Fig 2: SRSF1 binding renders the LPS-sensitivity of the target mRNAs for their nuclear export.(A) REMSA was performed to show binding of purified recombinant SRSF1 protein to the Cxcl1 3’UTR (nt 781–901), Cxcl1 3’UTR (nt Δ 790–840 in 781–901) and Cxcl1 3’UTR (800-855). (B–D) CMV-Luc- Cxcl1 3’UTR (721-940), (nt Δ 790–840 in 721–940) and (nt Δ 829–835 in 721–940) plasmids were transiently transfected in IRAK-deficient 293-IL1R (I1A) cells for 24 hr; followed by RNA immunoprecipitation with anti-SRSF1 antibody and RT-PCR analyses of the indicated mRNAs, the presented are the relative values normalized against IgG control (Material and methods) (B); Luciferase activity was measured (C); RT-PCR analysis of total mRNAs isolated from the cytoplasmic and nuclear fractions (D). (E) CMV-Luc- Cxcl1 3’UTR (721-940), (nt Δ 790–840 in 721–940) and (nt Δ 829–835 in 721–940) plasmid were transiently transfected in wild-type, IRAK2 KO and IRAK2 KI macrophages and treated with LPS for 12 hr, followed by luciferase assay. (F) For the transfected cells as described in Figure 6B–D, cell lysates were subjected to RNA immunoprecipitation with anti-ALYREF antibody and RT-PCR analyses of the indicated mRNAs. The presented are the relative values normalized against IgG control (Materials and methods). (G) REMSA was performed to show binding of purified recombinant ALYREF protein to the Cxcl1 3’UTR (nt 781–901), Cxcl1 3’UTR (nt Δ 790–840 in 781–901) and Cxcl1 3’UTR (800-855). Data represent mean ± SEM; *p<0.05 by Student’s t test.10.7554/eLife.29630.034Figure 6—source data 1.The numerical data for the graphs in Figure 6.(A) for Figure 6B. (B) for Figure 6C. (C) for Figure 6D. (D) for Figure 6E. (E) for Figure 6F.

Fig 3: LncRNA TUG1 suppressed H/R-induced ferroptosis in HK-2 cells by regulating SRSF1/ACSL4. A Cell viability was assessed by CCK-8 kits. B Cell death of HK-2 cells was measured by TUNEL assay. TUNEL-positive cell quantitative bar diagrams are shown. Scale bar: 50 µm. C The levels of MDA and GSH in the supernatants of HK-2 cells were detected by commercial kits. D The levels of Fe2+ in HK-2 cells were checked by commercial kits. E The levels of ROS in HK-2 cells were detected using DCFH-DA probe. F The protein levels of ACSL4, GPX4, FTH1, and COX2 in HK-2 cells were assessed by western blot and shown as blot image and quantitative bar diagram. N = 3 samples per groups. *p < 0.05, **p < 0.01, ***p < 0.001. The above data were all measurement data and expressed as means ± SD. All experiment was performed three times

Fig 4: USC-Exo ingested by kidney cells upregulate lncRNA TUG1 expression, which promotes ACSL4 mRNA decay by interacting with SRSF1, then suppresses ACSL4-mediated cell ferroptosis, and furtherly improves IRI-induced AKI

Fig 5: SRSF1 promotes bladder cancer tumorigenesis in part via regulating CD46-CYT2 levels(A) EJ-1 cells were infected with lentivirus expressing several indicated shRNAs to establish stably expressing cell lines. Semiquantitative RT-PCR analysis was performed to detect the CD46 exon 13 alternative splicing. The ratio for 13−/13+ is listed below the panel. (B) CD46 exon 13 splicing was measured by RT-PCR in EJ-1 cells stably expressing FLAG-Cherry, FLAG-SRSF1, or FLAG-hnRNPA1. (C) Relative expression of SRSF1 mRNA expression levels were evaluated by real-time PCR in 27 paired case specimens. Expression levels of SRSF1 were normalized to that of GAPDH. A bar value <1 indicates that SRSF1 is decreased in tumors. A bar value >1 indicates that SRSF10 is increased in tumors. (D) The positive correlation between the CD46 13−/13+ ratio and expression levels of SRSF1 was observed in bladder cancer samples. Relationships between these two variables were determined by Pearson’s correlation coefficients. The correlation was analyzed using GraphPad Prism 5 software. (E) A CCK-8 assay was utilized to quantify cell viability at each time point. Data are plotted as the mean ± SD of three independent experiments and were analyzed by two-way ANOVA. ∗∗∗p < 0.001. (F) (1) Representative images of cell culture plates following staining for colony formation of the indicated cell lines. (2) The number of colonies was quantified. The data represent mean ± SD and were analyzed by an unpaired two-tailed Student’s t test (n = 3). ∗p < 0.05. n.s., not significant. (G) (1) Migration assay for the indicated cell lines. (2) Number of migrated cells was quantified in five random images from each treatment group. The data represent mean ± SD and were analyzed by an unpaired two-tailed Student’s t test (n = 5). ∗∗p < 0.01, ∗∗∗p < 0.001.