Validating the demethylating effects of 5-aza-2'-deoxycytidine in insects requires a whole-genome approach (A reply to Ellers et al.)

We previously demonstrated that treatment with the demethylating agent 5-aza-2′-deoxycytidine (5-azadC) alters the offspring sex ratios produced by females of the parasitoid wasp Nasonia vitripennis (Cook et al. 2015). 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 amongst their sons for mates. Interestingly, treatment with 5-aza-dC did not ablate the facultative sex allocation response. Instead, sex ratios became less female-biased, a shift in the direction of the optimum sex ratio for paternally-inherited alleles according to genomic conflict theory. This was the first (albeit indirect) experimental evidence for genomic conflict over sex allocation. Ellers et al. (2019) have since assayed the effects of 5-aza-dC on DNA methylation in ten Nasonia genes, finding no evidence of demethylation in these 10 genes, from which they conclude that 5-aza-dC has no demethylating capability in Nasonia 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.