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Axonal Mrna Transport and Translation at a Glance Pabitra K © 2018. Published by The Company of Biologists Ltd | Journal of Cell Science (2018) 131, jcs196808. doi:10.1242/jcs.196808 CELL SCIENCE AT A GLANCE Axonal mRNA transport and translation at a glance Pabitra K. Sahoo1, Deanna S. Smith1, Nora Perrone-Bizzozero2 and Jeffery L. Twiss1,* ABSTRACT efforts are now uncovering new roles for locally synthesized proteins Localization and translation of mRNAs within different subcellular in neurological diseases and injury responses. In this Cell Science at domains provides an important mechanism to spatially and a Glance article and the accompanying poster, we provide an temporally introduce new proteins in polarized cells. Neurons make overview of how axonal mRNA transport and translation are use of this localized protein synthesis during initial growth, regulated, and discuss their emerging links to neurological regeneration and functional maintenance of their axons. Although disorders and neural repair. the first evidence for protein synthesis in axons dates back to 1960s, KEY WORDS: Axonal mRNA transport, Axonal mRNA translation, improved methodologies, including the ability to isolate axons to RNA granule, Post-transcriptional regulation, Protein synthesis, purity, highly sensitive RNA detection methods and imaging Ribonucleoprotein particle approaches, have shed new light on the complexity of the transcriptome of the axon and how it is regulated. Moreover, these Introduction Neurons are highly polarized cells with cytoplasmic extensions that 1Department of Biological Sciences, University of South Carolina, 715 Sumter St., can extend for millimeters, or even up to meters in large vertebrates. CLS 401, Columbia, SC 29208, USA. 2Department of Neurosciences, University of Axons and dendrites constitute the vast majority of the volume and New Mexico School of Medicine, 1 University of New Mexico, MSC08 4740, Albuquerque, NM 87131, USA. surface area of a neuron, and neurons use localized protein synthesis in these cytoplasmic extensions to spatially and temporally regulate *Author for correspondence ([email protected]) the protein content of these subcellular domains (Jung et al., 2014). D.S.S., 0000-0002-3014-641X; N.P., 0000-0002-7608-1332; J.L.T., 0000-0001- Axons provide long-range connections between neurons and their 7875-6682 targets that allow the brain, spinal cord and peripheral nerves to Journal of Cell Science 1 CELL SCIENCE AT A GLANCE Journal of Cell Science (2018) 131, jcs196808. doi:10.1242/jcs.196808 communicate. With the known transport rates of proteins, organelles 2013). Although mRNA protein-coding sequences (CDS) that and other macromolecules, the distal axon must respond to mediate axonal targeting have not yet been found for axonal environmental stimuli well before anything could be transported mRNAs, CDS motifs have been described in yeast (Kilchert and there from the cell body. Localized protein synthesis is one way to Spang, 2011), so it is likely that CDS motifs for axonal mRNA overcome this distance constraint, but the neuroscience community targeting will also be uncovered. In another commonality with largely overlooked the possibility of localized mRNA translation in dendrites and non-neuronal systems, RNA-binding proteins (RBPs) axons until recent years (Box 1). Generating proteins locally within binding to these motifs are necessary for axonal mRNA transport distal axons brings unique advantages for growth, survival and (Khalil et al., 2018; Korsak et al., 2016). Thus, the interaction of an function to these far reaches of the cytoplasm of a neuron, and recent mRNA with RBPs is sequence dependent, but consensus RNA unbiased analyses point to thousands of different mRNAs in axons. motifs unique to axonal mRNA targeting have yet to be found. In this Cell Science at a Glance article, we aim to summarize new Moreover, some axonal localization motifs can also target mRNAs advances in the field and point out where knowledge gaps exist. We into dendrites (Tiruchinapalli et al., 2003; Vuppalanchi et al., 2010), focus on the regulation of mRNA transport and translation and the which could reflect common uses for the encoded proteins in axons unique functions served from axonally synthesized proteins, and dendrites. highlighting how neuronal health is affected by these mechanisms The secondary structures of the RNA motifs are thought to where loss or gain of function result in disease or altered axon growth influence their interaction with RBPs (Gomes et al., 2014). The capacity. Obviously, it is not possible to cover the entire field in this ability to bioinformatically compare secondary structures across short article, so we refer the reader to several recent reviews for more RNA species is advancing, so common structural motifs may be detailed summaries (Batista and Hengst, 2016; Costa and Willis, discovered soon. Next-generation sequencing of RNAs from RBP 2017; Kar et al., 2018; Tasdemir-Yilmaz and Segal, 2016). immunoprecipitations also holds promise to uncover RBP- recognition motifs that are shared between axonal mRNAs, as How does the neuron know which mRNAs to localize into seen for recent work with motor axon transcriptomes (Rotem et al., axons? 2017). However, multiple RBPs can bind to the same mRNA and Just as RNAs are localized into dendrites and the subcellular regions impart different fates to the mRNA. Interactions of mRNAs with of non-neuronal cells, mRNA transport into axons is driven by RBPs likely begin in the nucleus, either co-transcriptionally or sequences inherent to the RNAs. These sequence motifs have most shortly after transcription, and the fate of an mRNA with regard to often been found in 3′ untranslated regions (UTR) of the mRNAs its subcellular mRNA localization is conferred by the sequential (Andreassi and Riccio, 2009; Gomes et al., 2014), but 5′UTR- binding of multiple RBPs. Evidence for this is the interaction of localization motifs have also been uncovered (Merianda et al., β-actin mRNA (ACTB) with zip code binding protein 2 (ZBP2, also called KHSRP and FUBP2) and ZBP1 (also called IGF2BP1 and IMP1) (Pan et al., 2007). Box 1. History of axonal mRNA translation As new knowledge of localization motifs emerges, it will need to be Early electron microscopy (EM) analyses of rodent brain showed interpreted in the context of multiple protein interactors and different evidence for polysomes at the base of dendritic spines in mature neuron types, as well as for the protein–protein interactions that occur hippocampus (Steward and Levy, 1982), which spurred decades of at different sites within the soma and along the axon. For example, research focusing on the role of dendritically synthesized proteins in recent work shows that γ-actin mRNA localizes into motor axons synaptic plasticity (Namjoshi and Raab-Graham, 2017). Those same (Moradi et al., 2017), whereas it appears to be restricted to the soma of early EM studies shed doubt on the possibility that axons can synthesize sensory and cortical neurons (Bassell et al., 1998; Zheng et al., 2001). proteins, as no polysomes were found in the axons of the mature hippocampus (Steward and Levy, 1982). Further doubt that translation In addition, CREB mRNA localizes to embryonic sensory axons, but occurs in axons came from instances of mRNAs having been detected in not axons of sympathetic neurons (Andreassi et al., 2010; Cox et al., hypothalamic and olfactory axons, yet ribosomes appeared to be lacking 2008). Axonal mRNA populations can similarly change with growth (Denis-Donini et al., 1998; Mohr and Richter, 1992). Thus, it was states and as the neuron matures (Gumy et al., 2011; Shigeoka et al., suggested that the translational machinery and mRNAs are excluded 2016; Taylor et al., 2009). In-depth axonal RNA profiles of different from vertebrate axons (Steward, 1997), even though biochemical neuronal subtypes will undoubtedly uncover more differences in evidence already pointed to the possibility of intra-axonal protein synthesis in the 1960s (Koenig, 1965a,b; Koenig, 1967a,b) and, axonal transcriptomes, physiological states and pathological states, shortly thereafter, EM evidence for ribosomes in axons of the PNS and it is important to keep in mind that differential gene expression as was published (Bunge, 1973; Tennyson, 1970; Yamada et al., 1971). well as combinations of RBPs could drive these. A series of critical studies taking advantage of the size of the squid giant axon as a tractable model to test for intra-axonal protein synthesis Mechanism for the regulation of axonal mRNA transport culminated in clear evidence for intra-axonal protein synthesis in those Just as with proteins and organelles, mRNAs are actively invertebrate neurons (Giuditta et al., 1980, 1986, 1991). Subsequent transported into axons by molecular motors. Because of the studies in vertebrates, and then mammals, showed that axons synthesize proteins even in adults (Perry and Fainzilber, 2014; Twiss unified polarity of microtubules in axons, the plus-end-directed and van Minnen, 2006). Fueled by recent technical and experimental kinesin motor proteins are used for long-range anterograde transport advances, the field has now moved from phenomenon to understanding in axons on microtubules, whereas myosin motor proteins are used the functions served by axonally synthesized proteins in neuronal for short-range transport on microfilaments (see poster) (Kalinski development, injury responses and disease states. Recent studies are et al., 2015b). Dynein motor-dependent retrograde movements have starting to uncover
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