Joint actions of diverse transcription factor families establish neuron-type identities and promote enhancer selectivity

Abstract

To systematically investigate the complexity of neuron-specification regulatory networks we performed an RNA interference (RNAi) screen against all 875 transcription factors (TFs) encoded in Caenorhabditis elegans genome and searched for defects in nine different neuron types of the monoaminergic (MA) superclass and two cholinergic motoneurons. We identified 91 TF candidates required for correct generation of these neuron types of which 28 were confirmed by mutant analysis. We found that correct reporter expression in each individual neuron type requires at least nine different TFs. Individual neuron types do not usually share TFs involved in their specification but share a common pattern of TFs belonging to the five most common TF families: Homeodomain (HD), basic Helix Loop Helix (bHLH), Zinc Finger (ZF), Basic Leucine Zipper Domain (bZIP) and Nuclear Hormone Receptors (NHR). HD TF members are over-represented, supporting a key role for this family in the establishment of neuronal identities. We found a similar TF family distribution for TF mutants with assigned phenotypes in any neuron type in C. elegans, Drosophila or mouse. In addition, we studied terminal differentiation complexity focusing on the dopaminergic terminal regulatory program. We found two HD TFs (UNC-62 and VAB-3) that work together with known dopaminergic terminal selectors (AST-1, CEH-43, CEH-20). Combined TF binding sites for these five TFs constitute a cis-regulatory signature enriched in the regulatory regions of dopaminergic effector genes. Our results provide new insights on neuron-type regulatory programs in C. elegans that could help better understand neuron specification and evolution of neuron types.

Widespread employment of conserved C. elegans homeobox genes in neuronal identity specification

“…Neuronal differentiation defects of the RIC neurons were previously reported in animals lacking two non-homeobox genes, zip-5 and nhr-2 (Jimeno-Martin et al, 2022), but no regulator of the homeobox family was previously known. RIC expresses the MEIS homeobox gene unc-62 (Reilly et al, 2020).…”

“…What is the underlying molecular basis for homeotic identity transformation in unc-86 mutants? The terminal differentiation of all C. elegans dopaminergic neurons requires a combinatorial regulatory signature composed of at least four transcription factors, the ETS domain transcription factor ast-1 , and the three homeobox genes ceh-43/Dlx, unc-62/Meis and ceh-20/Pbx (Doitsidou et al, 2013; Flames and Hobert, 2009; Jimeno-Martin et al, 2022)(Fig.10A). unc-62 and ceh-20 are expressed in all neurons of the anterior deirid lineage (Fig.10A) and the data described above suggests that these factors are all required for each individual neuron fate in this lineage, apparently in combination with more restrictively expressed genes, namely unc-86 in ADA, FLP, AIZ and RMG, and ast-1 and ceh-43 in ADE (Fig.10A).…”

“…Table S4: A regulatory map of transcription factors with a role in terminal neuron differentiation . The basis for this data is taken mainly from (Hobert, 2016a) and supplemented with data from this paper, as well as others, including (Berghoff et al, 2021; Gendrel et al, 2016; Jimeno-Martin et al, 2022; Lloret-Fernandez et al, 2018; Maicas et al, 2021; Reilly et al, 2020; Vidal et al, 2022; Zheng et al, 2022). Criteria to be included in this list is that the transcription factor is expressed throughout embryonic and postembryonic development of the respective neuron type, i.e.…”

“…What is the underlying molecular basis for homeotic identity transformation in unc-86 mutants? The terminal differentiation of all C. elegans dopaminergic neurons requires a combinatorial regulatory signature composed of at least four transcription factors, the ETS domain transcription factor ast-1, and the three homeobox genes ceh-43/Dlx, unc-62/Meis and ceh-20/Pbx (Doitsidou et al, 2013;Flames and Hobert, 2009;Jimeno-Martin et al, 2022)(Fig. 10A).…”

Consensus guidelines on the construct validity of rodent models of restless legs syndrome

“…HPO-9 and DOP-1 proteins function similarly in egg-laying and locomotor behaviors, and HPO-9 deficiency leads to increased transcription of dop-3 ( Lyu et al, 2020a ). Both unc-62 and hpo-9 ( Chen et al, 2022 ) are expressed in dopaminergic neurons, and suppression of unc-62 impairs dopaminergic neuron terminal differentiation and dopamine pathway gene expression ( Lyu et al, 2020b ; Jimeno-Martín et al, 2022 ). Furthermore, interference with unc-62 expression increases expression of ferritin, indicating a possible connection between the MEIS1 gene and iron metabolism in humans ( Catoire et al, 2011 ).…”