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1. Molecular analyses of zebrafish V0v spinal interneurons and identification of transcriptional regulators downstream of Evx1 and Evx2 in these cells

   https://doi.org/10.1186/s13064-023-00176-w  

   England, Samantha J. ; Rusnock, Amber K. ; Mujcic, Amra ; Kowalchuk, Angelica ; de Jager, Sarah ; Hilinski, William C. ; Juárez-Morales, José L. ; Smith, Matthew E. ; Grieb, Ginny ; Banerjee, Santanu ; et al ( November 2023 , Neural Development)

   V0v spinal interneurons are highly conserved, glutamatergic, commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However, we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells.

   To identify candidate members of V0v gene regulatory networks, we FAC-sorted wild-type andevx1;evx2double mutant zebrafish V0v spinal interneurons and expression-profiled them using microarrays and single cell RNA-seq. We also used in situ hybridization to compare expression of a subset of candidate genes inevx1;evx2double mutants and wild-type siblings.

   Our data reveal two molecularly distinct subtypes of zebrafish V0v spinal interneurons at 48 h and suggest that, by this stage of development,evx1;evx2double mutant cells transfate into either inhibitory spinal interneurons, or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons, plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes arehmx2andhmx3a. Intriguingly, we show that Hmx2/3a, repress dI2 interneuron expression ofskor1aandnefma, two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulateskor1aandnefmaexpression in V0v interneurons by repressing Hmx2/3a expression.

   This study identifies two molecularly distinct subsets of zebrafish V0v spinal interneurons, as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of these cells. Our data further suggest that in the absence of both Evx1 and Evx2, V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then, later, start to express markers of distinct types of inhibitory spinal interneurons, or motoneurons. Taken together, our findings significantly increase our knowledge of V0v and spinal development and move us closer towards the essential goal of identifying the complete gene regulatory networks that specify this crucial cell type.

    

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2. Ebf Activates Expression of a Cholinergic Locus in a Multipolar Motor Ganglion Interneuron Subtype in Ciona

   https://doi.org/10.3389/fnins.2021.784649  

   Popsuj, Sydney ; Stolfi, Alberto ( December 2021 , Frontiers in Neuroscience)

   Conserved transcription factors termed “terminal selectors” regulate neuronal sub-type specification and differentiation through combinatorial transcriptional regulation of terminal differentiation genes. The unique combinations of terminal differentiation gene products in turn contribute to the functional identities of each neuron. One well-characterized terminal selector is COE (Collier/Olf/Ebf), which has been shown to activate cholinergic gene batteries in C. elegans motor neurons. However, its functions in other metazoans, particularly chordates, is less clear. Here we show that the sole COE ortholog in the non-vertebrate chordate Ciona robusta , Ebf, controls the expression of the cholinergic locus VAChT/ChAT in a single dorsal interneuron of the larval Motor Ganglion, which is presumed to be homologous to the vertebrate spinal cord. We propose that, while the function of Ebf as a regulator of cholinergic neuron identity conserved across bilaterians, its exact role may have diverged in different cholinergic neuron subtypes (e.g., interneurons vs. motor neurons) in chordate-specific motor circuits. 

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3. Systematic expression profiling of Dpr and DIP genes reveals cell surface codes in Drosophila larval motor and sensory neurons

   https://doi.org/10.1242/dev.200355  

   Wang, Yupu ; Lobb-Rabe, Meike ; Ashley, James ; Chatterjee, Purujit ; Anand, Veera ; Bellen, Hugo J. ; Kanca, Oguz ; Carrillo, Robert A. ( May 2022 , Development)

   ABSTRACT In complex nervous systems, neurons must identify their correct partners to form synaptic connections. The prevailing model to ensure correct recognition posits that cell-surface proteins (CSPs) in individual neurons act as identification tags. Thus, knowing what cells express which CSPs would provide insights into neural development, synaptic connectivity, and nervous system evolution. Here, we investigated expression of Dpr and DIP genes, two CSP subfamilies belonging to the immunoglobulin superfamily, in Drosophila larval motor neurons (MNs), muscles, glia and sensory neurons (SNs) using a collection of GAL4 driver lines. We found that Dpr genes are more broadly expressed than DIP genes in MNs and SNs, and each examined neuron expresses a unique combination of Dpr and DIP genes. Interestingly, many Dpr and DIP genes are not robustly expressed, but are found instead in gradient and temporal expression patterns. In addition, the unique expression patterns of Dpr and DIP genes revealed three uncharacterized MNs. This study sets the stage for exploring the functions of Dpr and DIP genes in Drosophila MNs and SNs and provides genetic access to subsets of neurons. 

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4. Location, location, location: the organization and roles of potassium channels in mammalian motoneurons

   https://doi.org/10.1113/JP278675  

   Deardorff, Adam S. ; Romer, Shannon H. ; Fyffe, Robert E. W. ( January 2021 , The Journal of Physiology)

   The spatial and temporal balance of spinal α‐motoneuron (αMN) intrinsic membrane conductances underlies the neural output of the final common pathway for motor commands. Although the complete set and precise localization of αMN K⁺channels and their respective outward conductances remain unsettled, important K⁺channel subtypes have now been documented, including Kv1, Kv2, Kv7, TASK, HCN and SK isoforms. Unique kinetics and gating parameters allow these channels to differentially shape and/or modify αMN firing properties, and recent immunohistochemical localization of K⁺‐channel complexes reveals a framework in which their spatial distribution and/or focal clustering within different surface membrane compartments is highly tuned to their physiological function. Moreover, highly evolved regulatory mechanisms enable specific channels to operate over variable levels of αMN activity and contribute to either state‐dependent enhancement or diminution of firing. While recent data suggest an additional, non‐conducting role for clustered Kv2.1 channels in the formation of endoplasmic reticulum–plasma membrane junctions postsynaptic to C‐bouton synapses, electrophysiological evidence demonstrates that conducting Kv2.1 channels effectively regulate αMN firing, especially during periods of high activity in which the cholinergic C‐boutons are engaged. Intense αMN activity or cell injury rapidly disrupts the clustered organization of Kv2.1 channels in αMNs and further impacts their physiological role. Thus, αMN K⁺channels play a critical regulatory role in motor processing and are potential therapeutic targets for diseases affecting αMN excitability and motor output, including amyotrophic lateral sclerosis.image

    

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5. Suppressors of cGAS‐STING are downregulated during fin‐limb regeneration and aging in aquatic vertebrates

   https://doi.org/10.1002/jez.b.23227  

   Mathavarajah, Sabateeshan ; Thompson, Andrew W. ; Stoyek, Matthew R. ; Quinn, T. Alexander ; Roy, Stéphane ; Braasch, Ingo ; Dellaire, Graham ( October 2023 , Journal of Experimental Zoology Part B: Molecular and Developmental Evolution)

   During the early stages of limb and fin regeneration in aquatic vertebrates (i.e., fishes and amphibians), blastema undergo transcriptional rewiring of innate immune signaling pathways to promote immune cell recruitment. In mammals, a fundamental component of innate immune signaling is the cytosolic DNA sensing pathway, cGAS‐STING. However, to what extent the cGAS‐STING pathway influences regeneration in aquatic anamniotes is unknown. In jawed vertebrates, negative regulation of cGAS‐STING activity is accomplished by suppressors of cytosolic DNA such as Trex1, Pml, and PML‐like exon 9 (Plex9) exonucleases. Here, we examine the expression of these suppressors of cGAS‐STING, as well as inflammatory genes and cGAS activity during caudal fin and limb regeneration using the spotted gar (Lepisosteus oculatus) and axolotl (Ambystoma mexicanum) model species, and during age‐related senescence in zebrafish (Danio rerio). In the regenerative blastema of wounded gar and axolotl, we observe increased inflammatory gene expression, including interferon genes and interleukins 6 and 8. We also observed a decrease in axolotlTrex1and garpmlexpression during the early phases of wound healing which correlates with a dramatic increase in cGAS activity. In contrast, theplex9.1gene does not change in expression during wound healing in gar. However, we observed decreased expression ofplex9.1in the senescing cardiac tissue of aged zebrafish, where 2′3′‐cGAMP levels are elevated. Finally, we demonstrate a similar pattern ofTrex1,pml, andplex9.1gene regulation across species in response to exogenous 2′3′‐cGAMP. Thus, during the early stages of limb‐fin regeneration, Pml, Trex1, and Plex9.1 exonucleases are downregulated, presumably to allow an evolutionarily ancient cGAS‐STING activity to promote inflammation and the recruitment of immune cells.

    

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