The mosquitocidal toxin (MTX) from Bacillus sphaericus and the apoptosis-inducing pierisin-1 from the cabbage butterfly Pieris rapae are two of the most intriguing members of the family of ADP-ribosyltransferases. They are both ∼100 kDa proteins, composed of an N-terminal ADP-ribosyltransferase (∼27 kDa) and a C-terminal putative binding and translocation domain (∼70 kDa) consisting of four ricin-B-like domains. While they both share structural homologies, with an overall amino acid sequence identity of ∼30% that becomes ∼50% at the level of the catalytic core, and functional similarities, notably in terms of enzyme regulation, they seem to largely differ with regard to their targets or cell internalization mechanisms. MTX ADP-ribosylates numerous proteins in lysates of target insect cells at arginine residues, whereas pierisin-1 modifies DNA of insect and mammalian cells by ADP-ribosylation at 2′-deoxyguanosine residues resulting in DNA adducts, mutations and eventually apoptosis. This target specificity differentiates pierisin-1 from all other ADP-ribosyltransferases described so far, and implies that the enzyme must reach the nucleus of target cells. The recently solved crystal structure of MTX catalytic domain is helpful to reveal new insights into structural organization, catalytic mechanisms, proteolytic activation and autoinhibition of both enzymes. The uptake and processing of the ADP-ribosyltransferases is discussed.