CELF RNA binding proteins promote axon regeneration in C. elegans and mammals through alternative splicing of Syntaxins

Lizhen Chen,Andrew D Chisholm,Bing Zhou,Xiang-Dong Fu,Zhijie Liu,Yishi Jin,Yu Zhou,Michael G Rosenfeld,Chaoliang Wei

doi:10.7554/elife.16072

Lizhen Chen, Andrew D Chisholm + Show 7 more

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https://doi.org/10.7554/elife.16072

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Abstract

Axon injury triggers dramatic changes in gene expression. While transcriptional regulation of injury-induced gene expression is widely studied, less is known about the roles of RNA binding proteins (RBPs) in post-transcriptional regulation during axon regeneration. In C. elegans the CELF (CUGBP and Etr-3 Like Factor) family RBP UNC-75 is required for axon regeneration. Using crosslinking immunoprecipitation coupled with deep sequencing (CLIP-seq) we identify a set of genes involved in synaptic transmission as mRNA targets of UNC-75. In particular, we show that UNC-75 regulates alternative splicing of two mRNA isoforms of the SNARE Syntaxin/unc-64. In C. elegans mutants lacking unc-75 or its targets, regenerating axons form growth cones, yet are deficient in extension. Extending these findings to mammalian axon regeneration, we show that mouse Celf2 expression is upregulated after peripheral nerve injury and that Celf2 mutant mice are defective in axon regeneration. Further, mRNAs for several Syntaxins show CELF2 dependent regulation. Our data delineate a post-transcriptional regulatory pathway with a conserved role in regenerative axon extension.

Highlights

Axon regeneration requires coordinated gene expression at many levels (Benowitz et al, 1981; Gervasi et al, 2003; Glasgow et al, 1992; Skene and Willard, 1981)
Overexpression of UNC-64 in unc-75 null mutants can rescue axon regeneration defects, indicating that UNC-64 is a major target of UNC-75 in regenerating neurons. Extending these findings to mammals, we find that mouse Celf2 expression is induced by axon injury and that CELF2 is required for effective peripheral axon regeneration
Transgenic animals expressing Pmec-4-GFP::UNC-75DNLS(aa 1–472) showed diffuse fluorescence throughout the cell, as compared to full-length GFP::UNC-75, which localizes to neuronal nuclei (Figure 1—figure supplement 1B) (Loria et al, 2003), suggesting that nuclear localization may be critical for UNC-75 function in axon regeneration

Summary

Introduction

Axon regeneration requires coordinated gene expression at many levels (Benowitz et al, 1981; Gervasi et al, 2003; Glasgow et al, 1992; Skene and Willard, 1981). While much work has focused on injury-regulated gene transcription, increasing evidence points to roles for post-transcriptional regulation of mRNAs by RNA binding proteins (RBPs). The conserved RNA 3’-terminal phosphate cyclase has been identified as an inhibitor of axon regeneration in C. elegans, Drosophila and mouse, acting through RNA repair and splicing (Kosmaczewski et al, 2015; Song et al, 2015). Despite these advances, mechanistic understanding of the roles of RBPs in axon regeneration remains limited

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