Chinmo function in cockroaches provides new insights into the regulation and evolution of insect metamorphosis

by Jorge Escudero, Judit Gonzalvo, Maria-Dolors Piulachs, Xavier Belles

Insect metamorphosis occurs in two main forms, hemimetaboly (simple) and holometaboly (complete), both regulated by hormonal and genetic pathways involving the transcription factors Krüppel homolog 1 (Kr-h1), Broad-Complex (BR-C), and Ecdysone-induced protein 93F (E93). The BTB–zinc finger protein Chronologically inappropriate morphogenesis (Chinmo), recently identified in the fruit fly Drosophila melanogaster, an holometabolan, as a larval state maintainer, was studied here in the German cockroach, Blattella germanica, an hemimetabolan. We also examined another BTB transcription factor, Abrupt (Ab), based on findings in another holometabolan, the red flour beetle, Tribolium castaneum, suggesting a cooperative role. We characterized chinmo expression in B. germanica and found sustained transcript levels during the N4 and N5 nymphal instars, followed by a marked decline at the final N6 instar. RNA interference (RNAi) knockdown of chinmo at N4 induced precocious metamorphosis two molts later, accompanied by reduced Kr-h1 and elevated E93 expression. Combined knockdown of chinmo and E93 revealed that Chinmo primarily represses E93. Similarly, ab knockdown also triggered precocious metamorphosis, decreasing Kr-h1 and increasing E93 expression; double knockdown of ab and E93 indicated that Ab primarily promotes Kr-h1 expression. These results expand the MEKRE93 pathway by identifying Chinmo and Ab as additional regulators that help maintain the juvenile state in both hemimetabolan and holometabolan insects. Holometaboly likely evolved from hemimetabolan ancestors through the embryonic internalization of wing primordia into imaginal cells, which enabled the emergence of distinct larval forms. Key regulatory factors like Kr-h1, Chinmo, Ab, BR-C, and E93, already present in hemimetabolan lineages, were conserved and rewired in holometabolans. Crucial shifts in this evolutionary transition include Chinmo-mediated inhibition of BR-C and an inversion in the juvenile hormone effect on BR-C, from activation to repression.