C. elegans Males Integrate Food Signals and Biological Sex to Modulate State-Dependent Chemosensation and Behavioral Prioritization

Abstract

Dynamic integration of internal and external cues is essential for flexible, adaptive behavior. In C. elegans, biological sex and feeding state regulate expression of the food-associated chemoreceptor odr-10, contributing to plasticity in food detection and the decision between feeding and exploration. In adult hermaphrodites, odr-10 expression is high, but in well-fed adult males, odr-10 expression is low, promoting exploratory mate-searching behavior. Food-deprivation transiently activates male odr-10 expression, heightening food sensitivity and reducing food leaving. Here, we identify a neuroendocrine feedback loop that sex-specifically regulates odr-10 in response to food deprivation. In well-fed males, insulin-like (insulin/IGF-1 signaling [IIS]) and transforming growth factor β (TGF-β) signaling repress odr-10 expression. Upon food deprivation, odr-10 is directly activated by DAF-16/FoxO, the canonical C. elegans IIS effector. The TGF-β ligand DAF-7 likely acts upstream of IIS and links feeding to odr-10 only in males, due in part to the male-specific expression of daf-7 in ASJ. Surprisingly, these responses to food deprivation are not triggered by internal metabolic cues but rather by the loss of sensory signals associated with food. When males are starved in the presence of inedible food, they become nutritionally stressed, but odr-10 expression remains low and exploratory behavior is suppressed less than in starved control males. Food signals are detected by a small number of sensory neurons whose activity non-autonomously regulates daf-7 expression, IIS, and odr-10. Thus, adult C. elegans males employ a neuroendocrine feedback loop that integrates food detection and genetic sex to dynamically modulate chemoreceptor expression and influence the feeding-versus-exploration decision.

Keywords: C. elegans; TGF-β signaling; behavioral choice; chemoreceptor; chemosensation; feeding; insulin signaling; sex differences; starvation; state dependence.

Figure 1.. Insulin-like signaling regulates odr-10 expression males in response to feeding state.

(A) Representative images of ODR-10::GFP fluorescence in a well fed male, a food-deprived (“starved”) male, and a male re-fed after food deprivation. (B) ODR-10::GFP expression in WT fed and starved (stv) males and hermaphrodites, scored qualitatively on a four-point scale (see Methods). (C) ODR-10::GFP expression in male insulin-signaling mutants. (D) Chemotaxis to diacetyl (1:1000) in WT and daf-2 males. (E) ODR-10::GFP expression in fed and starved WT and daf-16 males. (F) Chemotaxis to diacetyl (1:1000) in WT and daf-16 males either well-fed or starved for 12 hours. (G) ODR-10::GFP expression in hermaphrodite IIS mutants. (H) ODR-10::GFP expression in fed and starved WT and daf-16 hermaphrodites. (I) Model for the regulation of odr-10 by food availability and IIS. In this and subsequent models, dotted lines indicate probable cell non-autonomous interactions. *0.01 < p < 0.05; **0.001 < p < 0.01; ***p < 0.001. Dotted gray brackets indicate p > 0.05. See also Figure S1.

Figure 2.. DAF-16 acts cell-autonomously, and likely directly, to regulate odr-10.

(A) DAF-16::GFP and AWA::mCherry expression. Upper panels show individual channels; lower panel shows a merged, pseudocolored image with a high-magnification inset. To bring about nuclear localization of DAF-16, this strain carried a daf-2 mutation and was grown at 25°C. (B) ODR-10::GFP expression in WT and daf-16 fed and starved (stv) males, with or without (–) AWA::daf-16f transgenes. Results are shown for two independent lines. (C) Diagram depicting the odr-10 genomic locus (upper) and odr-10 transcriptional reporters (middle and lower) with wild-type and mutant versions of the putative DAF-16 binding site. (D) Representative images of Podr-10^WT::GFP and Podr-10^ΔDAF-16::GFP in fed and starved males. Yellow ovals indicate the location of AWA. (E) Quantification of GFP fluorescence in Podr-10^WT::GFP and Podr-10^ΔDAF-16::GFP (two independent lines) in fed and starved males. *0.01 < p < 0.05; **0.001 < p < 0.01; ***p < 0.001. Dotted gray brackets indicate p > 0.05. See also Figure S2.

Figure 3.. odr-10 is sex-specifically regulated by daf-7 TGFβ.

(A) ODR-10::GFP expression in WT and daf-7 males and hermaphrodites. (B) ODR-10::GFP expression in WT and daf-7 hermaphrodites, with and without pan-neural masculinization by Prab-3::fem-3(+). (C) ODR-10::GFP expression in WT and ASJ-feminized (ASJ^f) males. Two independent lines are shown. (D) ODR-10::GFP expression in WT and ASJ-ablated (ASJ⊗) males. Two independent lines are shown. (E) ODR-10::GFP expression in WT and ASJ-ablated hermaphrodites. Two independent lines are shown. (F) ODR-10::GFP expression in WT, daf-7, daf-16, and daf-16; daf-7 mutant males. (G) ODR-10::GFP expression in WT and daf-8 mutants carrying the indicated daf-8 expression constructs. Two independent lines are shown for AWA::daf-8. (H) Model for the regulation of odr-10 by daf-7 and daf-2 (IIS) signaling. In this and following models, the dashed gray line indicates the possibility that daf-7 signaling could act in parallel to IIS. *0.01 < p < 0.05; **0.001 < p < 0.01; ***p < 0.001. Dotted gray brackets indicate p > 0.05. See also Figure S3.

Figure 4.. Sensory perception, not a metabolic cue, regulates odr-10 and daf-7 in males.

(A) ODR-10::GFP expression in males cultured on E. coli OP50, without food, or on aztreonam-treated OP50 for 16-18 h. (B) ODR-10::GFP expression in well-fed males, food-deprived males, and food-deprived males after 24 h recovery on control or aztreonam-treated OP50. (C) ODR-10::GFP expression in hermaphrodites cultured on OP50, without food, or on aztreonam-treated OP50 for 16-18 h. (D-E) ODR-10::GFP expression in males (D) and hermaphrodites (E) cultured on OP50, without food, or on heat-killed OP50 for 16-18 h. (F) ODR-10::GFP expression in WT and eat-2 males and hermaphrodites. (G-H) Pdaf-7::GFP fluorescence intensity in ASJ (G) and ASI (H) on males cultured on OP50, without food, or on aztreonam-treated OP50. (I) Pathway for the regulation of odr-10 by TGFβ and IIS in response to external food signals in males. *0.01 < p < 0.05; **0.001 < p < 0.01; ***p < 0.001. Dotted gray brackets indicate p > 0.05. See also Figure S4.

Figure 5.. Signals from TAX-2/TAX-4 neurons regulate odr-10 expression in males.

(A-B) ODR-10::GFP expression in tax-2, tax-4, daf-16, and daf-16; tax-4 males (A) and hermaphrodites (B). (C) Representative images of Pdaf-7::GFP expression in WT and tax-4 males. Yellow ovals indicate the locations of ASI and ASJ. (D) ODR-10::GFP (kyIs53) expression in WT and tax-4 males with the indicated tax-4 expression constructs. Two separate lines are shown for ASI+ASJ rescue. (E-F) Pdaf-7::GFP fluorescence in ASJ (E) and ASI (F) in tax-4 males with the indicated tax-4 expression constructs. Two separate lines are shown for ASI+ASJ rescue. (G) Food-leaving behavior of well-fed males (“OP50”), starved males (“no food”), and males starved in the presence of inedible food (“OP50+Az”). At the indicated time point, each plate was examined to determine the farthest distance between the food spot and the worm’s track, using the four categories shown in the cartoon to the right. “F” indicates the location of the food spot. (H) Pathway for the regulation of odr-10 by food signals, TGFβ signaling, and IIS in males. *0.01 < p < 0.05; **0.001 < p < 0.01; ***p < 0.001. Dotted gray brackets indicate p > 0.05. See also Figure S5.

Figure 6.. A sex-specific chemosensory feedback loop couples food detection, TGFβ signaling, and IIS to expression of the chemoreceptor odr-10 in adult C. elegans males.

The model shows the proposed mechanism by which chemosensory information about food availability is transmitted through a neuroendocrine loop to regulate odr-10 in AWA. The gray path between ASJ/ASI and AWA represents the possibility that daf-7 signaling may act independently of IIS. This would have to take place through an intermediate cell, as daf-8 does not appear to act in AWA. Pathways at the bottom depict the genetic architecture of the mechanism. For simplicity, the possibility of daf-16-independent functions of daf-7 signaling are not indicated. See Discussion for details.