Establishment of the paternal methylation imprint of the human H19 and MEST/PEG1 genes during spermatogenesis

Parental-specific epigenetic modifications are imprinted on a subset of gen es in the mammalian genome during germ cell maturation, However, the precis e timing of their establishment remains to be determined. Methylation of Cp G dinucleotides has been shown to be a part of the parental imprint, We hav e examined how the methylation pattern characteristic of the paternal allel e in germ cells are established during human spermatogenesis. Two represent ative imprinted genes, H19 and MEST/PEG1, were studied. The experiments wer e performed using the bisulphite sequencing method on microdissected indivi dual cells at different stages of male germ cell differentiation. We show t hat both genes are unmethylated in fetal spermatogonia, suggesting that all pre-existing methylation imprints are already erased by this stage, The ME ST/PEG1 gene remains unmethylated at all subsequent post-pubertal stages of spermatogenesis, including mature spermatozoa, The methylation of H19 typi cal of the paternal allele first appears in a subset of adult spermatogonia and then is maintained in spermatocytes, spermatids and mature spermatozoa , Our results suggest that the methylation imprint inherited from the paren ts is first erased in the male germ line at an early fetal stage. The pater nal-specific imprint is re-established only later, during spermatogonial di fferentiation in the adult testis.