Abstract:  【Aim】 DNA methylation is one of the important ways of gene modification, mainly resulting in 5mC and 6mA. At present, there have been many studies related to 5mC, while studies on 6mA in eukaryotes are rare. Wolbachia bacteria are endosymbionts frequently found in insects. They can mediate their hosts' reproduction via several strategies. Cytoplasmic incompatibility (CI) is the most common defect induced by Wolbachia, which arrests the development of embryos from uninfected females that are mated with Wolbachia-infected males. The molecular mechanisms of CI, however, are not clear. The aim of this study is to investigate the mechanisms of Wolbachia affecting the reproduction of Drosophila from the viewpoint of 6mA methylation. 【Methods】 The model organism D. melanogaster was used as the test insect. Quantitative RT-PCR was adopted to detect the expression levels of DNA 6mA demethylase gene (DMAD) in testes, ovaries and early embryos of Wolbachia-infected and uninfected D.melanogaster flies. The embryos examined were from three different crosses: TT (the control, with both males and females uninfected), TW (viable, males were uninfected while females were Wolbachia-infected), and WT (lethal CI embryos, with males Wolbachia-infected and females uninfected). 【Results】 Wolbachia infection significantly up-regulated the expression of DMAD in testes of 1-day-old flies, but had no effects on DMAD expression in ovaries. In early CI embryos (0.5 h, pre-midblastula transition), DMAD expressionin level was significantly higher than those of the control and TW embryos. In the embryos at midblastula transition (3 h), the transcriptional levels of DMAD in TW and CI were extremely higher than that in the control embryos. In 6 h embryos (post-midblastula transition), however, DMAD expressionin level was the lowest in CI embryos as compared with the control and TW embryos. 【Conclusion】 Wolbachia infection may significantly change 6mA level in testes of their Drosophila hosts, and thus result in death of early embryos derived from Wolbachia-infected males and uninfected females.

Key words: Drosophila melanogaster; Wolbachia, 6mA modification, DMAD, testis, ovary, embryo