Fig. 4: LHX1-DT acts as a ceRNA to absorb miR-590-5p.

A LHX1-DT was predicted to be located mainly in the cytoplasm using the bioinformatics tools in lncATLAS. B The qRT-PCR analysis of subcellular LHX1-DT expression in the nucleus and cytoplasm of RCC cells. C Subcellular localization of LHX1-DT in RCC cells detected by RNA-FISH. LHX1-DT was stained red (Cy3), and nuclei were stained blue (DAPI). D Schematic diagram of the Ago2-RIP process. E Fold enrichment of LHX1-DT in 786-O and Caki-1 cells. F Venn diagram showing the predicted target of LHX1-DT. G Relative expression of miR-590-5p in 52 paired RCC tissues compared with adjacent normal renal tissues by qRT-PCR. H RIP assay was performed using AGO2 antibody in 786-O and Caki-1 cells, then the enrichment of miR-590-5p was detected. I RIP assay was performed using AGO2 antibody in 786-O and Caki-1 cells transfected with miR-590-5p mimics or mimics NC, then the enrichment of LHX1-DT was detected. J The correlation between miR-590-5p and LHX1-DT was evaluated, and regression analysis was applied (r = −0.2984, P = 0.0317). K Expression level of miR-590-5p in 786-O and Caki-1 cells after transfection with overexpression plasmids of LHX1-DT. L Expression level of miR-590-5p in 786-O and Caki-1 cells treated with siRNAs of LHX1-DT. M The binding sites of LHX1-DT and miR-590-5p. N, O Luciferase reporter activity of wild-type (WT) or mutated (MUT) LHX1-DT co-transfected with miR-590-5p mimics in 786-O (N) and Caki-1 (O) cells. P LHX1-DT was pulled down with biotinylated miR-590-5p, and miR-590-5p was also pulled down with biotinylated LHX1-DT in 786-O and Caki-1 cells. Q, R The addition of functional FOXQ1-cDNA partially rescued the growth (Q) and invasion (R) of 786-O and Caki-1 cells transfected with miR-590-5p mimics. S, T The addition of LHX1-DT-shRNA partially rescued the growth (S) and invasion (T) of 786-O and Caki-1 cells transfected with an miR-590-5p inhibitor.