. PubMed.
4. Rossoll W, Kröning AK, Ohndorf UM, Steegborn C, Jablonka S, Sendtner M. Specific interaction of Smn, the spinal muscular atrophy determining gene product, with hnRNP-R and gry-rbp/hnRNP-Q: a role for Smn in RNA processing in motor axons?. Hum Mol Genet. 2002 Jan 1;11(1):93-105. PubMed.
5. Fan L, Simard LR. Survival motor neuron (SMN) protein: role in neurite outgrowth and neuromuscular maturation during neuronal differentiation and development. Hum Mol Genet. 2002 Jul 1;11(14):1605-14. PubMed.
6. Gubitz AK, Feng W, Dreyfuss G. The SMN complex. Exp Cell Res. 2004 May 15;296(1):51-6. PubMed.
7. Zhang HL, Pan F, Hong D, Shenoy SM, Singer RH, Bassell GJ. Active transport of the survival motor neuron protein and the role of exon-7 in cytoplasmic localization. J Neurosci. 2003 Jul 23;23(16):6627-37. PubMed.

News

1. Research Brief: Researchers Solicit SMN Understudy to Treat SMA 9 Mar 2011
2. Is Spinal Muscular Atrophy a Disease of Sensory Neurons? 11 Feb 2011
3. Motor Neuron Disease Risk Factors: Fresh miRNA Clue, KIFAP3 Letdown 29 Jun 2010
4. Baby Steps for Gene Therapy? Fatal Infant Disease Cured in Mice 2 Mar 2010
5. Infant Killer SMA a Splicing Disease? 16 May 2008
6. All Aboard? SMA Protein Predicted to Organize Axonal RNA Transport 11 Mar 2011
7. New Gene for ALS: RNA Regulation May Be Common Culprit 27 Feb 2009
8. Latest TDP-43 Mouse Unites ALS and SMA Pathways 8 Sep 2010
9. San Diego: TDP-43 Targets Loom Large—But Where’s the Bull’s Eye? 25 Nov 2010
10. CLIPs of TDP-43 Provide a Glimpse Into Pathology 9 Mar 2011