Epigenetic regulation of caspase-3 gene expression in rat brain development.

The expression levels of caspase-3, a major contributor to the execution of neuronal apoptosis, markedly decrease in the process of brain maturation. We have previously cloned the rat caspase-3 gene promoter and identified its essential regulatory elements. In the present study, we extended previous findings by examining transcriptional regulation of caspase-3 expression in the rat brain of two different ages, corresponding to the immature and mature brain. In particular, we determined that the rate of transcription initiation substantially declines during brain maturation. Furthermore, we established that mRNA levels of Ets1, Ets2, and Sp1 do not change in the brain with maturation, suggesting that these transcription factors do not contribute to age-dependent caspase-3 down-regulation. Hence, we examined a role of DNA methylation and histone modification in this process. Utilizing bisulfite DNA sequencing, we determined the presence of age-dependent differentially methylated fragments within the caspase-3 promoter region. Strikingly, differentially methylated CpG sites correspond to the predicted binding sites for a number of transcription factors that have been previously shown to be involved in neuronal development and differentiation. Moreover, using chromatin immunoprecipitation, we found that mature brains displayed significantly lower levels of histone 3 acetylated Lys14 and histone 4 acetylated Lys5, 8, 12, and 16. This observation is consistent with the decreased level of expression of caspase-3 in the mature brain. Together with our observation that histone deacetylase inhibitor, trichostatin A, increased the level of caspase-3 mRNA in cortical neurons in vitro, these results further indicate an important role of epigenetic factors in the regulation of caspase-3 gene expression.