(F) Confocal fluorescence sections of living D. mel. embryos (after ~40min at stage5 during embryonic development) expressing an optimized reporter mNeonGreen protein, and carrying hunchback anterior enhancer - tested core promoter - mNeon. The fluorescence signal of mNeonGreen can be seen (in false colors) in the nuclei at the embryo peripheries. The promoters tested correspond to motif knockouts or motif substitution with consensus sequences from the constitutive MED4 (on the left) and the developmental pain (on the right) promoters. The ko motifs or the type of mutations are indicated in white, together with the normalized expression levels (in bracket) measured with our Luciferase assay pipeline. Whereas MED4 promoters drive strong expression along the entire anterior (A) - posterior (P) axis of the embryo, pain embryos show weaker expression, consistently with the expression levels measured in the Luciferase assay. Noteworthy, in contrast to the homogenous AP expression with the constitutive MED4 gene, the A-P expressions patterns for developmental pain resemble the known AP gradient of expression typically observed for the Hb enhancer. The white arrows indicate the fluorescence signals of the nuclei in the anterior part of the embryos. 