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. Author manuscript; available in PMC: 2015 Jun 5.
Published in final edited form as: Cell. 2015 May 7;161(4):790–802. doi: 10.1016/j.cell.2015.02.053

Figure 5. YBX1 target transcripts and their response to changes in tRF levels.

Figure 5

(A) YBX1 interacts with the 3′UTRs of HMGA1, CD151, CD97, and TIMP3. The last exon of the indicated transcripts are shown with mapped reads from experimental replicates of YBX1 CLIP-seq. (B) Transfection of antisense LNAs against YBX1-binding tRFs resulted in the up-regulation of HMGA1, CD151, CD97, and TIMP3 transcripts, in a YBX1-dependent manner, as determined by qPCR measurements. (C) Similarly, transfecting antisense LNAs resulted in a significant stabilization of HMGA1, CD97, and TIMP3 transcripts in a YBX1-dependent manner. Whole-genome RNA stability measurements were perfomed using α-amanitin-mediated inhibition of RNA polymerase II (see Methods). (D) A GFP/mCherry dual-reporter assay was used to measure the effects of cloning HMGA1, CD97, and part of the TIMP3 3′ UTRs downstream of mCherry using qRT-PCR. The 3′ UTR of MAPK14, which is devoid of YBX1 tags, was included as a control. Consistent with our prior findings, LNA transfections resulted in a significant increase in relative mCherry expression in a YBX1-dependent manner. (E) Exogenously added tRF mimetics, while showing no effect on MAPK14 abundance, resulted in a significant depletion of HMGA1, CD97, and TIMP3 transcripts from the YBX1 co-immunoprecipitated RNA population. Statistical significance is measured using one-tailed Student’s t-test: *, p<0.05, **, p<0.01, and ***, p<0.001. Error bars in all panels indicate s.e.m. unless otherwise specified.