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. 2020 May 29;11:2677. doi: 10.1038/s41467-020-16504-1

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© The Author(s) 2020

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. 4 — a Mechanism of hypoxic induction for RBP-dependent translatomes. Global analysis of hypoxia-induced translational versus RNA-level changes in U87MG using RNA sequencing of ribosome density fractions for HuR-, hnRNP A2/B1-, and PCBP1-dependent targets. Color scheme: red, Class I proteins, normoxia-enriched; blue, Class III proteins, hypoxia-enriched. b Global analysis of HuR-dependent changes in mRNA translation efficiency and steady-state expression in U87MG using RNA sequencing of ribosome density fractions following siRNA-mediated HuR-silencing. c Hypoxic translation efficiency regulation of glycolytic effectors by the hypoxia-adaptive RBPs PCBP1, hnRNP A2/B1, and HuR, as determined by qRT-PCR of ribosome density fractions. Asterisk denotes statistical significance calculated using two-sided Student’s t-tests compared to non-silencing control. Exact p values: PCBP1 siRNA: GLUT1 (p = 0.02), GPI (p = 0.03), ALDOA (p = 0.001), ALDOC (p = 0.02), GAPDH (p = 0.007), PGK (p = 0.03), PGAM1 (p = 0.004), ENO1 (p = 0.047); hnRNP A2/B1 siRNA: GLUT1 (p = 0.007), HK2 (p = 0.004), PFKP (p = 0.04), ALDOA (p = 0.04), ALDOC (p = 0.048), GAPDH (p = 0.05), PGK (p = 0.03), ENO2 (p = 6.79e−05); HuR siRNA: GLUT1 (p = 0.04), HK2 (p = 0.04), ALDOC (p = 0.001), PKM (p = 0.0008). Data represent mean ± SEM (error bars) of three independent experiments (n = 3). Red box: observed decrease in protein output as determined by TMT-pSILAC. d Representative immunoblots of normoxic and hypoxic (6 h) U87MG treated with actinomycin D (+) or vehicle DMSO (−). NS: non-silencing. Quantitation represents mean of three independent experiments (n = 3). Source data are provided as a Source data file.