Validating the demethylating effects of 5-aza-20-deoxycytidine in insects requires a whole-genome approach (A reply to Ellers et al.)

We previously demonstrated that treatment with the de-methylating agent 5-aza-20-deoxycytidine (5-aza-dC) alters the offspring sex ratios produced by females of the parasitoid wasp Nasonia vitripennis. Females allocate offspring sex ratio in line with local mate competition theory, producing more or less female-biased sex ratios as the number of other females laying eggs on a patch varies, thereby reducing competition among their sons for mates. Interestingly, treatment with 5-aza-dC did not ablate the facultative sex allocation re-sponse. Instead, sex ratios became less female biased, a shift in the direction of the optimum sex ratio for paternally inherited alleles ac-cording to genomic conflict theory. This was the first (albeit indirect) experimental evidence for genomic conflict over sex allocation. In their comment, Ellers and colleagues assayed the effects of 5-aza-dC on DNA methylation in 10 Nasonia genes, finding no evidence of demeth-ylation in these 10 genes, from which they conclude that 5-aza-dC has no demethylating capability in N. vitripennis. Quantifying the efficacy of 5-aza-dC in terms of demethylation is indeed crucial to in-depth interpretation of studies using 5-aza-dC to link phenotypes to epigenetic regulation. Here we outline the mode of action of 5-aza-dC and demonstrate that determining the efficacy of 5-aza-dC in insect systems requires a whole-genome approach