The assembly of neural circuits is dependent on precise spatiotemporal expression of cell recognition molecules1-5. Factors controlling cell type specificity have been identified6-8, but how timing is determined remains unknown. Here we describe induction of a cascade of transcription factors by a steroid hormone (ecdysone) in all fly visual system neurons spanning target recognition and synaptogenesis. We demonstrate through single-cell sequencing that the ecdysone pathway regulates the expression of a common set of targets required for synaptic maturation and cell-type-specific targets enriched for cell-surface proteins regulating wiring specificity. Transcription factors in the cascade regulate the expression of the same wiring genes in complex ways, including activation in one cell type and repression in another. We show that disruption of the ecdysone pathway generates specific defects in dendritic and axonal processes and synaptic connectivity, with the order of transcription factor expression correlating with sequential steps in wiring. We also identify shared targets of a cell-type-specific transcription factor and the ecdysone pathway that regulate specificity. We propose that neurons integrate a global temporal transcriptional module with cell-type-specific transcription factors to generate different cell-type-specific patterns of cell recognition molecules regulating wiring.
