Fig. 6. TINCR undermine the stability of OAS1 through binding to STAU1.

A Mean centered, hierarchical clustering of transcripts altered in breast cancer cells treated with scrambled siRNA or si-TINCR 2#, with three repeats. B, C qRT-PCR analysis was used to validate the changes of several top regulated mRNAs upon TINCR depletion and overexpression in T47D and MCF-7 cells. D OAS1 mRNA expression upon STAU1 depletion, as detected by quantitative reverse transcription (qRT–PCR). E OAS1 mRNA expression was detected by qRT–PCR in pcDNA-TINCR and si-STAU1 co-transfected T47D and MCF-7 cells. F Protein level of OAS1 was detected by western blot assays in pcDNA-TINCR and si-STAU1 co-transfected T47D and MCF-7 cells. G Bioinformatics tool (http://bioinfo.bjmu.edu.cn/lncpro/) was used to evaluate the combination possibility between several top upregulated mRNAs and STAU1. Predictions with probabilities >0.5 were considered “positive”, indicating that the corresponding RNA and protein are likely to interact. OAS1 showed high possibility to interact with STAU1. H RIP assay was performed with IgG or STAU1 antibodies in T47D cells, and the coprecipitated RNA was subjected to qRT-PCR for OAS1. I The predicted TINCR binding site in OAS1 (OAS1-3′UTR-Wt) and the designed mutant sequence (YY2-30 UTR-Mut) are shown (up). Luciferase reporter gene assays of T47D and MCF-7 cells are shown (down) (OAS1-3′UTR-Wt + TINCR-NC group is seen as control). J, K TINCR and STAU1 control OAS1 mRNA stability. RNA stability assays were performed in T47D cells using Actinomycin D to disrupt RNA synthesis, and the degradation rates of the OAS1 mRNA were measured over 9 h. Values are shown as the mean ± s.d in three independent experiments. The mean values of two groups were compared via paired, two-tailed Student t tests.*P < 0.05; **P < 0.01.