Tlx3 Exerts Direct Control in Specifying Excitatory Over Inhibitory Neurons in the Dorsal Spinal Cord

Abstract

The spinal cord dorsal horn is a major station for integration and relay of somatosensory information and comprises both excitatory and inhibitory neuronal populations. The homeobox gene Tlx3 acts as a selector gene to control the development of late-born excitatory (dILB) neurons by specifying glutamatergic transmitter fate in dorsal spinal cord. However, since Tlx3 direct transcriptional targets remain largely unknown, it remains to be uncovered how Tlx3 functions to promote excitatory cell fate. Here we combined a genomics approach based on chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq) and expression profiling, with validation experiments in Tlx3 null embryos, to characterize the transcriptional program of Tlx3 in mouse embryonic dorsal spinal cord. We found most dILB neuron specific genes previously identified to be directly activated by Tlx3. Surprisingly, we found Tlx3 also directly represses many genes associated with the alternative inhibitory dILA neuronal fate. In both cases, direct targets include transcription factors and terminal differentiation genes, showing that Tlx3 directly controls cell identity at distinct levels. Our findings provide a molecular frame for the master regulatory role of Tlx3 in developing glutamatergic dILB neurons. In addition, they suggest a novel function for Tlx3 as direct repressor of GABAergic dILA identity, pointing to how generation of the two alternative cell fates being tightly coupled.

Long-Term Cultures of Spinal Cord Interneurons

“…Lmx1b+ neurons in our cultures were colocalized with majority of Tlx3 neurons (data not shown), which was also previously confirmed in vivo (Dai et al, 2008). Tlx3 transcription factor, present mostly in laminae I-II, has been identified to have direct control in specifying excitatory neurons in the dorsal horn (Monteiro et al, 2021). Apart from the previously mentioned, mostly excitatory, interneurons, we have also observed a large population of Pax2+ neurons in our cultures.…”

Developmental stage of transplanted neural progenitor cells influences anatomical and functional outcomes after spinal cord injury in mice

“…We also quantified the abundance of dorsal spinal cord postmitotic cells. dILA/dILB interneurons are late-born cells within the superficial dorsal spinal cord laminae I-IV, and include Lbx1 + GABAergic dILA neurons and Tlx3 + glutamatergic dILB neurons 42 . We found that Lbx1 + dILA neurons were significantly more abundant in E13.5 cultures versus E11.5 ( P = 0.0066) and E12.5 ( P = 0.0394) cultures (Fig.…”

Transplantation of Human-Fetal-Spinal-Cord-Derived NPCs Primed with a Polyglutamate-Conjugated Rho/Rock Inhibitor in Acute Spinal Cord Injury

“…We failed to detect GFP expression in any of the analyzed tissues (data not shown). GABAergic inhibitory Lbx1 and glutamatergic excitatory Tlx3 are transcription factors involved in neuronal fate determination of somatosensory interneuron populations located in the dorsal horns of the spinal cord, which modulate and integrate peripheral somatosensory inputs [ 46 ]. A low percentage of hfNPCs displayed Lbx1 and Tlx3 expression in the grafts, while priming with PGA-SS-FAS did not influence this cell fate determination, as we failed to find any significant differences between the groups ( Figure 4 G–I).…”