Preferential demethylation of DNA cytosine on the chromosomes restricted to germ cells in the spermatocytes but not the spermatogonia in the inshore hagfish, Eptatretus burgeri

Chromosome elimination and chromatin diminution occur in various species including single-cell ciliates and several multicellular animals. DNA methylcytosine (5mC) and histone modifications have been identified as markers of the eliminated DNA and chromatins in ciliate and finch. Here we examined the levels of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC; an intermediate of active DNA demethylation) in the male testicular cells of the inshore hagfish Eptatretus burgeri and simultaneously detected germline-restricted repetitive sequences (EEEb1) to identify the chromosomes restricted to germ cells (E-chromosomes). We detected 5mC and 5hmC signals at all chromosomes in the spermatogonia and in all of the interphase nuclei whereas 5mC signals were selectively located on the chromosomes without EEEb1 signals in the spermatocytes’ metaphase, suggesting no 5mC signal on the E-chromosomes. No significant difference in 5hmC levels between the E-chromosomes and the other chromosomes, was detected in the spermatocytes. This chromosome-specific hypomethylation has never been detected in mouse or zebrafish germ cells. These results therefore suggest that the DNA methylation pattern of the E-chromosomes, namely those presumptively eliminated in somatic differentiation, are altered just before or during meiosis. This exclusive alteration of the methylation pattern may play a key role in the chromosome elimination in hagfish species’ embryogenesis.