Yoda, S.

In insects, conspicuous larval pigmentation patterns serve critical ecological roles such as warning signals and mimicry, yet their underlying genetic regulation remains poorly understood. In this study, I investigated the molecular mechanisms underlying black and yellow pigmentation patterns in three distinct larval spot types of the silkworm Bombyx mori: large, diffuse L-spots of the Multilunar (L) mutant; small, sharply defined +p-spots of the Normal strain; and oval pM-hybrid spots of an interspecific hybrid with Bombyx mandarina. Each spot type comprises a yellowish center surrounded by a black periphery, forming crescent-shaped pigmentation patterns. Chemical treatments confirmed that both colors are melanin-based. Using quantitative PCR and RNA interference (RNAi), I analyzed six melanin synthesis genes (Tyrosine Hydroxylase, Dopa Decarboxylase, laccase2, yellow, tan, and ebony) and discovered that black pigmentation involves both dopa/dopamine- and NBAD-melanin synthesis, whereas yellow pigmentation primarily reflects only the latter. I further examined Wnt1 and apontic-like (apt-like) using qPCR, RNAi, and TALEN-mediated mosaic analysis. Wnt1 expression localized to presumptive spot areas, functioning dose-dependently to regulate both spot size and pigment composition: high Wnt1 levels induced larger spots with yellow centers, while reduced Wnt1 expression resulted in black pigmentation and smaller spots. Wnt1-activated transcription factor apt-like was required for pigmentation in all spot types without influencing spot size. Taken together, the results of this study reveal a morphogen-driven gene regulatory network in which Wnt1 dosage and downstream transcriptional cascades orchestrate pigment placement and patterning, offering new insights into the modular genetic control of insect pigmentation.