11 June 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 colocalize 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.
Akten B, Kye MJ, Hao LT, Wertz MH, Singh S, Nie D, Huang J, Merianda TT, Twiss JL, Beattie C, Steen JAJ, Sahin M. Interaction of survival of motor neuron (SMN) and HuD proteins with cpg15 mRNA rescues motor neuron axonal deficits. Proc Natl Acad Sci. 2011 Jun 7. Abstract