Chromatin and transcriptional regulators act in a cascade to establish a bilateral asymmetry of the C. elegans nervous system

Abstract

Neuroanatomical bilateral asymmetry is a widespread feature in both vertebrates and invertebrates. Although mostly bilaterally symmetric, the nervous system of Caenorhabditis elegans displays bilateral asymmetry. Bilateral asymmetry in C. elegans arises in part from left-right asymmetric cell lineages. The single left-right unpaired MI neuron is normally generated from the right side of an otherwise left-right symmetric cell lineage that on the left gives rise to the e3D epithelial cell. We performed genetic screens and isolated mutants that displayed symmetry in this normally asymmetric cell lineage, with the MI neuron transformed into an e3D-like cell. We identified that a C. elegans Otx homeodomain protein CEH-36 and two basic helix-loop-helix proteins NGN-1 and HLH-2 promote the generation of the MI neuron and are required to establish the bilateral asymmetry in this cell lineage. We found that CEH-36 is asymmetrically expressed and is present in an MI precursor cell on the right but not in an e3D precursor cell on the left. This bilaterally asymmetric CEH-36 expression in turn promotes asymmetric NGN-1 and HLH-2 expression, leading to the generation of the MI neuron on the right side of the cell lineage. The Otx/bHLH transcriptional cascade is evolutionarily conserved, and our results suggest that this transcriptional cascade plays a role in establishing neuroanatomical bilateral asymmetry in other animals. We also discovered that a mutation in a replication-dependent histone H3 gene his-9 transforms the MI neuron into an e3D-like cell. This mutant allele of his-9 causes an altered-function activity that is predicted to impair the interaction of the mutant HIS-9 protein with another histone H3 molecule and inhibit the formation of a histone H3-H4 tetramer. Replication-dependent histones H3-H4 are deposited onto replicating DNA by the heterotrimeric protein complex CAF-1. We observed that loss of function of each of three genes encoding members of the C. elegans CAF-1 complex transformed MI into an e3D-like cell. We propose that CAF-1-mediated nucleosome formation is impaired by the presence of mutant HIS-9 proteins that are unable to form the histone H3-H4 heterotetramer. We also found that two histone-modifying enzymes SET-16 and UTX-1 are required to establish the bilateral asymmetry in this cell lineage. set-16 encodes a protein homologous to the human MLL protein, a histone methyltransferase specific for histone H3 lysine 4, and utx-1 encodes a protein homologous to human UTX protein, a histone demethylase specific for histone H3 lysine 27. Our results reveal a novel mechanism of establishing neuroanatomical bilateral asymmetry and suggest that nucleosome formation and histone H3 modification are required to establish this bilateral asymmetry.