Acquisition of proper neuronal identity and position is critical for the formation of neural circuits. In the embryonic spinal cord, cardinal populations of interneurons diversify into specialized subsets and migrate to defined locations within the spinal parenchyma. However, the factors that control interneuron diversification and migration remain poorly characterized. Here, we show that the Onecut transcription factors are necessary for proper diversification and distribution of the V2 interneurons in the developing spinal cord. Furthermore, we uncover that these proteins restrict and moderate the expression of spinal isoforms of
Pou2f2
, a transcription factor known to regulate B-cell differentiation. By gain- or loss-of-function experiments, we show that Pou2f2 contribute to regulate the position of V2 populations in the developing spinal cord. Thus, we uncovered a genetic pathway that regulates the diversification and the distribution of V2 interneurons during embryonic development.
In the developing spinal cord, Onecut transcription factors control the diversification of motor neurons into distinct neuronal subsets by ensuring the maintenance of Isl1 expression during differentiation. However, other genes downstream of the Onecut proteins and involved in motor neuron diversification have remained unidentified. In the present study, we generated conditional mutant embryos carrying specific inactivation of Onecut genes in the developing motor neurons, performed RnA-sequencing to identify factors downstream of onecut proteins in this neuron population, and employed additional transgenic mouse models to assess the role of one specific Onecut-downstream target, the transcription factor Nkx6.2. Nkx6.2 expression was up-regulated in Onecut-deficient motor neurons, but strongly downregulated in Onecut-deficient V2a interneurons, indicating an opposite regulation of Nkx6.2 by onecut factors in distinct spinal neuron populations. Nkx6.2-null embryos, neonates and adult mice exhibited alterations of locomotor pattern and spinal locomotor network activity, likely resulting from defective survival of a subset of limb-innervating motor neurons and abnormal migration of V2a interneurons. Taken together, our results indicate that Nkx6.2 regulates the development of spinal neuronal populations and the formation of the spinal locomotor circuits downstream of the Onecut transcription factors. Movement is the principle expressive output of the central nervous system. Locomotor movements are initiated in the brain either cortically or subcortically, but are expressed by channeling activity to locomotor networks, located primarily in the ventral part of the spinal cord, that include rhythm-generating circuits called Central Pattern Generators (CPGs). The spinal locomotor networks ensure the integration of the multiple inputs that regulate motor activity and control the rhythm, speed, and coordination of the locomotor movements. CPGs are composed of interconnected spinal interneurons (INs) of diverse types that generate patterned activity that is then imposed on the motor neurons (MNs) that innervate skeletal muscles. Although many regulatory genes that direct the differentiation of these spinal neuron populations have been identified and the role of some of these genes has been extensively studied 1 , the complete genetic programs underlying the specification of the various neuron types within locomotor networks are far from fully deciphered.
32 Acquisition of proper neuronal identity and position is critical for the formation of neural 33 circuits. In the embryonic spinal cord, cardinal populations of interneurons diversify into 34 specialized subsets and migrate to defined locations within the spinal parenchyma. 35 However, the factors that control interneuron diversification and migration remain poorly 36 characterized. Here, we show that the Onecut transcription factors are necessary for proper 37 diversification and distribution of the V2 interneurons in the developing spinal cord. 38 Furthermore, we uncover that these proteins restrict and moderate the expression of spinal 39 isoforms of Pou2f2, a transcription factor known to regulate B-cell differentiation. By gain-40 or loss-of-function experiments, we show that Pou2f2 contribute to regulate the position of 41 V2 populations in the developing spinal cord. Thus, we uncovered a genetic pathway that 42 regulates the diversification and the distribution of V2 interneurons during embryonic 43 development. 44 45 46 Significance statement 47 In this study, we identify the Onecut and Pou2f2 transcription factors as regulators of spinal 48 interneuron diversification and migration, two events that are critical for proper CNS 49 development. 50 51
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