Establishment of Motor Neuron-V3 Interneuron Progenitor Domain Boundary in Ventral Spinal Cord Requires Groucho-Mediated Transcriptional Corepression

Abstract

BackgroundDorsoventral patterning of the developing spinal cord is important for the correct generation of spinal neuronal types. This process relies in part on cross-repressive interactions between specific transcription factors whose expression is regulated by Sonic hedgehog. Groucho/transducin-like Enhancer of split (TLE) proteins are transcriptional corepressors suggested to be recruited by at least certain Sonic hedgehog-controlled transcription factors to mediate the formation of spatially distinct progenitor domains within the ventral spinal cord. The aim of this study was to characterize the involvement of TLE in mechanisms regulating the establishment of the boundary between the most ventral spinal cord progenitor domains, termed pMN and p3. Because the pMN domain gives rise to somatic motor neurons while the p3 domain generates V3 interneurons, we also examined the involvement of TLE in the acquisition of these neuronal fates.Methodology and Principal FindingsA combination of in vivo loss- and gain-of-function studies in the developing chick spinal cord was performed to characterize the role of TLE in ventral progenitor domain formation. It is shown here that TLE overexpression causes increased numbers of p3 progenitors and promotes the V3 interneuron fate while suppressing the motor neuron fate. Conversely, dominant-inhibition of TLE increases the numbers of pMN progenitors and postmitotic motor neurons.ConclusionBased on these results, we propose that TLE is important to promote the formation of the p3 domain and subsequent generation of V3 interneurons.

Prolyl isomerase Pin1 and protein kinase HIPK2 cooperate to promote cortical neurogenesis by suppressing Groucho/TLE:Hes1-mediated inhibition of neuronal differentiation

“…We next determined the effect of HIPK2-mediated Gro/TLE1 hyperphosphorylation on the transcriptional repressor function the Gro/TLE1:Hes1 complex. We observed that the previously demonstrated [11][12][13] ability of Hes1 to repress transcription in cells in which Gro/TLE proteins are endogenously expressed was blocked by a dominant-negative form of Gro/TLE, termed Gro/TLE1DQ, 32 showing that Hes1-mediated transcriptional repression depends on Gro/TLE corepressor activity (Figure 4a To determine the mechanisms underlying this inhibitory effect, we examined whether HIPK2 might weaken the association of Gro/TLE1 with Hes1 (Gro/TLE binding-incompetent Hes1DWRPW was used as a negative control). Coimmunoprecipitation experiments showed that neither HIPK2 nor HIPK2(K221R) had a detectable effect on the formation of the Gro/TLE1:Hes1 complex ( Figure 4d, see lanes 4-6).…”

“…Coronal sections through E13.5 mouse brains were rinsed twice in HEPESbuffered saline and then preincubated for 1 h in blocking solution, which was provided by the 'mouse-on-mouse' (MOM) kit purchased from Vector Laboratories Inc (Burlingame, CA, USA). Sections were then incubated for 1-2 h at room temperature in blocking solution containing rat anti-Gro/TLE 11,12,32,38 (i.e., 'panTLE'; 1 : 20) and rabbit anti-Pin1 39 (1 : 500) antibodies.…”

Identification of Multiple Subsets of Ventral Interneurons and Differential Distribution along the Rostrocaudal Axis of the Developing Spinal Cord

“…V3 INs characterized by Sim1 and Nkx2.2 expression arise from p3 progenitors defined by Nkx2.2 [86], [87]. These excitatory commissural neurons diversify into V3 D and V3 V according to their location and their respective electrophysiological characteristics [52].…”

Deciphering principles of morphogenesis from temporal and spatial patterns on the integument

“…3C). Other boundary-sharpening mechanisms have been reported by studying spinal cord neuron specification (Briscoe et al, 2000;Todd et al, 2012).…”