Abstract
Purpose :
Histone deacetylases (HDACs) 1 and 2 are highly conserved nuclear enzymes that are the core components of the corepressor complexes. HDAC1 and 2 regulate the chromatin structure and function and play a key role in critical biological processes like aging, oxidative stress, autophagy, and proteinopathy. Although these cellular processes are risk factors for several age-related eye diseases, the role of HDAC 1 and 2 are not well defined in the context of the eye and eye-related disorders. The current study explores a previously unrecognized role for HDAC1 and HDAC2 in regulating chromatin modifying enzymes in mouse RPE.
Methods :
We used CRISPR/Cas9 genome editing system to generate single or combinatorial depletion of HDAC 1 and 2 in ARPE19 cells. Also, in vivo effects of HDAC 1 and 2 on chromatin modifying enzymes were determined using a transgenic mouse that expresses Cre recombinase in the RPE with floxed HDAC 1 and 2. RNA was isolated from both the ARPE19 and mouse RPE cells (at 4 and 8 weeks). qPCR was conducted to transcriptionally profile a panel of 84 epigenetic chromatin modifying genes that included DNA and histone methyltransferases, DNA and histone demethylases, histone acetyltransferases, deacetylases, phosphorylases, and set domain proteins.
Results :
Ablation of HDAC 1 or 2 leads to compensatory upregulation of other HDACs. Simultaneous deletion of HDAC 1 and 2 leads to repression of several chromatin modifying enzymes. The mouse RPE showed differential expression of approximately 16 % genes at 4-week, and this drastically increases to 50 % genes at 8-week. This is concordant with the qPCR data as 8-week old mouse RPE had lower levels of HDAC1 and 2 transcripts than 4-week old RPE. Interestingly, except class II MHC transactivator (Ciita) gene, all the other genes were downregulated. Some of the genes which display altered expression as early as 4 weeks and continue the trend in 8-week mouse RPE include Atf2, Kat2b, Ncoa1, Ube2b, Usp21, and Hsp90ab1. Interestingly, HDAC 1 and 2 depleted RPE cells show downregulation of many HDACs with the maximum depletion of HDAC11 amounting to nearly 50 % of control. Thus the data shows that combined depletion of HDAC1 and HDAC2 can alter the transcript levels of many chromatin modifying enzymes.
Conclusions :
In conclusion, these data show that HDAC1 and HDAC2 play an essential role in the transcriptional regulation of chromatin modifiers.
This is a 2021 ARVO Annual Meeting abstract.