miR-182 Regulates Slit2-Mediated Axon Guidance by Modulating the Local Translation of a Specific mRNA

Abstract

SummaryDuring brain wiring, cue-induced axon behaviors such as directional steering and branching are aided by localized mRNA translation. Different guidance cues elicit translation of subsets of mRNAs that differentially regulate the cytoskeleton, yet little is understood about how specific mRNAs are selected for translation. MicroRNAs (miRNAs) are critical translational regulators that act through a sequence-specific mechanism. Here, we investigate the local role of miRNAs in mRNA-specific translation during pathfinding of Xenopus laevis retinal ganglion cell (RGC) axons. Among a rich repertoire of axonal miRNAs, miR-182 is identified as the most abundant. Loss of miR-182 causes RGC axon targeting defects in vivo and impairs Slit2-induced growth cone (GC) repulsion. We find that miR-182 targets cofilin-1 mRNA, silencing its translation, and Slit2 rapidly relieves the repression without causing miR-182 degradation. Our data support a model whereby miR-182 reversibly gates the selection of transcripts for fast translation depending on the extrinsic cue.

Axonal precursor mi RNA s hitchhike on endosomes and locally regulate the development of neural circuits

“…Thermal denaturation assay (Fig E) revealed increased fluorescence levels at low temperatures in the presence of in vitro transcribed pre‐miR‐181a‐1 (Fig F) but not in the presence of (i) pre‐miR‐187 which is absent from RGCs (this study and Ref. (Bellon et al , )), (ii) a modified pre‐miR‐181a‐1 lacking the loop region, and (iii) short double‐stranded miR‐181 mimic (see Methods; Appendix Fig S1D–F). This confirms that our MB is able to specifically hybridize with and detect pre‐miR‐181a‐1 in vitro .…”

“…LCM, laser capture microdissection. B, CRT–PCR performed on RNA extracted from axons collected by LCM (Ax) or from eyes. (B) β‐Actin mRNA is present both in eyes and in axons, while MAP2 and H4 are present only in the eye sample, suggesting the absence of contamination from cell bodies or other cells in LCM axonal samples (Bellon et al , ). Ax, axonal sample; Eye, stage 37/38 eye; LCM, laser capture microdissection; –, PCR no template control; NT, RT no template control. DSchematic of MB, pre‐miR‐181a‐1, and their hybridization complex.…”

“…To address this, we used isolated axons, which yield pure axons and growth cones responsive to cues (Appendix Fig S4A and B). We exposed isolated axons to the well‐described repellent cues, Sema3A and Slit2, at concentrations that generate protein synthesis‐dependent growth cone responses (Bellon et al , ) (Appendix Fig S4C). These cues are expressed within the optic tectum, a midbrain structure targeted by Xenopus RGC axons, and act as chemorepellent for these axons (Campbell et al , ; Piper et al , ).…”

“…To investigate pre-miRNA transport mechanisms, we first established that these precursors are indeed located to axons. Analysis of our previously published dataset of small RNAs (Bellon et al, 2017) revealed the existence of sequences corresponding to the loop region of 42 pre-miRNAs in axons and growth cones of Xenopus laevis retinal ganglion cells (RGCs), our model system (Appendix Fig S1A). We validated, by RT-PCR, the presence of the abundant pre-miR-182, pre-miR-181a-1, and pre-miR-181a-2 in eyes, and pure axonal RNA that we collected by laser capture microdissection (LCM; Fig 1A-C and Appendix Fig S1A).…”

MicroRNA-182 Regulates Neurite Outgrowth Involving the PTEN/AKT Pathway

“…MiR-182 is highly expressed in the retina (Xu et al, 2007) and identified as the most abundant among axonal miRNAs of RGCs. Loss of miR-182 causes RGC axon targeting defects in vivo (Bellon et al, 2017). In a companion paper, we compared the expression levels of several microRNAs between NSCs and neurons, and found that miR-182 was enriched in neurons.…”