Axonal Protein Synthesis Provides a Mechanism for Localized Regulation at an Intermediate Target

Department of Molecular and Cell Biology, Harvard University, Cambridge, Massachusetts, United States
Cell (Impact Factor: 32.24). 08/2002; 110(2):223-35. DOI: 10.1016/S0092-8674(02)00813-9
Source: PubMed


As axons grow past intermediate targets, they change their responsiveness to guidance cues. Local upregulation of receptor expression is involved, but the mechanisms for this are not clear. Here protein synthesis is traced within individual axons by introducing RNAs encoding visualizable reporters. Individual severed axons and growth cones can translate proteins and also export them to the cell surface. As axons reach the spinal cord midline, EphA2 is among the receptors upregulated on at least some distal axon segments. Midline reporter upregulation is recapitulated by part of the EphA2 mRNA 3' untranslated region, which is highly conserved and includes known translational control sequences. These results show axons contain all the machinery for protein translation and cell surface expression, and they reveal a potentially general and flexible RNA-based mechanism for regulation localized within a subregion of the axon.

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    • "The regulatory elements that control mRNA localization must be encoded in the mRNA itself, and indeed, the 3′-UTR is where many localization elements lie. The cis-elements can be primary nucleotide sequences, such as those in the 3′-UTRs of β-actin, RhoA, EphA2, CoxIV, and Impa-1 mRNAs (Andreassi et al., 2010; Aschrafi et al., 2010; Bassell et al., 1998; Brittis et al., 2002; Zhang et al., 2001), or secondary structures such as the hairpins found in the 3′-UTR of bicoid mRNA in Drosophila oocytes (Macdonald and Struhl, 1988) and Ash1 mRNA in the budding yeast (Chartrand et al., 1999). Cis-elements, however, can also be localized to the 5′-UTRs as in kor mRNA (Tsai et al., 2007); to the protein-coding sequence as in the target mRNAs of the RBP fragile X mental retardation protein (FMRP) (which is encoded by Fmr1 gene) (Ascano et al., 2012; Darnell et al., 2011) and Robo3 mRNA (Kuwako et al., 2010); or to introns as in some dendritically targeted mRNAs (Buckley et al., 2011). "
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    ABSTRACT: The subcellular position of a protein is a key determinant of its function. Mounting evidence indicates that RNA localization, where specific mRNAs are transported subcellularly and subsequently translated in response to localized signals, is an evolutionarily conserved mechanism to control protein localization. On-site synthesis confers novel signaling properties to a protein and helps to maintain local proteome homeostasis. Local translation plays particularly important roles in distal neuronal compartments, and dysregulated RNA localization and translation cause defects in neuronal wiring and survival. Here, we discuss key findings in this area and possible implications of this adaptable and swift mechanism for spatial control of gene function.
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    • "Such local protein translation is used as a mechanism to generate a focal response to a restricted stimulus. In developing neurons, guidance cues and growth factors can stimulate the translation of proteins in axonal growth cones that are used to mediate turning responses, promote growth or serve as retrograde messengers (Campbell and Holt, 2001; Zheng et al., 2001; Brittis et al., 2002; Wu et al., 2005; Leung et al., 2006; Yao et al., 2006; Cox et al., 2008; Kundel et al., 2009; Perry et al., 2012). Outside of growth cones, synthesis in axons is poorly understood (Deglincerti and Jaffrey, 2012). "
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    ABSTRACT: Axonal growth cones synthesize proteins during development and in response to injury in adult animals. Proteins locally translated in axons are used to generate appropriate responses to guidance cues, contribute to axon growth, and can serve as retrograde messengers. In addition to growth cones, mRNAs and translational machinery are also found along the lengths of axons where synapses form en passant, but contributions of intra-axonal translation to developing synapses are poorly understood. Here, we engineered a subcellular-targeting translational repressor to inhibit mRNA translation in axons, and we used this strategy to investigate presynaptic contributions of cap-dependent protein translation to developing CNS synapses. Our data show that intra-axonal mRNA translation restrains synaptic vesicle recycling pool size and that one target of this regulation is p35, a Cdk5 activating protein. Cdk5/p35 signaling regulates the size of vesicle recycling pools, p35 levels diminish when cap-dependent translation is repressed, and restoring p35 levels rescues vesicle recycling pools from the effects of spatially targeted translation repression. Together our findings show that intra-axonal synthesis of p35 is required for normal vesicle recycling in developing neurons, and that targeted translational repression provides a novel strategy to investigate extrasomal protein synthesis in neurons. © 2013 Wiley Periodicals, Inc. Develop Neurobiol, 2013.
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    • "As shown in Figure 5A, punctate staining (about 200 foci per neurite) along neuritic processes was observed for each of the four mRNAs, indicating that they are efficiently transported in NSC-34 neurites. Under the same experimental conditions, only a few spots (about 10 per neurite) (data not shown) were observed with probes specific for H1f0 transcripts, whose localization is restricted to the cell body (Brittis et al. 2002). Variable fractions of the RNA spots were found to colocalize with the SMN protein. "
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    ABSTRACT: Spinal muscular atrophy is a neuromuscular disease resulting from mutations in the SMN1 gene, which encodes the survival motor neuron (SMN) protein. SMN is part of a large complex that is essential for the biogenesis of spliceosomal small nuclear RNPs. SMN also colocalizes with mRNAs in granules that are actively transported in neuronal processes, supporting the hypothesis that SMN is involved in axonal trafficking of mRNPs. Here, we have performed a genome-wide analysis of RNAs present in complexes containing the SMN protein and identified more than 200 mRNAs associated with SMN in differentiated NSC-34 motor neuron-like cells. Remarkably, ∼30% are described to localize in axons of different neuron types. In situ hybridization and immuno-fluorescence experiments performed on several candidates indicate that these mRNAs colocalize with the SMN protein in neurites and axons of differentiated NSC-34 cells. Moreover, they localize in cell processes in an SMN-dependent manner. Thus, low SMN levels might result in localization deficiencies of mRNAs required for axonogenesis.
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