Endoplasmic reticulum stress pathway required for immune homeostasis is neurally controlled by arrestin-1

Abstract

In response to pathogen infection, the host innate immune system activates microbial killing pathways and cellular stress pathways that need to be balanced because insufficient or excessive immune responses have deleterious consequences. Recent studies demonstrate that two G protein-coupled receptors (GPCRs) in the nervous system of Caenorhabditis elegans control immune homeostasis. To investigate further how GPCR signaling controls immune homeostasis at the organismal level, we studied arrestin-1 (ARR-1), which is the only GPCR adaptor protein in C. elegans. The results indicate that ARR-1 is required for GPCR signaling in ASH, ASI, AQR, PQR, and URX neurons, which control the unfolded protein response and a p38 mitogen-activated protein kinase signaling pathway required for innate immunity. ARR-1 activity also controlled immunity through ADF chemosensory and AFD thermosensory neurons that regulate longevity. Furthermore, we found that although ARR-1 played a key role in the control of immunity by AFD thermosensory neurons, it did not control longevity through these cells. However, ARR-1 partially controlled longevity through ADF neurons.

FIGURE 1.

ARR-1 regulates both pathogen resistance and lifespan extension. A–D, wild type (WT) and arr-1(ok401) animals were exposed to P. aeruginosa (A), full lawn of P. aeruginosa (B), S. enterica (C), and Y. pestis (D) and scored for survival. E, wild-type, arr-1(ok401), daf-16(mu86), and daf-16(mu86);arr-1(ok401) mutant animals were scored for lifespan. F, wild-type, arr-1(ok401), daf-16(mu86), and daf-16(mu86);arr-1(ok401) mutant animals were exposed to P. aeruginosa and scored for survival. A–D, differences were statistically significant in all cases (p values <0.0001). E, WT versus arr-1(ok401) (p < 0.0001), arr-1(ok401) versus daf-16(mu86);arr-1(ok401) (p < 0.0001). F, arr-1(ok401) versus daf-16(mu86);arr-1(ok401) (p = 0.0205) are shown. Results represent assays of three to five independent experiments; n = 100–120 animals/strain.

FIGURE 2.

ARR-1 functions in the nervous system to regulate innate immunity. A, wild type (WT), arr-1(ok401), and arr-1(ok401)[pVS3(parr-1::arr-1::gfp):pRF4(rol-6(su1006))] (AY103) animals exposed to P. aeruginosa and scored for survival. B, WT, arr-1(ok401), and AY103 animals exposed to P. aeruginosa/GFP for 24 h, and the cfu were quantified. C, confocal image of AY103 animal expressing ARR-1::GFP. D, WT, arr-1(ok401), and arr-1(ok401)[pVS4(punc-119::arr-1::gfp):pRF4(rol-6(su1006))] (AY104) animals exposed to P. aeruginosa and scored for survival. E, WT, arr-1(ok401), and AY104 animals exposed to P. aeruginosa/GFP for 24 h and the cfu quantified. F, confocal image of AY104 animals expressing ARR-1::GFP in the nervous system. A, WT versus arr-1(ok401) (p < 0.0001), WT versus AY103 (p = 0.7384), arr-1(ok401) versus AY103 (p < 0.0001). D, WT versus AY104 (p = 0.4647), arr-1(ok401) versus AY104 (p < 0.0001). A and D, representative assays of at least three independent experiments; n = 100–120 animals/strain. B and E, mean ± S.E. (error bars). **, p < 0.005; ***, p < 0.0005; n = 4 independent experiments.

FIGURE 3.

The ERP phenotype of arr-1(ok401) animals is independent of OCTR-1 in ASH and ASI neurons. A, WT, arr-1(ok401), octr-1(ok371), and arr-1(ok401);octr-1(ok371) animals were exposed to P. aeruginosa and scored for survival. B, WT, arr-1(ok401), and arr-1(ok401)[pVS5(psra-6::arr-1::gfp):pRF4(rol-6(su1006))] (AY105) animals were exposed to P. aeruginosa and scored for survival. C, WT, arr-1(ok401), and AY105 animals were exposed to P. aeruginosa/GFP for 24 h, and cfu were quantified. D, histogram shows qRT-PCR analysis of abu genes in WT, arr-1(ok401), and AY105 animals exposed to P. aeruginosa for 4 h. A, WT versus octr-1(ok371) (p < 0.0001), arr-1(ok401) versus arr-1(ok401);octr-1(ok371) (p = 0.0006), octr-1(ok371) versus arr-1(ok401);octr-1(ok371) (p < 0.0001). B, representative assays of at least three independent experiments show WT versus AY105 (p < 0.0001), arr-1(ok401) versus AY105 (p = 0.0925). Shown are representative assays of at least three independent experiments; n = 100 animals/strain. C shows mean ± S.E. (error bars); n = 4 independent experiments. D shows mean ± S.E.; n = 3 independent experiments.

FIGURE 4.

NPR-1 partially controls innate immunity through ARR-1 signaling in AQR, PQR, and URX neurons. A, WT, arr-1(ok401), npr-1(ad609), and arr-1(ok401);npr-1(ad609) animals were exposed to P. aeruginosa and scored for survival. B, WT, arr-1(ok401), and arr-1(ok401)[pVS6(pgcy-32::arr-1::gfp):pRF4(rol-6(su1006))] (AY106) animals were exposed to P. aeruginosa and scored for survival. C, WT, arr-1(ok401) and AY106 animals were exposed to P. aeruginosa/GFP for 24 h, and the cfu were quantified. D, histogram shows qRT-PCR analysis of abu genes in WT, arr-1(ok401) and AY106 animals exposed to P. aeruginosa for 4 h. A, WT versus npr-1(ad609) (p = 0.001), WT versus arr-1(ok401);npr-1(ad609) (p < 0.0001), arr-1(ok401) versus arr-1(ok401);npr-1(ad609) (p < 0.0001). B, WT versus AY106 (p < 0.0001), arr-1(ok401) versus AY106 (p < 0.0001). A and B, representative assays of at least three independent experiments; n = 100 animals/strain. C, mean ± S.E. (error bars); n = 4 independent experiments. D, mean ± S.E.; n = 3 independent experiments. **, p < 0.005.

FIGURE 5.

ARR-1 signaling in ADF neurons suppresses innate immunity and longevity. A, WT, arr-1(ok401), and arr-1(ok401)[pVS7(psrh-142::arr-1::gfp):pRF4(rol-6(su1006))] (AY107) animals were scored for lifespan. B, WT, arr-1(ok401), and AY107 animals were exposed to P. aeruginosa and scored for survival. C, WT, arr-1(ok401), and AY107 animals were exposed to P. aeruginosa/GFP for 24 h, and cfu were quantified. D, histogram shows qRT-PCR analysis of abu genes in WT, arr-1(ok401), and AY107 animals exposed to P. aeruginosa for 4 h. A, WT versus AY107 (p < 0.0001), arr-1(ok401) versus AY107 (p = 0.0483). B, WT versus AY107 (p < 0.0001), arr-1(ok401) versus AY107 (p < 0.0001). A and B, representative assays of three independent experiments; n = 100 animals per strain. C, mean ± S.E. (error bars); n = 4 independent experiments. D, mean ± S.E. *, p < 0.05; **, p < 0.005; n = 3 independent experiments.

FIGURE 6.

ARR-1 signaling in AFD neurons suppresses innate immunity. A, WT, arr-1(ok401), and arr-1(ok401)[pVS7(psrh-142::arr-1::gfp):pRF4(rol-6(su1006))] (AY108) animals were scored for lifespan. B, WT, arr-1(ok401), and AY108 animals were exposed to P. aeruginosa and scored for survival. C, histogram shows qRT-PCR analysis of abu genes in WT, arr-1(ok401), and AY108 animals exposed to P. aeruginosa for 4 h. D, WT, arr-1(ok401), and AY108 animals were exposed to P. aeruginosa/GFP for 24 h, and the cfu were quantified. A, WT versus AY108 (p < 0.0001), arr-1(ok401) versus AY108 (p = 0.3224). B, WT versus AY108 (p = 0.0021); arr-1(ok401) versus AY108 (p < 0.0001). A and B, representative assays of at least three independent experiments; n = 100 animals/strain. C, mean ± S.E. (error bars); n = 3 independent experiments. D, mean ± S.E.; n = 4 independent experiments. *, p < 0.05; **, p < 0.005.

FIGURE 7.

Schematic of neural control of immune homeostasis by ARR-1 signaling. In addition to OCTR-1, at least another receptor may signal through ARR-1 in ASH and ASI neurons to control abu gene expression. ARR-1 function in AQR, PQR, and URX neurons appears to play only a small role in the control of abu genes, whereas ARR-1 function in ADF and AFD neurons plays a more important role. ARR-1 signaling in ADF neurons also partially regulates longevity. Chemosensory ADF and thermosensory AFD neurons have unidentified receptors that may be regulated by ARR-1 to control abu gene expression and immune homeostasis.