Article Title: HSF1 Stimulates Glutamine Transport by Super-Enhancer-Driven lncRNA LINC00857 in Colorectal Cancer
Figure Lengend Snippet: The LINC00857/ANXA11 axis promotes SLC1A5-mediated glutamine transport. ( A ) Score plot of principal components analysis (PCA) between control and LINC00857 knockdown group after LC–MS (liquid chromatography mass spectrometry) analysis. ( B ) Heatmap of 22 amino acids’ alteration after LINC00857 knockdown. ( C ) Association analysis between different amino acid. ( D ) Concentration of Glu (glutamic acid) and Asp (aspartic acid) after silencing LINC00857. ( E ) Correlation between ANXA11 and SLC1A5 expression using the GEPIA database. ( F ) The effect of ANXA11 knockdown on SLC1A5 expression in CRC cells, as explored by Western blotting. ( G , H ) Bioinformatics tools predicted a series of miRNAs complementary to ANXA11 mRNA and the SLC1A5 3′UTR region. ( I ) Dual-luciferase assays of the ANXA11 mRNA/SLC1A5 3′UTR constructions with intact or mutated seed sequences for miR-122-5p. ( J ) The effect of miR-122-5p mimics on SLC1A5 protein level, as explored by western blotting. ( K ) The effect of miR-122-5p inhibitors on ANXA11 knockdown-induced SLC1A5 downregulation, as explored by Western blotting. ( L , M ) RIP (RNA immunoprecipitation) assays were performed using the Ago2 antibody. The relative expression levels of ANXA11 and miR-122-5p were determined by qPCR. All data represent the mean ± SD, n = 3. * p < 0.05; ** p < 0.01; **** p < 0.0001.
Article Snippet: The following antibodies were used in this study: a-Tubulin (T5168, Sigma-Aldrich, St. Louis, MO, USA), HSF1 (ab52757, Abcam, Cambridge, UK), Histone H3 (acetyl K27) (ab177178, Abcam), ANXA11 (SC46686, Santa Cruz, CA, USA), p-mTOR (ab109268, Abcam), p-P70S6K (9234S, CST, Danvers, MA, USA), SLC1A5 (ab237704, Abcam), P300 (ab275378, Abcam), Pol II (ab26721, Abcam), BRD4 (A301-985A100, Bethyl Laboratories, Montgomery, TX, USA), β-actin (4970L, CST).
Techniques: Liquid Chromatography with Mass Spectroscopy, Liquid Chromatography, Mass Spectrometry, Concentration Assay, Expressing, Western Blot, Luciferase, Immunoprecipitation