Transcriptional control of compartmental boundary positioning during Drosophila wing development

Abstract

AbstractThe establishment of tissue axes is fundamental during embryonic development. In the Drosophila wing, the anterior/posterior (AP) and the dorsal/ventral (DV) compartment boundaries provide the basic coordinates around which the tissue develops. These boundaries arise as a result of two lineage decisions, the acquisition of posterior fate by the selector geneengrailed(en)and of dorsal fate by the selector geneapterous(ap). Whileenexpression domain is set up during embryogenesis,apexpression only starts during early wing development. Thus, the correct establishment ofapexpression pattern with respect toenmust be tightly controlled. Here we have functionally investigated the transcriptional inputs integrated by the “early”apenhancer (apE) and their requirement for correct boundary positioning. Detailed mutational analyses using CRISPR/Cas revealed a role of apE in positioning the DV boundary with respect to the AP boundary, with apE mutants often displaying mirror-image anterior wing duplications. We then accomplished tissue-specific enhancer disruption via dCas9 expression. This approach allowed us to dissect the spatio-temporal requirement of apE function, challenging the mechanism by which apE miss-regulation leads to AP defects. Base-pair resolution analyses of apE uncovered a single HOX binding site essential for wing development, which, when mutated, led to wingless flies. Along these lines, we found that the HOX gene Antennapedia (Antp) is fundamental forapexpression. In addition, we demonstrated that the transcription factors Pointed (Pnt), Homothorax (Hth) and Grain (Grn) are necessary for apE function. Together, our results provide a comprehensive molecular basis of earlyapactivation and the developmental consequences of its miss-regulation, shedding light on how compartmental boundaries are be set up during development.