View Program - 63rd Annual Drosophila Research Conference

Hox family transcription factors are selectively expressed in distinct segments along the neuraxis of the Drosophila central nervous system. This regional specificity of expression, together with the functional importance of Hox gene-expressing neurons in motor behaviors, makes Hox genes attractive candidates for gene- and region-specific targeting methods. The “Trojan-exon” strategy has been broadly successful in producing gene-specific drivers, but producing such drivers for developmentally important transcription factors, such as Hox genes, has been problematic (Diao F et al, 2015). This may be because these transcription factors are particularly sensitive to truncation of the protein translated from the allele containing the Trojan exon. Truncation necessarily lowers the level of transcription factor expression, but may also result in dominant-negative translation products that suppress transcriptional activity. To address these problems, we have developed a novel type of Trojan exon that incorporates split inteins. The protein fragments derived from a gene whose translation is normally interrupted by the Trojan exon will thus be re-ligated into a full-length active gene product, while at the same time allowing for expression of Gal4 or Split Gal4 components. Using this method, we have successfully made Gal4 drivers for the labial (lab), Proboscipedia (Pb), deformed (dfd), Antenapedia (Antp), Ultrabithorax (Ubx) and Abdominal-A (Abd-A) genes. In addition, we have made Split Gal4 hemidrivers for the lab, dfd, Abd-A and Sex Combs-reduced (Scr) genes. Gal4-driven UAS-reporter expression in the CNS is consistent with the known expression patterns of these genes. In addition, functional manipulation of previously characterized cell types yields the anticipated results. Dfd and other Hox genes play well-established roles in motor function, and we find that our Dfd- and Scr-specific hemidrivers express in glutamatergic neurons, including putative motor neurons. Consistent with this, suppression of neurons targeted by Dfd-Gal4 at the larval stage impairs feeding by preventing the mouth-hooks from being elevated. Conversely, stimulation of these neurons leaves the mouth-hooks constitutively elevated. In adults, stimulation of neurons in the Dfd-Gal4 pattern also alters feeding behavior by eliciting vigorous proboscis extension. Our results suggest that the new Hox-gene specific drivers express faithfully in their respective cell types and will be useful in developmental, physiological, and neural circuit-mapping studies.

Making Hox Gene-specific Drivers Using a Modified Trojan-exon Strategy