ASE Left Research Articles

A Gustatory Neural Circuit of Caenorhabditis elegans Generates Memory-Dependent Behaviors in Na+ Chemotaxis.

Animals show various behaviors in response to environmental chemicals. These behaviors are often plastic depending on previous experiences. Caenorhabditis elegans, which has highly developed chemosensory system with a limited number of sensory neurons, is an ideal model for analyzing the role of each neuron in innate and learned behaviors. Here, we report a new type of memory-dependent behavioral plasticity in Na+ chemotaxis generated by the left member of bilateral gustatory neuron pair ASE (ASEL neuron). When worms were cultivated in the presence of Na+, they showed positive chemotaxis toward Na+, but when cultivated under Na+-free conditions, they showed no preference regarding Na+ concentration. Both channelrhodopsin-2 (ChR2) activation with blue light and up-steps of Na+ concentration activated ASEL only after cultivation with Na+, as judged by increase in intracellular Ca2+ Under cultivation conditions with Na+, photoactivation of ASEL caused activation of its downstream interneurons AIY and AIA, which stimulate forward locomotion, and inhibition of its downstream interneuron AIB, which inhibits the turning/reversal behavior, and overall drove worms toward higher Na+ concentrations. We also found that the Gq signaling pathway and the neurotransmitter glutamate are both involved in the behavioral response generated by ASEL.SIGNIFICANCE STATEMENT Animals have acquired various types of behavioral plasticity during their long evolutionary history. Caenorhabditis elegans prefers odors associated with food, but plastically changes its behavioral response according to previous experience. Here, we report a new type of behavioral response generated by a single gustatory sensory neuron, the ASE-left (ASEL) neuron. ASEL did not respond to photostimulation or upsteps of Na+ concentration when worms were cultivated in Na+-free conditions; however, when worms were cultivated with Na+, ASEL responded and inhibited AIB to avoid turning and stimulated AIY and AIA to promote forward locomotion, which collectively drove worms toward higher Na+ concentrations. Glutamate and the Gq signaling pathway are essential for driving worms toward higher Na+ concentrations.

Development of left/right asymmetry in the Caenorhabditis elegans nervous system: From zygote to postmitotic neuron

Despite their gross morphological symmetry, animal nervous systems can perceive and process information in a left/right asymmetric manner. How left/right asymmetric functional features develop in the context of a bilaterally symmetric structure is a very poorly understood problem, in part because very few morphological or molecular correlates of functional asymmetries have been identified so far in vertebrate or invertebrate nervous systems. One of the very few systems in which a molecular correlate for functional lateralization has been uncovered is the taste sensory system of the nematode Caenorhabditis elegans, which is composed of a pair of bilaterally symmetric neurons, ASE left (ASEL) and ASE right (ASER). ASEL and ASER are similar in morphology, connectivity, and molecular composition, but they express distinct members of a putative chemoreceptor gene family and respond in a fundamentally distinct manner to taste cues. Extensive forward and reverse genetic analysis has uncovered a complex gene regulatory network, composed of transcription factors, miRNAs, chromatin regulators, and intercellular signals, that instruct the asymmetric features of these two neurons. In this review, this system is described in detail, drawing a relatively complete picture of asymmetry control in a nervous system.

From genes to circuits and behaviors: Neuropeptides expand the coding potential of the nervous system.

Neuropeptide signaling remodels the composition of a chemosensory circuit and shapes behavior in Caenorhabditis elegans. We reported that the ASE left (ASEL) salt sensory neuron uses a proprotein convertase, BLI-4, to cleave the insulin-like peptide INS-6. INS-6 peptides are released from the ASEL neuron in response to large, but not small changes in salt stimuli. Fast INS-6 signaling functionally transforms the AWC olfactory sensory neuron into an interneuron in the neural circuit for high salt. This new circuit configuration potentiates behavioral attraction to high salt. Here, in the context of genes, circuits, and behaviors, we discuss the diverse modes of neuropeptide processing and signaling, which expand the coding potential of the nervous system. First, neuropeptide processing and release genes prepare insulin peptides to signal in the nervous system. Second, this neuropeptide signaling diversifies the communication of neural circuits and introduces circuit-level flexibility. Finally, the resulting multisensory neurons and circuits drive finely tuned behavioral choices.

Two distinct types of neuronal asymmetries are controlled by the Caenorhabditis elegans zinc finger transcription factor die-1

Left/right asymmetric features of animals are either randomly distributed on either the left or right side within a population ("antisymmetries") or found stereotypically on one particular side of an animal ("directional asymmetries"). Both types of asymmetries can be found in nervous systems, but whether the regulatory programs that establish these asymmetries share any mechanistic features is not known. We describe here an unprecedented molecular link between these two types of asymmetries in Caenorhabditis elegans. The zinc finger transcription factor die-1 is expressed in a directionally asymmetric manner in the gustatory neuron pair ASE left (ASEL) and ASE right (ASER), while it is expressed in an antisymmetric manner in the olfactory neuron pair AWC left (AWCL) and AWC right (AWCR). Asymmetric die-1 expression is controlled in a fundamentally distinct manner in these two neuron pairs. Importantly, asymmetric die-1 expression controls the directionally asymmetric expression of gustatory receptor proteins in the ASE neurons and the antisymmetric expression of olfactory receptor proteins in the AWC neurons. These asymmetries serve to increase the ability of the animal to discriminate distinct chemosensory inputs.

Genetic Screens forCaenorhabditis elegansMutants Defective in Left/Right Asymmetric Neuronal Fate Specification

We describe here the results of genetic screens for Caenorhabditis elegans mutants in which a single neuronal fate decision is inappropriately executed. In wild-type animals, the two morphologically bilaterally symmetric gustatory neurons ASE left (ASEL) and ASE right (ASER) undergo a left/right asymmetric diversification in cell fate, manifested by the differential expression of a class of putative chemoreceptors and neuropeptides. Using single cell-specific gfp reporters and screening through a total of almost 120,000 haploid genomes, we isolated 161 mutants that define at least six different classes of mutant phenotypes in which ASEL/R fate is disrupted. Each mutant phenotypic class encompasses one to nine different complementation groups. Besides many alleles of 10 previously described genes, we have identified at least 16 novel "lsy" genes ("laterally symmetric"). Among mutations in known genes, we retrieved four alleles of the miRNA lsy-6 and a gain-of-function mutation in the 3'-UTR of a target of lsy-6, the cog-1 homeobox gene. Using newly found temperature-sensitive alleles of cog-1, we determined that a bistable feedback loop controlling ASEL vs. ASER fate, of which cog-1 is a component, is only transiently required to initiate but not to maintain ASEL and ASER fate. Taken together, our mutant screens identified a broad catalog of genes whose molecular characterization is expected to provide more insight into the complex genetic architecture of a left/right asymmetric neuronal cell fate decision.

Early Embryonic Programming of Neuronal Left/Right Asymmetry in C. elegans

Nervous systems are largely bilaterally symmetric on a morphological level but often display striking degrees of functional left/right (L/R) asymmetry. How L/R asymmetric functional features are superimposed onto an essentially bilaterally symmetric structure and how nervous-system laterality relates to the L/R asymmetry of internal organs are poorly understood. We address these questions here by using the establishment of L/R asymmetry in the ASE chemosensory neurons of C. elegans as a paradigm. This bilaterally symmetric neuron pair is functionally lateralized in that it senses a distinct class of chemosensory cues and expresses a putative chemoreceptor family in a L/R asymmetric manner. We show that the directionality of the asymmetry of the two postmitotic ASE neurons ASE left (ASEL) and ASE right (ASER) in adults is dependent on a L-/R-symmetry-breaking event at a very early embryonic stage, the six-cell stage, which also establishes the L/R asymmetric placement of internal organs. However, the L/R asymmetry of the ASE neurons per se is dependent on an even earlier anterior-posterior (A/P) Notch signal that specifies embryonic ABa/ABp blastomere identities at the four-cell stage. This Notch signal, which functions through two T box genes, acts genetically upstream of a miRNA-controlled bistable feedback loop that regulates the L/R asymmetric gene-expression program in the postmitotic ASE cells. Our results link adult neuronal laterality to the generation of the A/P axis at the two-cell stage and raise the possibility that neural asymmetries observed across the animal kingdom are similarly established by very early embryonic interactions.

Searching for neuronal left/right asymmetry: genomewide analysis of nematode receptor-type guanylyl cyclases.

Functional left/right asymmetry ("laterality") is a fundamental feature of many nervous systems, but only very few molecular correlates to functional laterality are known. At least two classes of chemosensory neurons in the nematode Caenorhabditis elegans are functionally lateralized. The gustatory neurons ASE left (ASEL) and ASE right (ASER) are two bilaterally symmetric neurons that sense distinct chemosensory cues and express a distinct set of four known chemoreceptors of the guanylyl cyclase (gcy) gene family. To examine the extent of lateralization of gcy gene expression patterns in the ASE neurons, we have undertaken a genomewide analysis of all gcy genes. We report the existence of a total of 27 gcy genes encoding receptor-type guanylyl cyclases and of 7 gcy genes encoding soluble guanylyl cyclases in the complete genome sequence of C. elegans. We describe the expression pattern of all previously uncharacterized receptor-type guanylyl cyclases and find them to be highly biased but not exclusively restricted to the nervous system. We find that >41% (11/27) of all receptor-type guanylyl cyclases are expressed in the ASE gustatory neurons and that one-third of all gcy genes (9/27) are expressed in a lateral, left/right asymmetric manner in the ASE neurons. The expression of all laterally expressed gcy genes is under the control of a gene regulatory network composed of several transcription factors and miRNAs. The complement of gcy genes in the related nematode C. briggsae differs from C. elegans as evidenced by differences in chromosomal localization, number of gcy genes, and expression patterns. Differences in gcy expression patterns in the ASE neurons of C. briggsae arise from a difference in cis-regulatory elements and trans-acting factors that control ASE laterality. In sum, our results indicate the existence of a surprising multitude of putative chemoreceptors in the gustatory ASE neurons and suggest the existence of a substantial degree of laterality in gustatory signaling mechanisms in nematodes.

A novelC. eleganszinc finger transcription factor,lsy-2, required for the cell type-specific expression of thelsy-6microRNA

The two Caenorhabditis elegans gustatory neurons, ASE left (ASEL) and ASE right (ASER) are morphologically bilaterally symmetric, yet left/right asymmetric in function and in the expression of specific chemosensory signaling molecules. The ASEL versus ASER cell-fate decision is controlled by a complex gene regulatory network composed of microRNAs (miRNAs) and transcription factors. Alterations in the activities of each of these regulatory factors cause a complete lateral cell-fate switch. Here, we describe lsy-2, a novel C2H2 zinc finger transcription factor that is required for the execution of the ASEL stable state. In lsy-2 null mutants, the ASEL neuron adopts the complete ASER gene expression profile, including both upstream regulatory and terminal effector genes. The normally left/right asymmetric ASE neurons are therefore ;symmetrized' in lsy-2 mutants. Cell-specific rescue experiments indicate that lsy-2 is required autonomously in ASEL for the activation of ASEL-specifying factors and the repression of ASER-specifying factors. Genetic epistasis experiments demonstrate that lsy-2 exerts its activity by regulating the transcription of the lsy-6 miRNA in the ASEL neuron, thereby making lsy-2 one of the few factors known to control the cell-type specificity of miRNA gene expression.

Fateful Feedback

Control of cell differentiation is thought to result from bistable regulatory networks that allow a transient developmental signal to cause cells to permanently adopt a differentiated state. In the nematode Caenorhabditis elegans , Johnston et al. examined such a network that determines the alternative fates of two taste receptor neurons, known as ASE left (ASEL) and ASE right (ASER). These neurons are bilaterally symmetrical but express distinct sets of chemoreceptors that are necessary for the worm's navigation in search of food. The authors find through genetic analysis that two key transcriptions factors, DIE-1 and COG-1, which promote expression of genes specific to ASEL and ASER neurons, respectively, act in a feedback loop in which they are linked by two microRNAs encoded by the lsy-6 and mir-273 genes. Expression of lsy-6 is enhanced by DIE-1 transcription factor, and the lsy-6 miRNA inhibits expression of ASER-promoting factor COG-1. COG-1 in turn promotes the expression of mir-273 miRNA, which inhibits expression of the ASEL-inducing gene die-1 . Although the stimulus that causes switching of this loop to favor production of one or the other transcription factor remains unknown, the results provide the essence of a miRNA-containing transcriptional feedback loop that can account for the stabilized expression of terminal cell fate in the ASER and ASEL neurons. R. J. Johnston Jr., S. Chang, J. F. Etchberger, C. O. Ortiz, O. Hobert, MicroRNAs acting in a double-negative feedback loop to control a neuronal cell fate decision. Proc. Natl. Acad. Sci. U.S.A. 102 , 12449-12454 (2005). [Abstract] [Full Text]

The Elegance of the MicroRNAs: A Neuronal Perspective

As knowledge of microRNAs (miRNA) grows from a compendium of sequences to annotated functional data it has become increasingly clear that a highly significant segment of regulatory biology depends on these approximately 22 nucleotide noncoding transcripts. The expression of many miRNAs in the nervous system, some with a high degree of temporal and spatial specificity, suggests that understanding miRNAs in the nervous system will yield rewarding neurobiological insights. High on the list of insights that microRNAs promise is a deeper understanding of the remarkable cellular diversity found among neurons. This review examines the interface between an emerging biology of miRNAs and their role in nervous systems.

MicroRNAs act sequentially and asymmetrically to control chemosensory laterality in the nematode.

Animal microRNAs (miRNAs) are gene regulatory factors that prevent the expression of specific messenger RNA targets by binding to their 3' untranslated region. The Caenorhabditis elegans lsy-6 miRNA (for lateral symmetry defective) is required for the left/right asymmetric expression of guanyl cyclase (gcy) genes in two chemosensory neurons termed ASE left (ASEL) and ASE right (ASER). The asymmetric expression of these putative chemoreceptors in turn correlates with the functional lateralization of the ASE neurons. Here we find that a mutation in the die-1 zinc-finger transcription factor disrupts both the chemosensory laterality and left/right asymmetric expression of chemoreceptor genes in the ASE neurons. die-1 controls chemosensory laterality by activating the expression of lsy-6 specifically in ASEL, but not in ASER, where die-1 expression is downregulated through two sites in its 3' untranslated region. These two sites are complementary to mir-273, a previously uncharacterized miRNA, whose expression is strongly biased towards ASER. Forced bilateral expression of mir-273 in ASEL and ASER causes a loss of asymmetric die-1 expression and ASE laterality. Thus, an inverse distribution of two sequentially acting miRNAs in two bilaterally symmetric neurons controls laterality of the nematode chemosensory system.

A microRNA controlling left/right neuronal asymmetry in Caenorhabditis elegans.

How left/right functional asymmetry is layered on top of an anatomically symmetrical nervous system is poorly understood. In the nematode Caenorhabditis elegans, two morphologically bilateral taste receptor neurons, ASE left (ASEL) and ASE right (ASER), display a left/right asymmetrical expression pattern of putative chemoreceptor genes that correlates with a diversification of chemosensory specificities. Here we show that a previously undefined microRNA termed lsy-6 controls this neuronal left/right asymmetry of chemosensory receptor expression. lsy-6 mutants that we retrieved from a genetic screen for defects in neuronal left/right asymmetry display a loss of the ASEL-specific chemoreceptor expression profile with a concomitant gain of the ASER-specific profile. A lsy-6 reporter gene construct is expressed in less than ten neurons including ASEL, but not ASER. lsy-6 exerts its effects on ASEL through repression of cog-1, an Nkx-type homeobox gene, which contains a lsy-6 complementary site in its 3' untranslated region and that has been shown to control ASE-specific chemoreceptor expression profiles. lsy-6 is the first microRNA to our knowledge with a role in neuronal patterning, providing new insights into left/right axis formation.

A transcriptional regulatory cascade that controls left/right asymmetry in chemosensory neurons of C. elegans.

The molecular mechanisms of differential pattern formation along the left/right (L/R) axis in the nervous system are poorly understood. The nervous system of the nematode Caenorhabditis elegans displays several examples of L/R asymmetry, including the directional asymmetry displayed by the two ASE taste receptor neurons, ASE left (ASEL) and ASE right (ASER). Although bilaterally symmetric in regard to all known morphological criteria, these two neurons display distinct chemosensory capacities that correlate with the L/R asymmetric expression of three putative sensory receptor genes, gcy-5, expressed only in ASER, and gcy-6 and gcy-7, expressed only in ASEL. In order to understand the genetic basis of L/R asymmetry establishment, we screened for mutants in which patterns of asymmetric gcy gene expression are disrupted, and we identified a cascade of several symmetrically and asymmetrically expressed transcription factors that are sequentially required to restrict gcy gene expression to either the left or right ASE cell. These factors include the zinc finger transcription factor che-1; the homeobox genes cog-1, ceh-36, and lim-6; and the transcriptional cofactors unc-37/Groucho and lin-49. Specific features of this regulatory hierarchy are sequentially acting repressive interactions and the finely balanced activity of antagonizing positive and negative regulatory factors. A key trigger for asymmetry is the L/R differential expression of the Nkx6-type COG-1 homeodomain protein. Our studies have thus identified transcriptional mediators of a putative L/R-asymmetric signaling event and suggest that vertebrate homologs of these proteins may have similar functions in regulating vertebrate brain asymmetries.