Fig. 5: SOX2 enhances histone acetylation in part by inhibiting long chain fatty acid synthesis via repression of ACSL5 expression.

a Heatmap analyses showing SOX2 regulates the expression of various lipid metabolism-related enzymes in K450 cells, signals were z-scaled and DESeq2 normalized. b RT-PCR analysis confirming the regulation of various fatty acid metabolism-related genes by SOX2 in K450 cells. c RT-PCR analysis confirming that SOX2 knockdown led to significant upregulation of ACSL5 and ACSL6 in K450 cells. d RT-PCR analysis showing that ectopic SOX2 overexpression led to significant downregulation of ACSL5 and ACSL6 in K450 cells. e, f Oil red staining showing elevated lipid droplet level in SOX2 knockdown K30 and K140 cells (e) and in SOX2 knockout K450 cells (f). g Oil red staining showing substantial reduction of lipid droplet level in SOX2 overexpressing K30 cells. h Oil red staining comparing the levels of lipid droplets in SOX2-positive K70 and K450 cells and SOX2-negative K150 and K410 cells. i Western blot analysis showing that SOX2 also negatively controlled ACSL5 protein level but had no effect on ACC1 and FASN. Antibodies against SOX2, ACTIN, ACSL5, ACC1 were probed on the same gel. The samples originate from the same experiment, FASN were processed on separate gels in parallel. j Oil red staining showing accumulation of lipid droplets in K450 cells with ectopic overexpression of ACSL5 and marked reduction of lipid droplet level in K410 cells with knockdown of ACSL5. k Western blot analysis showing upregulation of histone acetylation level in K450 and K410 cells with ACSL5 knockdown. Note that ACSL5 knockdown upregulated SOX2 protein level in SOX2 positive K450 cells. Antibodies against SOX2, ACTIN, ACSL5, H3 were probed on the same gel. The samples originate from the same experiment, H3ac and Kac were processed on separate gels in parallel. l Western blot analysis showing that overexpression of ACSL5 downregulated histone acetylation levels in both SOX2-positive K30 and K450 cells and SOX2-negative K150 and K410 cells. Note that overexpression of ACSL5 slightly downregulated SOX2 protein level in SOX2 positive K30 and K450 cells. m Western blot analysis showing that overexpression of ACSL5 downregulated global non-histone acetylation level in cells SOX2-positive K450 cells and SOX2-negative K410 cells. In k, l antibodies against SOX2, ACTIN, ACSL5, H3 were probed on the same gel. The samples originate from the same experiment, H3ac, H4ac and AcK were processed on separate gels in parallel. n RT-PCR analysis showing that overexpression of ACSL5 downregulated the expression of multiple SOX2 target genes. RT-PCR data (b, c, d, n) are represented as mean +/- SD from 3 biological replicates, n = 3. Western blot data (l, m) were repeated independently at least 3 times with similar results. Quantifications of Oil red staining are shown in (e, f, g, h, j), 3 independent experiments. Scale bar, 20 μm. Data represented as mean ± SD; ns, P  >  0.05, *P  <  0.05, **P  <  0.01, ***P  <  0.001. Student’s t test (unpaired, two-tailed). Source data are provided as a Source Data file.