Analysis of Genomewide DNA Methylation Reveals Differences in DNA Methylation Levels between Dormant and Naturally as well as Artificially Potentiated Pedicle Periosteum of Sika Deer (Cervus nippon).

Abstract

Deer antlers are the only mammalian appendages that can fully regenerate each year from the permanent bony protuberances of the frontal bones, called pedicles. Pedicle periosteum (PP) is the key tissue for antler regeneration and the source of antler stem cells. The distal one third of the PP has acquired the ability to regenerate antlers and is termed the potentiated PP (PPP), whereas the proximal two thirds of the PP requires further interactions within its niche to launch antler regeneration and is termed the dormant PP (DPP). However, the molecular mechanisms underlying the process of potentiation from the DPP to the PPP are unknown. In this study, we used the fluorescence-labeled methylation-sensitive amplified polymorphism method to assess the levels of DNA methylation in both cells and tissues of the PPP and the DPP. The results showed that the levels of DNA methylation were significantly lower in the PPP compared to the DPP (P < 0.05). Therefore, DNA demethylation may be involved in the process of this potentiation. This involvement was further confirmed by functional testing by artificially creating a potentiated PP (aPPP) from DPP tissue. Moreover, we identified 15 methylated fragments by the methylation sensitive amplified polymorphism method that are either unique to the PPP or the DPP, which were further confirmed by Southern blot analysis. Taken together, our data suggest that DNA demethylation is involved in the process of PP potentiation, which is a prerequisite step for the initiation of antler regeneration. These findings provide the first experimental evidence to link epigenetic regulation and mammalian appendage regeneration.