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. 2018 Jan 2;9:10. doi: 10.1038/s41467-017-02449-5

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 6 — The Poly(A) SPOT-ON reduces nascent protein synthesis and axonal translation in DRG neurons. a Cultured DRG neurons are incubated with SPOT-ONs (10 μM) or homoharrintonine (50 μM) for 3 h prior to addition of puromycin (1 μM) for an additional 15 min. Incubation with Poly(A) SPOT-ON, but not scrambled SPOT-ON or vehicle, significantly reduces nascent protein synthesis in DRG neurons. Staining is shown from top to bottom for puromycin (green), peripherin (red), or a merge. b Quantification of a. n = 6. *P < 0.05, **P < 0.01, significantly different from vehicle + puro group analyzed by one-way ANOVA followed by Bonferroni post hoc test. c Cultured DRG neurons are incubated with vehicle, SPOT-ONs, or hippuristanol for 3 h followed by emetine incubation (200 μM) for 5 min and puromycin (100 μM) for an additional 5 min. Incubation with Poly(A) SPOT-ON (10 μM), but not scrambled SPOT-ON or vehicle, significantly reduces proximal axonal translation (around 20–25 μM from the cell body) in peripherin-positive DRG axons. As in a, staining is shown from top to bottom for puromycin (green), peripherin (red), or a merge. d Representative images showing distal axonal ribopuromycylation (more than 25 μM from the cell body; randomly selected) in peripherin-positive DRG axons under identical conditions as described in c. e Quantification of images in c. n = 20. *P < 0.05, **P < 0.01, significantly different from vehicle+E+P group analyzed by one-way ANOVA followed by Bonferroni post hoc test. f Quantification of images in d. n = 9. *P < 0.05, **P < 0.01, significantly different from vehicle +E + P group analyzed by one-way ANOVA followed by Bonferroni post hoc test. For all graphs shown in the figure, data are plotted as mean ± s.e.m.