Fig 2: HDAC1/2 ablation causes major changes in transcriptional regulation in the retina. (a) Volcano plots illustrating global transcriptional changes in intact and injured (2 dpi) retinae of HDAC1/2 mutants and control. Each dot represents one gene. The log2 fold change is represented on the x axis, and the y axis shows the log10 of the p value. All genes with a p value lower or equal to .01 and a fold change greater than or equal to1.5 are represented in red. (b) Venn diagram showing the total upregulated and downregulated genes in the control and HDAC1/2 mutant retinae, with or without injury (p < .05). (c) Heat map depicting changes in expression of the most significantly regulated genes (p < .001, fold change ≥ 1.5) in control retinae following optic nerve transection and compared with expression changes between HDAC1/2 mutant intact and injured retinae. (d) The JNK-p53 stress-activated pathway was identified as a major interaction network involved after optic nerve transection.

Fig 3: MAP kinase pathways are activated comparably in HDAC1/2 mutants and controls following injury. (a) RNA-seq analysis of the JNK targets Jun, p53, and Hrk. Error bars indicate SEM (n = 3, *p < .05, **p < .01, ***p < .001). (b) Retinae lysates of control and HDAC1/2 mutants, with or without injury, were analyzed by immunoblotting at 2 dpi and 4 dpi to assess activation of MAP kinase pathways (JNK, c-jun, and Erk1/2). Alpha-tubulin was used as control. Error bars indicate SEM (n = 3, *p < .05, **p < .01, ***p < .001). Int = intact, not injured. Ctl = control.

Fig 4: Ablation of HDAC1/2 reduces p53-dependent transcription of PUMA and p21 following injury. (a) RNA-seq analysis of p53 targets in HDAC1/2-ablated retinae. Error bars indicate SEM (n = 3, **p < .01, ***p < .001). (b) Confirmation of RNA-seq analysis by qRT-PCR, with extension to 4 dpi. Note: Fas mRNA upregulation at 4 dpi but not at 2 dpi. Error bars indicate SEM (n = 3, *p < .05, **p < .01). (c) Western blot analysis of main p53 targets revealed patterns comparable to those observed at the mRNA level. Error bars indicate SEM (n = 3, *p < .05, **p < .01). (d) Retinal immunohistochemistry showing increased expression of the proapoptotic protein PUMA in retinal ganglion cells following injury (4 dpi) in the control and reduced signal in HDAC1/2 mutant. Scale bar: 250 µm.