Neuronal NPR-15 controls the interplay between molecular and behavioral immune responses through the amphid sensory neuron-intestinal axis in C. elegans

Benson Otarigho
Anna Frances Butts
Alejandro Aballay author has email address

Department of Molecular Microbiology & Immunology, Oregon Health & Science University, Portland, OR 97239, USA

https://doi.org/10.7554/eLife.90051.1

Open access
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Figures and data

NPR-15 loss-of-function enhanced pathogen resistance and inhibit avoidance behavior.
A. Wild type (WT), npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals were exposed to P. aeruginosa partial lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Pnp-15::npr-15, P=NS.
B. Colonization of WT, npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals by P. aeruginosa-GFP after 24 hours at 25°C. Scale bar, 200 μm
C. Colony-forming units per animal [WT, npr-15(tm12539), npr-15(tm12539);Pnp-15::npr-15] grown on P. aeruginosa –GFP for 24 hours at 25°C. Bars represent means while error bars indicate SD of three independent experiments; ***p < 0.001.
D. WT, npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals were exposed to S. enterica partial lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Pnp-15::npr-15, P = NS.
E. WT, npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals were exposed to E. faecalis partial lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Pnp-15::npr-15, P = NS.
F. WT, npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals were exposed to S. aureus partial lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Pnp-15::npr-15, P = NS.
G. WT, npr-15(tm12539), and npr-15(tm12539);Pnp-15::npr-15 animals were exposed to live E. coli and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Pnp-15::npr-15, P = NS.
H. WT and npr-15(tm12539) animals were exposed to UV-killed E. coli and scored for survival. WT vs npr-15(tm12539), P=NS.
I. Schematics of avoidance behavior assay on S. aureus.
J. Lawn occupancy of WT C. elegans and npr-15(tm12539) animals on a partial lawn of S. aureus at 24 h. Bars represent means while error bars indicate SD; **p < 0.001, and NS = not significant.

NPR-15 inhibits the expression of immune and aversion-related genes/pathways.
A. Volcano plot of upregulated and downregulated genes in npr-15(tm12539) vs WT animals. Red and blue dots represent significant upregulated and downregulated genes respectively, while the grey dots represent not significant genes.
B. Gene ontology analysis of upregulated genes in npr-15(tm12539) vs WT animals. The cutoff is based on the filtering thresholds of P < 0.05 and arranged according to the representation factor.
C. Representation factors of immune pathways for the upregulated immune genes in npr-15(tm12539) vs WT animals.
D. Venn diagram showing the upregulated immune genes in each pathway in npr-15(tm12539) vs WT animals.
E. qRT-PCR analysis of ELT-2-depenent immune gene expression in WT and npr-15(tm12539) animals. Bars represent means while error bars indicate SD of three independent experiments; *p < 0.05, **p < 0.001 and ***p < 0.0001.
F. qRT-PCR analysis of HLH-30-depenent immune gene expression in WT and npr-15(tm12539) animals. Bars represent means while error bars indicate SD of three independent experiments; *p < 0.05, **p < 0.001 and ***p < 0.0001.

NPR-15 loss-of-function enhances immunity via ELT-2 and HLH-30 when exposed to S. aureus.
A. WT and npr-15(tm12539) animals fed with elt-2 RNAi were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001; elt-2 RNAi, P <0.0001; npr-15(tm12539);elt-2 RNAi, P=NS. elt-2 RNAi vs npr-15(tm12539); elt-2 RNAi, P<0.001.
B. WT and npr-15(tm12539) animals fed with hlh-30 RNAi were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001; hlh-30 RNAi, P <0.05; npr-15(tm12539);hlh-30 RNAi, P<0.05. hlh-30 RNAi vs npr-15(tm12539); hlh-30 RNAi, P<0.0001.
C. WT and npr-15(tm12539) animals fed with hlh-30 and elt-2 RNAi were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001; npr-15(tm12539);hlh-30;elt-2 RNAi, P <0.001. WT;elt-2 RNAi vs npr-15(tm12539);hlh-30;elt-2 RNAi, P=NS.
D. WT and npr-15(tm12539) animals fed with pmk-1 RNAi and animals were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001; pmk-1 RNAi, P <0.0001; npr-15(tm12539);pmk-1 RNAi, P< 0.0001. pmk-1 RNAi vs npr-15(tm12539);pmk-1 RNAi, P<0.0001.
E. WT and npr-15(tm12539) animals fed with daf-16 RNAi were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001; daf-16 RNAi, P =NS; npr-15(tm12539);daf-16 RNAi, P<0.0001. daf-16 RNAi vs npr-15(tm12539);daf-16 RNAi, P<0.0001.
F. WT and npr-15(tm12539) animals fed with skn-1 RNAi were exposed to S. aureus full lawn and scored for survival. EV, empty vector RNAi control. WT EV vs. npr-15(tm12539) EV, P < 0.0001;skn-1 RNAi, P =NS; npr-15(tm12539);skn-1 RNAi, P<0.0001. skn-1 RNAi vs npr-15(tm12539);skn-1 RNAi, P<0.0001.

NPR-15 suppresses molecular immune response via sensory neurons, ASJ.
A. Neuronal connectome of NPR-15-expressing neurons.
B. WT, npr-15(tm12539), and npr-15(tm12539);Prab-3::npr-15 animals were exposed to S. aureus full lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Prab-3::npr-15, P=NS.
C. WT, npr-15(tm12539), ASJ(-), and ASJ(-);npr-15(tm12539) animals were exposed to S. aureus full lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; ASJ(-), P < 0.001; ASJ(-);npr-15(tm12539), P < 0.0001. ASJ(-) vs. ASJ(-);npr-15(tm12539), P =NS.
D. WT, npr-15(tm12539), ASG(-), and ASG(-);npr-15(tm12539) animals were exposed to S. aureus full lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; ASG(-), P < 0.001; ASG(-);npr-15(tm12539), P < 0.0001. ASG(-) vs. ASG(-);npr-15(tm12539), P < 0.05.
E. WT, npr-15(tm12539), ASE(-), and ASE(-);npr-15(tm12539) animals were exposed to S. aureus full lawn and scored for survival. WT vs npr-15(tm12539), P < 0.0001; ASE(-), P < 0.001; ASE(-);npr-15(tm12539), P < 0.0001. ASE(-) vs. ASG(-);npr-15(tm12539), P < 0.05.
F. WT, npr-15(tm12539), npr-15(tm12539);Ptrx-1::npr-15 animals were exposed to S. aureus full lawn and scored for survival. WT vs npr-15(tm12539);Ptrx-1::npr-15, P = NS.
G. qRT-PCR analysis of ELT-2 and HLH-30-depenent immune gene expression in WT and ASJ(-) animals. Bars represent means while error bars indicate SD of three independent experiments; *p < 0.05, **p < 0.001 and ***p < 0.0001.

NPR-15 loss-of-function inhibits pathogen avoidance behavior in an ASJ-dependent manner.
A. WT, npr-15(tm12539), and npr-15(tm12539);Prab-3::npr-15 animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Prab-3::npr-15, P=NS.
B. WT, npr-15(tm12539), ASJ(-), and ASJ(-);npr-15(tm12539) animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.0001; ASJ(-), P < 0.001; ASJ(-);npr-15(tm12539), P < 0.0001. ASJ(-) vs. ASJ(-);npr-15(tm12539), P =NS.
C. WT, npr-15(tm12539), and npr-15(tm12539);Ptrx-1::npr-15 animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.0001; npr-15(tm12539);Ptrx-1::npr-15, P=NS.
D. WT, npr-15(tm12539), ASE(-), and ASE(-);npr-15(tm12539) animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.0001; ASE(-), P < 0.001; ASE(-);npr-15(tm12539), P < 0.05. ASE(-) vs. ASE(-);npr-15(tm12539), P < 0.0001.
E. WT, npr-15(tm12539), ASI(-), and ASI(-);npr-15(tm12539) animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.001; ASI(-), P < 0.001; ASI(-);npr-15(tm12539), P < 0.0001. ASI(-) vs. ASI(-);npr-15(tm12539), P < 0.001.
F. WT, npr-15(tm12539), and AFD(-) animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.001; AFD(-), P =NS.
G. WT, npr-15(tm12539), and AWC(-) animals were exposed to partial lawn of S. aureus and scored for lawn occupancy. WT vs npr-15(tm12539), P < 0.001; AWC(-), P =NS.
H. WT and npr-15(tm12539) fed with gon-2 RNAi and were exposed to the partial lawn of S. aureus and scored for lawn occupancy. EV, empty vector RNAi control. WT EV vs WT;gon-2, P<0.0001; npr-15(tm12539);gon-2, P<0.0001. WT;gon-2 vs npr-15(tm12539);gon-2, P=NS.
I. WT and npr-15(tm12539) fed with gtl-2 RNAi and were exposed to the partial lawn of S. aureus and scored for lawn occupancy. EV, empty vector RNAi control. WT EV vs WT;gtl-2, P<0.001; npr-15(tm12539);gtl-2, P<0.0001. WT;gtl-2 vs npr-15(tm12539);gtl-2, P<0.001.
J. WT, npr-15(tm12539), RNAi intestine-specific strain MGH171, MGH171; npr-15(tm12539) fed with gon-2 RNAi and were exposed to the partial lawn of S. aureus and scored for lawn occupancy. EV, empty vector RNAi control. MGH171 EV vs MGH171;gon-2, P<0.0001. MHG171;gon-2 vs npr-15(tm12539);gon-2, P=NS.
K. WT, npr-15(tm12539), ASJ(-), ASJ(-);npr-15(tm12539) fed with gon-2 RNAi and were exposed to the partial lawn of S. aureus and scored for lawn occupancy. EV, empty vector RNAi control. WT;gon-2 RNAi vs ASJ(-);gon-2, P=NS.

GPCR/NPR-15 suppressed immune response and avoidance behavior via sensory neurons, ASJ. The immune response control is dependent on ELT-2 and HLH-30 transcription factors. While NPR-15 controls avoidance behavior to S. aureus via intestinal-expressed TRPM channel, GON-2.

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