Abstract
Investigations into the epigenetic status of individual cells within tissues can produce both epigenetic data for different cell types and positional information of the cells. Thus, these investigations are important for understanding the intra- and inter-cellular control systems of developmental and environmental responses in plants. However, a simple method to detect epigenetic modifications of individual cells in plant tissues is not yet available because detection of the modifications requires immunohistochemistry using specific antibodies. In this study, we developed a simple immunohistochemical method that does not require sectioning to investigate epigenetic modifications. This method uses a clearing system to detect methylated histones, acetylated histones, methylated DNA and/or centromeric histone H3 variants. Analyses of four dicots and five monocots indicated that this method provides a universal technique to investigate epigenetic modifications in diverse plant species.
Highlights
Investigations into the epigenetic status of individual cells within tissues can produce both epigenetic data for different cell types and positional information of the cells
chromatin immunoprecipitation (ChIP) analyses are unable to elucidate the epigenetic modifications in individual cells because the analyses are performed for a mixture of cells, and the results appear as averages of the modification status on chromosomal loci among the cells analyzed
We developed a clearing method for immunohistochemical analyses
Summary
Investigations into the epigenetic status of individual cells within tissues can produce both epigenetic data for different cell types and positional information of the cells. Loci on individual chromosomes are occasionally controlled differentially by different epigenetic modifications depending on the environmental conditions and cell positions, even in the same organism. To elucidate these phenomena, the understandings of the epigenetic modifications in each cell and of the spatio-temporal relationships of the cells are necessary. A number of clearing methods that are not harmful to fluorescent proteins have been developed to investigate the neural networks of animal brains[7,8,9,10] These clearing methods are not easy to apply directly to plant tissues given the existence of chloroplasts and cell walls, both of which emit strong autofluorescence[11]. Rigid cell walls obstruct antibody penetration into the cells[12]
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