Abstract
Purpose:
The epigenetic state of any cell is, in part, regulated by the interaction of DNA with nuclear histones. Histone tails can be modified in a number of ways that impact on the availability of DNA to interact with transcriptional complexes, including methylation, acetylation, phosphorylation, ubiquituination, and sumoylation. Histones are acetylated by a large family of enzymes, histone acetyl transferases (HATs), and deacetylated by the histone deacetylases. Acetylated histones are generally considered markers of genomics regions that are actively being transcribed, whereas deacetylated and methylated histones are markers of regions that are inactive.The goal of the present study was to determine the epigenetic state of retinal neurons in general and elucidate which of the classical HDACs might be playing a role in the differentiation of retinal ganglion cells.
Methods:
Antibodies against acetylated histones, methylated histones and HDACs1-11 were tested by western blot on extracts of developing retinal tissue in a previous study on chick and mouse optic nerve. Expression patterns in vivo were characterized by double-label immunofluorescence for HDACs and retinal ganglion cell-specific Brn 3b or RA4 on cryosections through embryonic day 3 (E3), E5, E8 and E18 in chick retina.Immunolabeled sections were visualized with an Olympus confocal microscope.
Results:
A basic pattern emerged in which acetylated and methylated histones and HDACs were found throughout the developing retina, in a gradient that was highest in the ganglion cell layer and lowest in the outer layers of the retina.This pattern was consistent from E3 to E8, but analysis at the mature stage of the chick retina (E18) indicated that, for the most part, HDAC label was weak or absent in the outer nuclear layer and outer portion of the inner nuclear layer.Further, HDACs 1-6, 9 and 11 labeled a subpopulation of cells in the inner half of the inner nuclear layer, whereas HDACs 8 and 10 were completely absent.
Conclusions:
Our work suggests that the models in which neuronal cells are globally acetylated during differentiation are incorrect and a model in which both acetylation and deacetylation is critical for the proper differentiation of the neuronal populations of the retina. Furthermore, both acetylation and deacetylation appears to be necessary for the maintenance and/or survival of retinal ganglion cells in the mature retina.
Keywords: 698 retinal development