Figure 2

Reproductive sterility using a homing endonuclease controlled by the inducible Q-system in combination with site-specific recombination. The proposed reproductive sterility system is based on the inducible binary expression system Q [48], in which quinic acid (QA) acts as an inducer that hinders the repressor QS from complexing the transcriptional activator QF, which can activate its target genes by binding to a Q upstream activation sequence (QUAS). To generate male reproductive sterility systems the spermatogenesis-specific promoter of the β2 tubulin (β2t) gene can be suitably used to affect either the sperm itself or the progeny sired by the sperm. The Q system can be combined with a recombinase mediated transcription regulation system to render the induction of an effector gene expression permanent and independent of the presence of the inducer QA. In this dual system, QF drives the expression of a site-specific recombinase (FLP) that can in turn remove a flp-out cassette [57], which contains a transcriptional terminator (SV40) and is flanked by flp recombinant target sites (FRTs) in direct orientation. After the removal of the transcriptional terminator, the directed expression of an effector gene is mediated by the tissue-specific promoter 5' to the FRT. Since the Q system components are superfluous after the activation of the effector gene, they can also be placed into the flp-out cassette. To make sure that both components of the Q system are translated in a bi-cistronic messenger RNA, they will be separated by an internal ribosome entry site (IRES). A homing endonuclease targeting the progeny genome can be employed as an effector that would kill the progeny but not the sperm itself [34]. During regular rearing this male reproductive sterility would be kept in an OFF state, as at the absence of QA the repressor QS will mask QF and block its activation potential. Only after the addition of QA to the food in the release generation, QS will be inactivated and QF thereby allowed to activate the expression of the flp recombinase (FLP), which in turn would remove the Q system regulators and at the same time mediate the expression of the homing endonuclease that could block development of the next generation and thus cause male reproductive sterility.