Mutations in \u3ci\u3eDMRT3\u3c/i\u3e Affect Locomotion in Horses and Spinal Circuit Function in Mice

Locomotion in mammals relies on a central pattern-generating circuitry of spinal interneurons established during development that coordinates limb movement. These networks produce left–right alternation of limbs as well as coordinated activation of flexor and extensor muscles. Here we show that a premature stop codon in the DMRT3 gene has a major effect on the pattern of locomotion in horses. The mutation is permissive for the ability to perform alternate gaits and has a favorable effect on harness racing performance. Examination of wild-type and Dmrt3-null mice demonstrates that Dmrt3 is expressed in the dI6 subdivision of spinal cord neurons, takes part in neuronal specification within this subdivision, and is critical for the normal development of a coordinated locomotor network controlling limb movements. Our discovery positions Dmrt3 in a pivotal role for configuring the spinal circuits controlling stride in vertebrates. The DMRT3 mutation has had a major effect on the diversification of the domestic horse, as the altered gait characteristics of a number of breeds apparently require this mutation

GABAergic and glutamatergic identities of developing midbrain Pitx2 neurons

Pitx2 , a paired-like homeodomain transcription factor, is expressed in post-mitotic neurons within highly restricted domains of the embryonic mouse brain. Previous reports identified critical roles for PITX2 in histogenesis of the hypothalamus and midbrain, but the cellular identities of PITX2-positive neurons in these regions were not fully explored. This study characterizes Pitx2 expression with respect to midbrain transcription factor and neurotransmitter phenotypes in mid-to-late mouse gestation. In the dorsal midbrain, we identified Pitx2 -positive neurons in the stratum griseum intermedium (SGI) as GABAergic and observed a requirement for PITX2 in GABAergic differentiation. We also identified two Pitx2 -positive neuronal populations in the ventral midbrain, the red nucleus, and a ventromedial population, both of which contain glutamatergic precursors. Our data suggest that PITX2 is present in regionally restricted subpopulations of midbrain neurons and may have unique functions that promote GABAergic and glutamatergic differentiation. Developmental Dynamics 240:333–346, 2011. © 2011 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/79425/1/22532_ftp.pd

Characterization of Nkx6-2-Derived Neocortical Interneuron Lineages

Ventral telencephalic progenitors expressing the homeodomain transcription factor Nkx6-2 have been shown to give rise to a multitude of cortical interneuron subtypes usually associated with origin in either the medial ganglionic eminence or the caudal ganglionic eminence. The function of Nkx6-2 in directing the fate of those progenitors has, however, not been thoroughly analyzed. We used a combination of genetic inducible fate mapping and in vivo loss-of-function to analyze the requirement of Nkx6-2 in determining the fate of cortical interneurons. We have found that interneuron subtypes are born with a characteristic temporal pattern. Furthermore, we extend the characterization of interneurons from the Nkx6-2 lineage through the application of electrophysiological methods. Analysis of these populations in Nkx6-2 null mice suggests that there is a small and partially penetrant loss of delayed non-fast spiking somatostatin/calretinin double positive cortical interneurons in the absence of Nkx6-2 gene function

Induction of Olig2+ Precursors by FGF Involves BMP Signalling Blockade at the Smad Level

During normal development oligodendrocyte precursors (OPCs) are generated in the ventral spinal cord in response to Sonic hedgehog (Shh) signalling. There is also a second, late wave of oligodendrogenesis in the dorsal spinal cord independent of Shh activity. Two signalling pathways, controlled by bone morphogenetic protein and fibroblast growth factor (FGF), are active players in dorsal spinal cord specification. In particular, BMP signalling from the roof plate has a crucial role in setting up dorsal neural identity and its inhibition is sufficient to generate OPCs both in vitro and in vivo. In contrast, FGF signalling can induce OPC production from dorsal spinal cord cultures in vitro. In this study, we examined the cross-talk between mitogen-activated protein kinase (MAPK) and BMP signalling in embryonic dorsal spinal cord cultures at the SMAD1/5/8 (SMAD1) transcription factor level, the main effectors of BMP activity. We have previously shown that FGF2 treatment of neural precursor cells (NPCs) derived from rat E14 dorsal spinal cord is sufficient to generate OPCs in vitro. Utilising the same system, we now show that FGF prevents BMP-induced nuclear localisation of SMAD1-phosphorylated at the C-terminus (C-term-pSMAD1). This nuclear exclusion of C-term-pSMAD1 is dependent on MAPK activity and correlates with OLIG2 upregulation, the obligate transcription factor for oligodendrogenesis. Furthermore, inhibition of the MAPK pathway abolishes OLIG2 expression. We also show that SMAD4, which acts as a common partner for receptor-regulated Smads including SMAD1, associates with a Smad binding site in the Olig2 promoter and dissociates from it upon differentiation. Taken together, these results suggest that FGF can promote OPC generation from embryonic NPCs by counteracting BMP signalling at the Smad1 transcription factor level and that Smad-containing transcriptional complexes may be involved in direct regulation of the Olig2 promoter

Oligodendrocytes: biology and pathology

Oligodendrocytes are the myelinating cells of the central nervous system (CNS). They are the end product of a cell lineage which has to undergo a complex and precisely timed program of proliferation, migration, differentiation, and myelination to finally produce the insulating sheath of axons. Due to this complex differentiation program, and due to their unique metabolism/physiology, oligodendrocytes count among the most vulnerable cells of the CNS. In this review, we first describe the different steps eventually culminating in the formation of mature oligodendrocytes and myelin sheaths, as they were revealed by studies in rodents. We will then show differences and similarities of human oligodendrocyte development. Finally, we will lay out the different pathways leading to oligodendrocyte and myelin loss in human CNS diseases, and we will reveal the different principles leading to the restoration of myelin sheaths or to a failure to do so

The role of Nkx proteins neuronal and glial specification

During development, different classes of neurons and glia are generated from proliferative progenitor cells lining the ventricles of the brain and the lumen of the spinal cord. A central issue in developmental neuroscience is to understand the mechanisms by which these cells are generated in space and over time. In the ventral spinal cord, the expression profile of homeodomain (HD) proteins defines five progenitor domains that each will give rise to a distinct type of neuron. Two closely related HD transcriptional repressors, Nkx6.1 and Nkx6.2 (Nkx6), are expressed by progenitors of the ventral spinal cord. We provide evidence that different levels of Nkx6 repressor activity in progenitor cells are a critical determinant of ventral neuronal fate, assigning both redundant and specific roles for these proteins in neuronal specification. A reduction in Nkx6 activity further permits V0 interneurons to be generated from progenitors that lack HD proteins normally required for their generation, providing direct evidence for a model where HD proteins direct specific cell fates by actively repressing the expression of transcription factors that direct alternative fates. In the ventral spinal cord, sMNs and oligodendrocyte precursors (OLPs) are sequentially generated from a domain defined by the expression of Olig2. We show that the generation of sMNs and OLPs in the ventral spinal cord is essentially missing in mice lacking Nkx6 function. In contrast, the same HD proteins instead act to suppress OLP specification in the ventral hindbrain. The divergent roles for Nkx6 proteins seem to reflect that OLPs in the spinal cord and hindbrain are produced by distinct ventral progenitor domains. While a ventral specification of OLPs is well established, it has remained unclear whether also more dorsal progenitor cells give rise to oligodendrocytes. We provide in vivo and in vitro evidence that oligodendrocytes are produced also by dorsal progenitors in the spinal cord and hindbrain and that the specification of these cells may result from the progressive evasion of dorsal BUT signalling over time. Together, our data suggest that oligodendrocytes are generated from multiple dorsoventral origins in the spinal cord and hindbrain, and indicate that the activation of Olig2 at different positions is controlled by distinct genetic programs. The observation that the loss of sMNs in the spinal cord of Nkx6 mutant mice correlates with an extinguished expression of the sMN determinant Olig2 has led to a model where Nkx6 proteins act strictly upstream of Olig2. However, in the hindbrain of Nkx6 mutant mice the initial expression of Olig2 is intact and despite this all sMNs are missing, indicating a parallel requirement for Nkx6 and Olig2 proteins in the generation of sMNs. Visceral motor neurons (vMNs) are generated immediately ventral to sMNs. The transcription factor Phox2b has been found to be an important determinant of these cells, but other factors involved have not been identified. We show that the HD protein Nkx2.2 is sufficient to mediate the expression of Phox2b. Furthermore, while the activities of Nkx6.1 and Nkx6.2 are dispensable for the initial generation of vMNs, they are required to prevent a parallel program of more dorsal interneuronal differentiation and to ensure a proper migration and axonal projection formation of vMNs. Thus, Nkx2 and Nkx6 proteins appear to have complementary roles in the establishment of vMN identity in the hindbrain. Taken together, our data suggest that both visceral and somatic motor neuron differentiation rely on the combined activity of cell intrinsic determinants, rather than on a singe key determinant of neuronal cell fate. Neuronal diversity is established by mechanisms that operate in space and over time. Advances have been made in regard to the mechanisms that restrict and direct neuronal generation in space, but less is known about the mechanisms that underlie how neural progenitors produce distinct types of neurons in a specific temporal order. We addressed this issue by studying a population of progenitor cells in the ventral hindbrain that gives rise to vMNs and serotonergic (S) neurons. Each hindbrain segment, or rhombomere (r), initially generates vMNs, but all the rhombomeres except for r4 switch to producing S neurons at a defined time point. We found that the temporal and spatial generation of vMNs and S neurons critically relies on the integrated activity of Nkx- and Hox-class HD proteins. A primary function of these proteins is to coordinate the activation of Phox2b in space and time. Phox2b, in turn, functions as a binary switch in deciding whether progenitors differentiate into vMNs or serotonergic neurons. Taken together, these data indicate that determinants that control spatial patterning may be associated also with temporal patterning and require that expression patterns are dynamic and modulated over time

Complementary roles for Nkx6 and Nkx2 class proteins in the establishment of motoneuron identity in the hindbrain

The genetic program that underlies the generation of visceral motoneurons in the developing hindbrain remains poorly defined. We have examined the role of Nkx6 and Nkx2 class homeodomain proteins in this process, and provide evidence that these proteins mediate complementary roles in the specification of visceral motoneuron fate. The expression of Nkx2.2 in hindbrain progenitor cells is sufficient to mediate the activation of Phox2b, a homeodomain protein required for the generation of hindbrain visceral motoneurons. The redundant activities of Nkx6.1 and Nkx6.2, in turn, are dispensable for visceral motoneuron generation but are necessary to prevent these cells from adopting a parallel program of interneuron differentiation. The expression of Nkx6.1 and Nkx6.2 is further maintained in differentiating visceral motoneurons, and consistent with this the migration and axonal projection properties of visceral motoneurons are impaired in mice lacking Nkx6.1 and/or Nkx6.2 function. Our analysis provides insight also into the role of Nkx6 proteins in the generation of somatic motoneurons. Studies in the spinal cord have shown that Nkx6.1 and Nkx6.2 are required for the generation of somatic motoneurons, and that the loss of motoneurons at this level correlates with the extinguished expression of the motoneuron determinant Olig2. Unexpectedly, we find that the initial expression of Olig2 is left intact in the caudal hindbrain of Nkx6.1/Nkx6.2 compound mutants, and despite this, all somatic motoneurons are missing. These data argue against models in which Nkx6 proteins and Olig2 operate in a linear pathway, and instead indicate a parallel requirement for these proteins in the progression of somatic motoneuron differentiation. Thus, both visceral and somatic motoneuron differentiation appear to rely on the combined activity of cell intrinsic determinants, rather than on a single key determinant of neuronal cell fate

Different levels of repressor activity assign redundant and specific roles to Nkx6 genes in motor neuron and interneuron specification

Specification of neuronal fate in the vertebrate central nervous system depends on the profile of transcription factor expression by neural progenitor cells, but the precise roles of such factors in neurogenesis remain poorly characterized. Two closely related transcriptional repressors, Nkx6.2 and Nkx6.1, are expressed by progenitors in overlapping domains of the ventral spinal cord. We provide genetic evidence that differences in the level of repressor activity of these homeodomain proteins underlies the diversification of interneuron subtypes, and provides a fail-safe mechanism during motor neuron generation. A reduction in Nkx6 activity further permits V0 neurons to be generated from progenitors that lack homeodomain proteins normally required for their generation, providing direct evidence for a model in which progenitor homeodomain proteins direct specific cell fates by actively suppressing the expression of transcription factors that direct alternative fates