When given the problem of identifying elements in a biochemical pathway, a geneticist will usually take a genetic approach, one that has worked in previous attempts in other systems. The tactic is to disclose enhancers which exacerbate known mutations. An enhancer is a mutation in the sought-for gene that engenders a more severe phenotype than that caused by the mutation of a single, already characterized gene. Two types of experiments were carried out. Mutations that act as enhancers of such DPP limited embryos cause embryonic lethality.
Mad mutations are also enhancers of mutant dpp appendage phenotypes.
Thus Mad mutants produce a further reduction in wing blade size, a slight reduction in the eye , and loss of tarsal claws. These Mad mutant phenotypes exhibit a close correspondence to dpp mutant phenotypes. Homozygous Mad mutant larvae also show midgut defects and a greatly reduced gastric caecae. DPP signaling from visceral mesoderm to midgut endoderm is required for proper extension of the gastric caecae in parasegment 4 and for the induction of the homeotic gene labial in the adjacent endoderm of parasegment 7. Homozygous Mad mutant embryos lack labial expression and have defects in midgut constriction engendered by labial expression.