Journal: Cancer Communications
Article Title: TIMELESS Promotes LUAD Growth via Suppressing Transferrin-Mediated Ferroptosis and Reprograms the Tumor Microenvironment against Anti-PD-1 Immunotherapy
doi: 10.34133/cancomm.0009
Figure Lengend Snippet: Dysregulated RNA-binding proteins in LUAD: from multicohort screening to functional validation. (A to D) Volcano plots showing the DE RBPs in the following 4 LUAD cohorts: The Cancer Genome Atlas Lung Adenocarcinoma (TCGA-LUAD) dataset ( n = 568; A), GSE32863 ( n = 116; B), GSE40419 ( n = 164; C), and GSE75037 ( n = 166; D). The significantly DE RBPs ( P adj < 0.05, |log 2 FC| > 1) are shown in blue, and the nonsignificantly DE RBPs ( P adj > 0.05, |log 2 FC| > 1) are shown in dark gray. The other non-RBP genes are shown in light gray. (E and F) Venn diagram showing the intersection of up-regulated (E) and down-regulated (F) DE RBPs across TCGA-LUAD and the 3 GEO datasets. (G) The volcano plot displayed the hazard ratios for patients stratified by the median expression level of RBP genes in the TCGA-LUAD cohort (red: up-regulated RBPs in tumors; blue: down-regulated RBPs in tumors). (H) Heatmap of significantly dysregulated RBPs identified in (E) and (F). The top 10 highest-expressed RBPs in tumors are highlighted (ranked by average tumor z score). (I) Expression levels of the top 10 LUAD-up-regulated RBPs in LUAD cell lines (A549 and H1975) with or without siRNA-mediated knockdown ( n = 4 each group). (J) Calcein-AM/PI staining showed the percentage of viable cells in the negative control group and following the knockdown of the RBP gene ( n = 4 each group). (K) CCK-8 assay of cell viability in RBP gene knockdown and control cells at the indicated time points. The statistical analysis was performed using a 2-tailed Student’s t test (I and J) or 2-way ANOVA (K). *P < 0.05, **P < 0.01, ***P < 0.001, data without statistically significant differences are not labeled (J and K). Abbreviations: ALDH18A1, aldehyde dehydrogenase 18 family member A1; BZW2, basic leucine zipper and W2 domains 2; CCK-8, cell counting kit-8; CENPF, centromere protein F; DE, differentially expressed; FAM83A, family with sequence similarity 83 member A; FC, fold change; FDR, false discovery rate; GEO, Gene Expression Omnibus; IF, immunofluorescence; IHC, immunohistochemistry; KIF20A, kinesin family member 20A; LUAD, lung adenocarcinoma; OS, overall survival; PDIA4, protein disulfide isomerase family A member 4; PRC1, protein regulator of cytokinesis 1; RBP, RNA-binding protein; SRPK1, SRSF protein kinase 1; TCGA, The Cancer Genome Atlas; TMA, tissue microarray; TOP2A, DNA topoisomerase II alpha.
Article Snippet: Cell counting kit-8 (CCK-8) reagent (MedChemExpress, Cat. HY-K0301) was added to each well, and the absorbance at 450 nm was measured using a microplate reader (TECAN, Infinite 200 PRO).
Techniques: RNA Binding Assay, Functional Assay, Biomarker Discovery, Expressing, Knockdown, Staining, Negative Control, CCK-8 Assay, Control, Labeling, Cell Counting, Sequencing, Gene Expression, Immunofluorescence, Immunohistochemistry, Microarray
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