A novel temporal identity window generates alternating cardinal motor neuron subtypes in a single progenitor lineage

Abstract

11Spatial patterning specifies neural progenitor identity, with further diversity generated by 12 temporal patterning within individual progenitor lineages. These mechanisms generate cardinal 13 classes of motor neurons (sharing a transcription factor identity and common muscle group 14 targets). In Drosophila, two cardinal classes are Even-skipped (Eve)+ motor neurons projecting 15 to dorsal muscles and Nkx6+ motor neurons projecting to ventral muscles. The Drosophila 16 neuroblast 7-1 (NB7-1) lineage generates distinct Eve+ motor neurons via the temporal 17 transcription factor (TTF) cascade Hunchback (Hb)-Krüppel (Kr)-Pdm-Castor (Cas). Here we 18 show that a newly discovered Kr/Pdm temporal identity window gives rise to an Nkx6+ Eve-19 motor neuron projecting to ventral oblique muscles, resulting in alternation of cardinal motor 20 neuron subtypes from a single progenitor (Eve>Nkx6>Eve). We show that co-overexpression of 21 Kr/Pdm generates ectopic VO motor neurons within the NB7-1 lineage and that Kr/Pdm act via 22 Nkx6, which itself is necessary and sufficient for VO motor neuron identity. Lastly, Nkx6 is 23 required for ventral oblique muscle targeting, thereby linking temporal patterning to motor 24 neuron morphology and synaptic target selection. In conclusion, we show that one neuroblast 25 lineage generates interleaved cardinal motor neurons fates; that the Kr/Pdm TTFs form a novel 26 temporal identity window that promotes expression of Nkx6; and that the Kr/Pdm>Nkx6 27 pathway is necessary and sufficient to specify VO motor neuron identity and morphology. 28 29 Introduction 30 31Neural diversity from flies to mice arises from two major developmental mechanisms. First, neural 32 progenitors acquire a unique and heritable spatial identity based on their position along the 33 rostrocaudal or dorsoventral body axes (Kohwi and Doe, 2013; Sagner and Briscoe, 2019). 34Second, temporal patterning based on neuronal birth-order results in individual progenitors 35 producing a diverse array of neurons and glia (Holguera and Desplan, 2018; Kohwi and Doe, 2013; 36 Miyares and Lee, 2019). Temporal patterning is best characterized in Drosophila; neural 37 progenitors (neuroblasts) located in the ventral nerve cord, central brain, and optic lobes all 38 undergo temporal patterning, in which the neuroblast sequentially expresses a cascade of TTFs 39 that specify distinct neuronal identities (Allan and Thor, 2015; Doe, 2017; Holguera and Desplan, 40 2018; Miyares and Lee, 2019). Although all neuroblasts undergo temporal patterning, the TTFs are 41 different in each region of the brain (Allan and Thor, 2015; Doe, 2017; Holguera and Desplan, 2018; 42 2Miyares and Lee, 2019). Similar mechanisms are used in the mammalian cortex, retina, and spinal 43 cord, although many TTFs remain to be identified (Alsiö et al., 2013; Delile et al., 2019; Elliott et al., 44 2008; Mattar et al., 2015; Sockanathan and Jessell, 1998; Stam et al., 2011). 45A major open question is how transient expression of TTFs like Kr and ...