Antennapedia and optix regulate metallic silver wing scale development and cell shape in Bicyclus anynana butterflies

Butterfly wing scales can develop intricate cuticular nanostructures that produce silver colors, but the underlying genetic and physical basis of such colors is mostly unexplored. Here, we characterize different types of wild-type silver scales in Bicyclus anynana butterflies and show that the varying thickness of the air layer between two cuticular laminas is most important for producing silvery broadband reflectance. We then address the function of five genes-apterous A, Ultrabithorax, doublesex, Antennapedia, and optix-in silver scale development by examining crispants with either ectopic gains or losses of silver scales. Simultaneous transformations of three parameters-loss of the upper lamina, increased lower lamina thickness, and increased pigmentation-occur when silver scales become brown and vice versa when brown scales become silver. Antennapedia and optix are high-level regulators of different silver scale types and determine cell shape in both sexes. Moreover, Antennapedia is involved in determining ridge and crossrib orientation.