Abstract
Paternal environmental perturbations, including cocaine intake, can affect the development and behavior of the offspring through epigenetic inheritance. However, the mechanism by which cocaine alters the male germ cells epigenome is almost unexplored. Here, we report that cocaine-treated male mice showed alterations on specific histone post-translational modifications (PTMs) including increased silent chromatin marks H3K9me3 and H3K27me3 and decreased active enhancer and promoter marks H3K27ac and H3K4me3 in isolated germ cells. Also, cocaine increased H3K9ac and H4K16ac levels, involved in the replacement of histones by protamines that take place at round spermatid stage. Cocaine also altered histones H3/H4 epigenetic enzymes by increasing acetyltransferase KAT8/MOF, deacetylase SIRT1 and methyltransferase KMT1C/G9A, and decreasing deacetylases HDAC1/2 and demethylase KDM1A/LSD1 protein levels. Moreover, a pre-treatment with dopamine receptor 1 (DRD1) antagonist SCH23390 (SCH) blocked cocaine effects on H3K4me3, H3K27me3, and H4K16ac epigenetic marks. Interestingly, treatment with SCH-only was able to modify most of the histone marks tested here, pointing to a dopamine role in controlling histone PTMs in germ cells. Taken together, our data suggest a key role for DRD1 in mediating cocaine-triggered epigenetic modifications related to the silencing of gene transcription and the histone-to-protamine replacement that controls chromatin architecture of maturing sperm cells, and pinpoints a novel role of the dopaminergic system in the regulation of male germ cells reprogramming.
Highlights
In the last years, there has been special interest in the characterization of epigenetic mechanisms during spermatogenesis that control the reprogramming of the paternal genome, due to the possible trans-generational transmission of acquired traits (Lacal and Ventura, 2018; Galan et al, 2019)
We have previously reported that male mice spermatogonia express DRD1, and that cocaine administration affects the testicular dopaminergic system decreasing DRD1 mRNA (González et al, 2015)
We evaluated the effect of COC, SCH, and SCH administered min before COC (SCH-COC) treatments on protein expression levels of specific H3/H4 post-translational modifications (PTMs) related to the epigenetic regulation of gene expression and chromatin remodeling during spermatogenesis: (i) H3K9me3 and H3K27me3 as silent chromatin marks, (ii) H3K27ac and H3K4me3 as active enhancer and promoter marks, and (iii) H3K9ac and H4K16ac as marks of open chromatin states and the replacement of histones by protamines
Summary
There has been special interest in the characterization of epigenetic mechanisms during spermatogenesis that control the reprogramming of the paternal genome, due to the possible trans-generational transmission of acquired traits (Lacal and Ventura, 2018; Galan et al, 2019). DNA methylation, and changes in small non-coding RNAs that modulate gene expression in response to basal transcriptional programs and environmental signals (Jenkins and Carrell, 2012). Histones PTMs differentially signal chromatin states such as open/transcription-permissive or closed/repressed, as well as regulatory elements in DNA including active enhancers and promoters (Miller and Grant, 2013). The spermatogenesis in particular is characterized by an epigenetic program that enables the multiple chromatin reorganizations and unique transcriptional regulation that are required for proper meiotic divisions and sperm maturation. It is important to point out that, once paternal DNA compaction has occurred, epigenetic marks may not be altered, creating windows of vulnerability in male germs cells to environmental reprogramming during spermatogenesis (Bale, 2015)
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