Fig 2: SPHK2 inhibits acetylation of IFN-β promoter through TET3 and HDAC1.(A) A549 cells and sgSPHK2-1 A549 cells were infected with IAV (WSN), MOI = 0.5, 24 h post infection, IFN-β promoter enrichment for H3ac, H4ac and HDAC1 antibodies was detected by ChIP-qPCR. (B) A549 cells transfected with pCMV-Myc or Myc-SPHK2 were infected with IAV (WSN), MOI = 0.5, 24 h post infection, IFN-β promoter enrichment for H3ac, H4ac and HDAC1 antibodies was detected by ChIP-qPCR. (C) A549 cells were treated with solvent (-) or TSA (5, 10, 20 or 50 nM) for 72 h, cell viability was detected by Cell Counting Kit-8. (D) A549 cells were treated with TSA (5, 10 and 20 nM), then transfected with Myc-SPHK2 and infected with WSN, MOI = 0.5, after 24 h, IFN-β mRNA level was measured by qPCR, and normalized to GAPDH. (E) A549 cells were transfected with TET3-siRNA, then transfected with Myc-SPHK2 and infected with WSN, MOI = 0.5, after 24 h, cells IFN-β mRNA level was measured by qPCR. Data are representative of three independent experiments. ns, no significance; *, P<0.05; **, P<0.01; ***, P<0.001.

Fig 3: The correlation of HDAC1 and CREPT in human tumors. (A). CREPT expression is positively correlated to HDAC1 at the protein level in colon cancer cells. The endogenous protein levels of HDAC1 and CREPT in colon cancer cell lines (SW620, SW480, HCT116, DLD1) and epithelial cells (NCM460) are shown. β-actin was used as a loading control. (B,C). Both HDAC1 (B) and CREPT (C) are highly expressed in colon cancer cells when compared with the normal cells at the mRNA level. The mRNA levels were normalized to the fold change relative to β-actin. (D). Both HDAC1 and CREPT are elevated in the tumor tissues compared with the adjacent normal tissues from human colon cancers. Western blot analyses were based on β-actin as a loading control. N represents paired adjacent normal tissue and T represents tumor tissue from the same patient. The bands were quantified using ImageJ and presented as a value normalized according to the moderate level of a band in each blot. (E). The relative level of CREPT and HDAC1 in adjacent normal and tumor tissues are shown. The ratio of CREPT or HDAC1 vs actin was calculated by quantifying the bands from the Western blot in (D). The closed circle represents normal tissue for CREPT/actin, the square represents tumor tissue for CREPT/actin, the triangle represents normal tissue for HDAC1/actin and the inverted triangle represents tumor tissue for HDAC1/actin. (F). HDAC1 and CREPT were co-stained in human colon tumor tissue. Immunohistochemical staining was performed with an antibody against CREPT or HDAC1. (G). A graphical presentation of correlation of CREPT and HDAC1 in colon cancer. The levels of CREPT and HDAC1 in tumor tissue and the adjacent normal tissue were quantified using Image J. The ratio of CREPT (X-axis) or HDAC1 (Y-axis) level in tumor tissue to the adjacent normal tissue was calculated. SPSS was employed to figure out the correlation coefficient between the two proteins. (* p < 0.05, ** p < 0.01, *** p < 0.001).

Fig 4: Hub genes selection and identification. (A) Venn diagram, two comparison groups showed an overlap of 142 different expressed genes. (B) The protein-protein interaction (PPI) network of overlapped different expressed genes, HDAC1 and Nrf2 were considered the key genes due to the most nodes. (C) HDAC1 mRNA expression levels in kidney tissues by quantitative real-time PCR (qRT-PCR) (n = 6). (D) Protein expression of HDAC1 and H3K27AC in kidney tissues (n = 3). Values are the mean ± SD that are significantly different indicated by asterisks as follows: **P < 0.01. Con, control; Col, colistin.

Fig 5: Activation of the ClpP protease along with inhibition of HDAC1/2 regulates the expression of antiapoptotic Bcl-2 family members. A and B, Standard Western blots of the antiapoptotic Bcl-2 family members (Mcl-1, Bcl-xL) in U251, GBM14, GBM12, and NCH644 cells treated with 10 µmol/L ONC206/ONC12 and 0.2 µmol/L panobinostat/2.5 nmol/L romidepsin for 24 hours. C, NCH644 and GBM14 cells were transduced with empty vector (EV) or Mcl1 adenovirus for 24 hours, treated with 10 µmol/L ONC206 and 0.2 µmol/L panobinostat/2.5 nmol/L romidepsin for 48 hours, and were labeled with Annexin/PI dye for flow cytometry analysis. Shown is the quantification of apoptotic cells (n = 3). D, NCH644 and GBM14 cells were transduced with empty vector (EV) or Bcl-xL adenovirus for 24 hours, treated with 10 µmol/L ONC206 and 0.2 µmol/L panobinostat/2.5 nmol/L romidepsin for 48 hours, and were labeled with Annexin/PI dye for flow cytometry analysis. Shown is the quantification of apoptotic cells (n = 3). Statistical significance was assessed by two-tailed Student t test. ****, P < 0.001.