Comparative analysis of Corazonin-encoding genes (Crz's) in Drosophila species and functional insights into Crz-expressing neurons

To gain insight into regulatory mechanisms of tissue-specific Corazonin (Crz) gene expression and its functions in Drosophila, we cloned the Crz genes from four Drosophila species (D. melanogaster, D. simulans, D. erecta, and D. virilis) and performed comparative analyses of Crz gene sequences and expression patterns using in situ hybridization and immunohistochemistry. Although Crz gene sequences showed a great deal of diversity, its expression patterns in the CNS were highly conserved in the Drosophila species examined here. In D. melanogaster larva, Crz expression was found in four pairs of neurons per cerebral lobe and in eight pairs of bilateral neurons in the ventral nerve cord; in adult, the number of Crz-producing neurons increased to 6-8 in the pars lateralis of each brain lobe, whereas neurons in the ventral nerve cord were no longer detectable. Crz transcripts were also found in the optic lobes; however, these mRNAs do not seem to be translated. Such adult-like Crz expression patterns were established within 48 hours after pupation. Somata of Crz-neurons in the pars lateralis are located in the vicinity of terminals emanating from PDF-containing pacemaking neurons, indicating a functional connection between the two peptidergic nervous systems. A subset of Crz neurons coexpressed the period clock gene; however, normal Crz transcription was unaffected by central clockworks. Two pairs of ectopic Crz cells were detected in the adult brains of behaviorally arrhythmic Clock(Jrk) or cycle(02) mutants, suggesting that CLOCK and CYCLE proteins negatively regulate Crz transcription in a cell-specific manner.