Abstract
Injury to axons in the peripheral nervous system induces rapid and local regenerative responses to form a new growth cone, and to generate a retrogradely transporting injury signal. The evidence for essential roles of intra-axonal protein synthesis during regeneration is now compelling. MicroRNA (miRNA) has recently been recognized as a prominent player in post-transcriptional regulation of axonal protein synthesis. Here, we directly contrast temporal changes of miRNA levels in the sciatic nerve following injury, as compared to those in an uninjured nerve using deep sequencing. Small RNAs (<200 nucleotides in length) were fractionated from the proximal nerve stumps to improve the representation of differential miRNA levels. Of 141 axoplasmic miRNAs annotated, 63 rat miRNAs showed significantly differential levels at five time points following injury, compared to an uninjured nerve. The differential changes in miRNA levels responding to injury were processed for hierarchical clustering analyses, and used to predict target mRNAs by Targetscan and miRanda. By overlapping these predicted targets with 2,924 axonally localizing transcripts previously reported, the overlapping set of 214 transcripts was further analyzed by the Gene Ontology enrichment and Ingenuity Pathway Analyses. These results suggest the possibility that the potential targets for these miRNAs play key roles in numerous neurological functions involved in ER stress response, cytoskeleton dynamics, vesicle formation, and neuro-degeneration and-regeneration. Finally, our results suggest that miRNAs could play a direct role in regenerative response and may be manipulated to promote regenerative ability of injured nerves.
Highlights
Axonal injury to the peripheral nervous system (PNS) triggers active translation of the localized transcripts that have been transported into axons from the cell body [1,2,3,4]
We find that axoplasmic miRNA levels in rat sciatic nerve are altered, upon injury
By integrating predicted potential target mRNAs and 2,924 transcripts known to be localized to distal axons of DRG neurons [27], we suggest the possibility that the targets of the intra-axonal miRNAs are directly related to multiple biological and neurological functions including regenerative responses following injury
Summary
Axonal injury to the peripheral nervous system (PNS) triggers active translation of the localized transcripts that have been transported into axons from the cell body [1,2,3,4]. Verma et al [13] has shown that axonally synthesized proteins are needed for the reformation of growth cones, which are critical for subsequent regeneration of injured axons These localized mRNAs are translationally dormant in the axon until recruited into translation machinery following injury. MicroRNAs (miRNAs) are a class of small regulatory non-coding RNAs (~22 nt long) They have been proposed to negatively regulate translation in the nervous system at the posttranscriptional level in a sequence-specific manner [14,15,16,17,18,19,20]. Several groups have further shown temporal changes of miRNA expression in the spinal cord, as well as in the sciatic nerve following injury [20, 24,25,26] These studies suggest direct roles of miRNAs in nerve regeneration via regulating protein synthesis. By integrating predicted potential target mRNAs and 2,924 transcripts known to be localized to distal axons of DRG neurons [27], we suggest the possibility that the targets of the intra-axonal miRNAs are directly related to multiple biological and neurological functions including regenerative responses following injury
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