A Novel Role for the Zinc-Finger Transcription Factor EGL-46 in the Differentiation of Gas-Sensing Neurons in Caenorhabditis elegans

Abstract

Oxygen (O 2 ) and carbon dioxide (CO 2 ) provoke distinct olfactory behaviors via specialized sensory neurons across metazoa. In the nematode C. elegans, the BAG sensory neurons are specialized to sense changes in both O 2 and CO 2 levels in the environment. The precise functionality of these neurons is specified by the coexpression of a membrane-bound receptor-type guanylyl cyclase GCY-9 that is required for responses to CO 2 upshifts and the soluble guanylyl cyclases GCY-31 and GCY-33 that mediate responses to downshifts in O 2 . Expression of these gas-sensing molecules in the BAG neurons is partially, although not completely, controlled by ETS-5, an ETS-domain-containing transcription factor, and EGL-13, a Sox transcription factor. We report here the identification of EGL-46, a zinc-finger transcription factor, which regulates BAG gas-sensing fate in partially parallel pathways to ETS-5 and EGL-13. Thereby, three conserved transcription factors collaborate to ensure neuron type-specific identity features of the BAG gas-sensing neurons.T HE acquisition of specific neuronal fates is controlled by regulatory transcription factors that activate the expression of neuron type-specific terminal differentiation genes (Hobert 2008(Hobert , 2011. Specific individual transcription factors may act to control the expression of the entire terminal gene battery of a specific neuron, or alternatively, multiple transcription factors may act in parallel to control distinct aspects of terminal fates . The use of combinatorial transcription factors to control aspects of terminal fates may provide flexibility to polymodal neurons that need to rapidly respond to changes in a specific environmental cue. We study the BAG neurons in Caenorhabditis elegans that have a dual function where they are activated by separate mechanisms when carbon dioxide (CO 2 ) levels increase or when oxygen (O 2 ) levels decrease (Zimmer et al. 2009). The membrane-bound receptor-type guanylyl cyclase GCY-9 is expressed specifically in the BAG neurons to mediate CO 2 avoidance behavior . Whereas the soluble guanylyl cyclases GCY-31 and GCY-33 are expressed in the BAG neurons to mediate behavioral responses when O 2 levels decrease (Zimmer et al. 2009). Previous studies have shown that the ETS domain-containing transcription factor ETS-5 is critical for CO 2 -sensing neuron fate through direct regulation of GCY-9 expression in the BAG neurons (Guillermin et al. 2011; Brandt et al. 2012;Gramstrup Petersen et al. 2013). In addition, the Sox transcription factor EGL-13 is partially required for GCY-9 expression in the BAG neurons (Gramstrup Petersen et al. 2013). Loss of either of these transcription factors causes defects in CO 2 avoidance behavior (Guillermin et al. 2011; Brandt et al. 2012;Gramstrup Petersen et al. 2013). The O 2 -sensing neuron fate of the BAG neurons, as assessed by expression of gcy-31 and gcy-33, is also defective in ets-5 and egl-13 mutant animals (Gramstrup Petersen et al. 2013). However, the expression of these and other B...

Separate transcriptionally regulated pathways specify distinct classes of sister dendrites in a nociceptive neuron

“…Because other studies have shown that the TEA domain transcription factor EGL-44 typically functions in concert with EGL-46 to specify cell-specific traits in C. elegans (Rojo Romanos et al, 2015; Wu et al, 2001; Yu et al, 2003), we tested an egl-44 mutant for PVD defects. This experiment revealed that the 2° branching defect of egl-44 is virtually identical to that of egl-46 (p>0.05, one-way ANOVA with Tukey’s multiple comparison test) and also results in a modest but significant reduction (~25%) in PVD 2° branches (p<0.001 vs wt, one-way ANOVA with Tukey’s multiple comparison test) (Figure 1e and Supplemental Figure 1e).…”

“…Experiments with additional PVD-specific markers are needed to validate this hypothesis. The strong conservation of MEC-3-related LIM homeodomain proteins and of EGL-44 and EGL-46 in vertebrate genomes argues that similar transcriptional networks may regulate neuronal morphogenesis in the brain (Anbanandam et al, 2006; Desai et al, 1988; Feng et al, 2013; Hunter and Rhodes, 2005; Rojo Romanos et al, 2015; Rosenbaum et al, 2011). …”

“…EGL-46 and its binding partner, the TEA domain transcription enhancer factor EGL-44, function together in C. elegans to regulate cell cycle exit in neural progenitors (Feng et al, 2013; Wu et al, 2001) and to define cell-specific traits in postmitotic neurons (Desai et al, 1988; Rojo Romanos et al, 2015; Wu et al, 2001; Yu et al, 2003). Our work previously revealed an additional role for EGL-46 in dendritic branching: egl-46 mutant PVD neurons show fewer lateral branches than wild type (Smith et al, 2013; 2010; 2012).…”

Behavioral Assays to Study Oxygen and Carbon Dioxide Sensing in Caenorhabditis elegans

“…Using the population behavioral assay, we describe in this protocol how these changes of speed are measurable to enable evaluation of the gas-sensing capability of a population of worms. We have previously used these assays to determine the functional importance of multiple transcription factors required for the development of gas-sensing neurons in C. elegans (Brandt et al , 2012; Gramstrup Petersen et al , 2013; Rojo Romanos et al , 2015 and 2017). In Figure 6, we illustrate examples of results obtained: we show how wild type worms decrease their speed in response to a BAG neuron-mediated downshift (21%-10%) or a URX neuron-mediated upshift (10%-21%) in O 2 concentration (Figure 6A).…”

Pharyngeal pumping inhibition and avoidance by acute exposure to high CO2levels are both regulated by the BAG neurons via different molecular pathways

“…15, 16 More recently, EGL-13 and EGL-46 were also found to be important for the differentiation of the CO 2 -sensing BAG neurons. 17,18 Further studies are required to fully understand the molecular mechanisms induced by high levels of CO 2 and how these mechanisms regulate physiologic responses to elevated CO 2 . We discuss here our recent study of a previously uncharacterized behavioral response of C. elegans to elevated CO 2 , and a new component that is involved in C. elegans CO 2 signaling.…”