The Caenorhabditis elegans germ line regulates distinct signaling pathways to control lifespan and innate immunity

Abstract

The relationship between the mechanisms that control an organism's lifespan and its ability to respond to environmental challenges are poorly understood. In Caenorhabditis elegans, an insulin-like signaling pathway modulates lifespan and the innate immune response to bacterial pathogens via a common mechanism involving transcriptional regulation by the DAF-16/FOXO transcription factor. The C. elegans germ line also modulates lifespan in a daf-16-dependent manner. Here, we show that the germ line controls the innate immune response of C. elegans somatic cells to two different Gram-negative bacteria. In contrast to the insulin-like signaling pathway, the germ line acts via distinct signaling pathways to control lifespan and innate immunity. Under standard nematode culture conditions, the germ line regulates innate immunity in parallel to a known p38 MAPK signaling pathway, via a daf-16-independent pathway. Our findings indicate that a complex regulatory network integrates inputs from insulin-like signaling, p38 MAPK signaling, and germ line stem cells to control innate immunity in C. elegans. We also confirm that innate immunity and lifespan in C. elegans are distinct processes, as nonoverlapping regulatory networks control survival in the presence of pathogenic and nonpathogenic bacteria. Finally, we demonstrate that the p38 MAPK pathway in C. elegans is activated to a similar extent by both pathogenic and nonpathogenic bacteria, suggesting that both can induce the nematode innate immune response.

FIGURE 1.

Mutations that prevent germ line proliferation in C. elegans increase resistance to pathogenic bacteria. Depicted are representative survival plots for the indicated C. elegans wild-type or mutant strains, which were exposed to either P. aeruginosa (A–D) or S. marcescens (E–G). Survival studies were initiated with nematodes in the late L4 larval stage. The tested alleles and further statistical data (medians, n, and p values) are listed in supplemental Table 1. The glp-1(e2124) allele was used in all experiments except for B and F, where the glp-1(q224) allele was used. mes-1 F refers to fertile nematodes; mes-1 S refers to sterile nematodes. Some wild-type (w.t.) curves in individual panels are from the same experiment as in other panels. All survival studies were carried out at least three times; the data is representative of these multiple experiments.

FIGURE 2.

The resistance of sterile nematodes to P. aeruginosa is independent of daf-16 under standard nematode culture conditions. Depicted are representative survival plots for the indicated C. elegans wild-type (w.t.) or mutant strains, which were exposed to either E. coli (A and B) or P. aeruginosa (C–H). Survival studies were carried out as described in the legend to Fig. 1, and the alleles tested and further statistical data (medians, n, and p values) are listed in supplemental Table 1. The mu86 (A–D and G) and mgDf47 (E–H) alleles of daf-16 were used. In G and H, survival analysis was carried out under slightly different conditions (modified NGM and different bacterial growth temperature in G, standard NGM and different bacterial growth temperature in H) as described under “Experimental Procedures” and “Results.”

FIGURE 3.

The interaction between the germ line and p38 MAPK pathway. Depicted are representative survival plots for the indicated C. elegans wild-type (w.t.) or mutant strains, which were exposed to either P. aeruginosa (A and C), or E. coli (B and D). Survival studies were carried out as described in the legend to Figs. 1 and 2, and the alleles tested and further statistical data (medians, n, and p values) are listed in supplemental Table 1.

FIGURE 4.

The glp-1 mutation does not alter the expression of genes that are regulated by the p38 MAPK pathway. A, wild-type (N2) or glp-1(e2141) mutant animals were exposed to E. coli or P. aeruginosa, RNA was collected, and antimicrobial gene expression was assayed by quantitative PCR using mlc-1 to normalize RNA concentration. Expression was measured relative to N2 grown in the presence of E. coli. Depicted are the results (±S.E.) of three independent experiments. B, as a control, wild-type and vhp-1(sa366) mutant nematodes were treated similarly, and antimicrobial gene expression was monitored. Expression levels that were significantly different from control (p < 0.05, t test) are indicated with an asterisk.

FIGURE 5.

Mutations that prevent production of the C. elegans germ line do not increase phosphorylation of PMK-1. Production of PMK-1 (A) and phospho-PMK-1 (B and C) in the indicated wild-type (w.t.) or mutant strains and in the presence of the indicated bacteria was quantitated using antibodies specific to the protein and the phosphorylated form of the protein and normalized using antibodies specific for β-tubulin as a reference as described under “Experimental Procedures.” Depicted are the results (±S.E.) of three independent experiments. Alleles used were: glp-1(e2141), glp-4(bn2), and nsy-1(ok593). As indicates wild-type nematodes that were treated with arsenic as described under “Experimental Procedures.” Phospho-PMK-1 levels that were significantly different from wild-type (p < 0.05, t test) are indicated with an asterisk.

FIGURE 6.

A model for the regulation of C. elegans lifespan and innate immunity. A, a schematic demonstrating that daf-2 regulates both nematode lifespan and the innate immune response in a daf-16-dependent manner (based largely on the work in Refs. 3, 4, 13, 52). In contrast, as depicted in B, while the germ line acts through daf-16 to control lifespan (1, 2) under standard nematode growth conditions, the germ line acts independently of daf-16 and in parallel to the p38 MAPK pathway (this work). Under modified nematode growth conditions (depicted with an asterisk), daf-16 does mediate the effect of the germ line on host defense (12, 14). The p38 MAPK pathway also plays a role in lifespan regulation in wild-type (52) but not germ line-deficient nematodes; for simplicity, this is not shown in these models.