Fig 2: HDAC11 was involved in biological function of ST8SIA6-AS1 in HCC. a pcDNA3-HDAC11 increased HDAC11 protein expression in Huh7 cells. b, c Overexpression of HDAC11 slightly increased cell proliferation and attenuated the anti-proliferation effect of ST8SIA6-AS1 siRNA in Huh7 (b) and Hep3B (c) cells. d, e Overexpression of HDAC11 did not affect cell apoptosis but attenuated the pro-apoptotic effect of ST8SIA6-AS1 siRNA in Huh7 (d) and Hep3B (e) cells. ***vs. control siRNA + pcDNA3, p < 0.001; ## vs. ST8SIA6-AS1 siRNA + pcDNA3, p < 0.01; ### vs. ST8SIA6-AS1 siRNA + pcDNA3, p < 0.001; && vs. control siRNA + pcDNA3, p < 0.01; &&& vs. control siRNA + pcDNA3, p < 0.001

Fig 3: HDAC2 inhibition induced BRN3A expression in melanocytes and melanoma cells. (A) Basal protein expression of HDAC1, HDAC2, HDAC3, and HDAC11, in HM (3 donors), Hermes, and melanoma cell lines (1205Lu, WM165, WM1232, and WM35). (B) Protein expression of BRN3A and HDAC2 48 h after siRNA-mediated transient knockdown of HDAC2 in melanocytes (HM, Hermes) and melanoma cells (1205Lu). HDAC2 #1: HDAC2 siRNA. BRN3A induction upon HDAC2 inhibition was also confirmed by a second siRNA (HDAC2 #2, see Supplementary Figure S3A); control: non-targeting control siRNA; Ø: transfection reagent only (HM, Hermes: JetPrime; 1205Lu: Lipofectamine RNAiMAX). (C) Protein expression of BRN3A as well as HDAC1, HDAC3, and HDAC11 48 h after knockdown of HDAC1, 3, and 11, respectively, in melanocytes (HM, Hermes) and melanoma cells (1205Lu). HDAC1, HDAC3, and HDAC11: specific siRNAs targeting HDAC1, 3, and 11, respectively; control: non-targeting control siRNA; Ø: transfection reagent only (HM, Hermes: JetPrime; 1205Lu: Lipofectamine RNAiMAX). Representative immunoblots are shown.

Fig 4: HDAC11 affects mitophagy and intracellular mtROS clearance via mTOR. BMDMs were infected with S. aureus (MOI = 10) for 6 and 24 h, in conditions with or without overexpression of HDAC11 and treatment with Rapamycin (100nM). (A and B) JC-1 staining and flow cytometry analysis were used to assess changes in mitochondrial membrane potential. Quantitative analysis of the percentage of JC-1 monomer-positive cells. (C and D) Flow cytometry analysis of mtROS levels in BMDMs infected with S. aureus (MOI = 10) at various time points. Quantitative analysis based on Mean Fluorescence Intensity (MFI) indicating intracellular mtROS levels. (E) Confocal images show mitochondria (Mito-tracker, red), lysosomes (Lyso-tracker, green), and nuclei (Hochst, blue) within BMDMs (Scale bar: 10 μm). (F, G, H, and I) Western blot analysis of LC3, P62, mTOR, and p-mTOR protein expression levels in each group. GAPDH used as a reference protein for quantitative analysis of P62 expression levels and changes in LC3ii/LC3i, p-mTOR/mTOR ratios. (J, K, and L) Mitochondrial preparations were analyzed using western blotting to assess Pink-1 and Parkin protein expression levels in each group. COX IV was used as a reference protein to quantify the relative expression levels of Pink-1 and Parkin. Statistical analysis was performed using a two-tailed Student’s unpaired t-test: ns, P > 0.05; * P < 0.05; ** P < 0.01; *** P < 0.001. The data represent the mean ± SD from at least three independent experiments

Fig 5: HDAC11 expression affects intracellular survival of S. aureus via IL10/mTOR. (A, B, and C) Detection of HDAC11 and H3K9ac protein levels in BMDMs overexpressing HDAC11 using Western blot. GAPDH is used as a reference protein to quantify HDAC11 expression, and H3 to quantify H3K9ac expression. (D) Validation of IL10 production levels induced by 24-hour intracellular S. aureus infection in BMDMs overexpressing HDAC11, using ELISA. (E and F) Bacterial plating and (G and H) flow cytometry to assess the impact of the mTOR inhibitor Rapamycin on intracellular bacterial load in BMDMs overexpressing HDAC11, with or without inhibitor treatment. Quantitative analysis of intracellular bacterial counts at 6 h and 24 h, and the proportion of BMDMs infected with GFP-S. aureus. Statistical analysis was performed using a two-tailed Student’s unpaired t-test: ns, P > 0.05; * P < 0.05; ** P < 0.01; *** P < 0.001. The data are presented as the mean ± SD from at least three independent experiments