Temporal transcription factors determine circuit membership by permanently altering motor neuron-to-muscle synaptic partnerships.

How circuit wiring is specified is a key question in developmental neurobiology. Previously, using the Drosophila motor system as a model, we found the classic temporal transcription factor Hunchback acts in NB7-1 neuronal stem cells to control the number of NB7-1 neuronal progeny form functional synapses on dorsal muscles (Meng et al., 2019). However, it is unknown to what extent control of motor neuron-to-muscle synaptic partnerships is a general feature of temporal transcription factors. Here, we perform additional temporal transcription factor manipulations-prolonging expression of Hunchback in NB3-1, as well as precociously expressing Pdm and Castor in NB7-1. We use confocal microscopy, calcium imaging, and electrophysiology to show that in every manipulation there are permanent alterations in neuromuscular synaptic partnerships. Our data show temporal transcription factors, as a group of molecules, are potent determinants of synaptic partner choice and therefore ultimately control circuit membership.