Fig 1: 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.
Fig 2: QCSL suppresses tumor growth in the BC xenograft mouse model. (a) Tumor volumes of vehicle, 200 mg/kg, and 400 mg/kg QCSL groups. ***P < 0.001 vs. control. (b) Image of tumors of each group. (c) TUNEL assay performed using the tumor samples of each group. (d) The protein levels of GLS1, SLC1A5, and c-Myc measured using the tumor samples of each group. ***P < 0.001 vs. vehicle.
Fig 3: ASCT2 is a direct target of miR-125b-5p. (A) The potential binding sites of miR-125b-5p and ASCT2 were predicted by TargetScan. (B) A luciferase reporter assay was performed in CCC-HIE-2 cells to confirm the interaction between miR-125b-5p and ASCT2. (C) Effects of miR-125b-5p mimic or inhibitor on ASCT2 mRNA expression levels. (D) Representative blot images and (E) quantification of ASCT2 protein expression levels following transfection with miR-125b-5p mimic or inhibitor. Data are expressed as the mean ± SD (n=5). ###P<0.001 compared with miR-NC; *P<0.05 and ***P<0.001 vs. LPS-alone. ASCT2, alanine serine cysteine-preferring transporter 2; miR, microRNA; NC, negative control; UTR, untranslated region; WT, wild-type; MUT, mutant; LPS, lipopolysaccharide.
Fig 4: hsa_miR-370-3p negatively regulated SLC1A5. A–K SKOV3 and A2780 cells were transfected with miR-NC, hsa_miR-370-3p, hsa_miR-370-3p + pcDNA, or hsa_miR-370-3p + SLC1A5. A SLC1A5 protein expression was examined using western blot. B and C CCK-8 and EdU assays assessed proliferation ability. D Apoptosis, invasion, and angiogenesis capacity was measured using flow cytometry, transwell, and tube formation assays. G and H c-Myc and MMP9 protein levels were analyzed via western blot. I–K Glutamine metabolism was evaluated via the corresponding assay kits. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001
Fig 5: SLC1A5 possesses a great prognostic value. (A) OS difference between high- and low-SLC1A5-expression groups. (B) PFS difference. (C) ROCs. (D) Time-independent ROCs. (E) The relationship between SLC1A5 expression and clinicopathological features of PAAD. (F,G) Independent prognostic analyses based on the multivariate and univariate cox regression methods. (H) DCA results. “Model A” represents the traditional prognostic model consisting of age, gender, histological grade, and clinical stage (blue line). “Improved model A” represents the prognostic model consisting of age, gender, histological grade, clinical stage, and SLC7A11 expression (red line). (I–S) Clinical subgroup analyses. OS, overall survival; PFS, progression-free survival; ROC, receiver operating characteristic curve; PAAD, pancreatic adenocarcinoma; DCA, decision curve analysis.
Supplier Page from Abcam for Anti-SLC1A5/ASCT2 antibody [CAL33]