Fig. 8
L-lactate-activated Hif-1α promotes DCBLD1 transcription. A DCBLD1 mRNA levels in SiHa cells, treated with the addition of sodium L-lactate (25 mM) for varying durations, were determined using qPCR. B, C Immunoblotting was performed to determine the Hif-1α level after a 6 h treatment with sodium L-lactate (25 mM). GAPDH was used as a loading control. C Immunoblotting was performed to determine Hif-1α, G6PD, and DCBLD1 levels in SiHa cells transfected with DCBLD1 shRNA vector, both with or without treatment with sodium L-lactate (25 mM). D, E DCBLD1 mRNA levels were determined using qPCR. F JASPAR was used to predict Hif-1α binding sites to the DCBLD1 promoter. G Chip assay for Hif1α binding to the DCBLD1 promoter, with/without sodium L-lactate (25 mM) treatment. H Construction of plasmids for wild-type and mutant (at the 1147–1156) luciferase reporter genes region of the DCBLD1 promoter. I A luciferase reporter gene assay was performed to validate the Hif1α binding site on the DCBLD1 promoter. Data are presented as the mean ± SD. Statistical significance was assessed using a one-way ANOVA with the Brown-Forsythe test (A), an unpaired t test (D and E), or a two-way ANOVA with Šídák's multiple comparisons test (I). (****, p < 0.0001; ***, p < 0.001; **, p < 0.01; *, p < 0.05)