Butterfly eyespot patterns: evidence for specification by a morphogen diffusion gradient

In this paper we describe a test for Nijhout's (1978, 1980a) hypothesis that the eyespot patterns on butterfly wings are the result of a threshold reaction of the epidermal cells to a concentration gradient of a diffusing degradable morphogen produced by focal cells at the centre of the future eyespot. The wings of the nymphalid butterfly, Bicyclus anynana, have a series of eyespots. each composed of a white pupil, a black disc and a gold outer ring. In earlier extirpation and transplantation experiments (Nijhout 1980a; French and Brakefield, 1995) it has been established that these eyespots are indeed organised around groups of signalling cells active during the first hours of pupal development. If these cells were to supply the positional information for eyespot formation in accordance with Nijhout's diffusion-degradation gradient model, then, when two foci are close together. the signals should sum, and this effect should be apparent in the detailed shape of the resulting pigment pattern. We give an equation for the form of the contours that would be obtained in this manner. We use this to test the morphogen gradient hypothesis on measurements of the outlines of fused eyespots obtained either by grafting focal cells close together, or by using a mutation (Spotty) that produces adjacent fused eyespots. The contours of the fused patterns were found to satisfy our equation, thus corroborating Nijhout's hypothesis to the extent possible with this particular type of experiment.