Definition of the molecular and structural aspects of sex determining splicing regulators: new opportunities for the population control of agricultural insect pests

The control of agricultural insect pests represents an excellent alternative to the use of pesticides which may produce severe environmental problems. Effective strategies to control the insect population that produces heavy damage to cultivations consist in inducing its local reduction by the Sterile Insect Technique (SIT). However, the success of these approaches relies also on understanding the molecular basis underlying the sex determination process in these species. Although information about the molecular mechanism that drives the sex destiny is extremely poor, in all these invasive species an important role is played by alternative splicing processes responsible for specifying male or female development. In this scenario, the present project (TRA2TRA) aims at characterizing the interaction between crucial splicing factors encoded by transformer (tra) and transformer-2 (tra2) genes, as this partnership is fundamental for female sex determination in several agricultural pests such as C. capitata (medfly), B. oleae (olive fly) and B. dorsalis (oriental fly). Here, we initially plan to unravel the structural basis of their recognition in medfly using a novel approach based on recombinant proteins and synthetic peptides. Then, synthetic peptides able to compete with the physiological TRA-TRA2 interaction, will be designed and characterized in-vitro. The functional ability of these peptides to masculinize C. capitata XX embryos will be tested in-vivo. Moreover, the abundance of recurrent Arg-Gly motifs in TRA and TRA2 sequences raises the intriguing possibility that the TRA2/TRA binding may generate functional condensates. The propensity of complexes formed by TRA-TRA2 and their fragments to induce phase separation will be explored. Finally, to overcome previous failures, novel CcTRA2 variants with reduced thermal stability at high temperatures will be developed to facilitate the temperature-dependent sex-reversal toward maleness experiments on these insects.