NK2-specific domain is responsible for cell death upon ectopic expression of VND in various Drosophila tissues

Drosophila ventral nervous system defective (VND) is a transcription factor containing tinman domain, homeodomain, and NK2-specific domain. The VND plays an essential role during the development of embryonic nervous system, however, little is known about its roles beyond embryonic stage. In this study, the expression patterns and levels of the VND during larval and adult stages were examined by using GAL4/UAS binary expression system and real-time PCR. The VND protein was expressed in larval brain, eye, and wing imaginal discs, implying possible roles of this protein in the larval tissues other than embryonic nervous system. Gain-of-function phenotypes of vnd were analyzed by crossing various GAL4 drivers with UAS - vnd transgenic line. Interestingly, we observed the massive cell death in the tissues including eye and wing discs where GAL4 is expressed. To down-regulate endogenous vnd transcripts in the eye disc, UAS - vnd - IR (inverted repeat) was crossed with ey3.5 - GAL4 line. Loss-of-function phenotypes of vnd also revealed the black ommatidial spots indicating apoptotic cell death in the transgenic eye tissues. To identify a domain within the VND involved in apoptosis, VND mutant proteins were ectopically expressed in the eye disc. In contrast to VND-Δtinman and VND-Y54M mutant proteins, overexpression of VND-ΔNK2 did not lead to cell death, indicating that NK2-specific domain is responsible for apoptosis. Taken together, our results suggest that VND might be involved in the development of the various tissues after embryonic stages and in apoptosis via NK2-specific domain.