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Stimulation of host immune defenses by a small molecule protects C. elegans from bacterial infection.

Pukkila-Worley R, Feinbaum R, Kirienko NV, Larkins-Ford J, Conery AL, Ausubel FM - PLoS Genet. (2012)

Bottom Line: We previously used a bacterial infection assay in C. elegans to identify 119 compounds that affect host-microbe interactions among 37,214 tested.Using transcriptome profiling, epistasis pathway analyses with C. elegans mutants, and an RNAi screen, we show that RPW-24 promotes resistance to Pseudomonas aeruginosa infection by inducing the transcription of a remarkably small number of C. elegans genes (∼1.3% of all genes) in a manner that partially depends on the evolutionarily-conserved p38 MAP kinase pathway and the transcription factor ATF-7.These data show that the immunostimulatory activity of RPW-24 is required for its efficacy and define a novel C. elegans-based strategy to identify compounds with activity against antibiotic-resistant bacterial pathogens.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The nematode Caenorhabditis elegans offers currently untapped potential for carrying out high-throughput, live-animal screens of low molecular weight compound libraries to identify molecules that target a variety of cellular processes. We previously used a bacterial infection assay in C. elegans to identify 119 compounds that affect host-microbe interactions among 37,214 tested. Here we show that one of these small molecules, RPW-24, protects C. elegans from bacterial infection by stimulating the host immune response of the nematode. Using transcriptome profiling, epistasis pathway analyses with C. elegans mutants, and an RNAi screen, we show that RPW-24 promotes resistance to Pseudomonas aeruginosa infection by inducing the transcription of a remarkably small number of C. elegans genes (∼1.3% of all genes) in a manner that partially depends on the evolutionarily-conserved p38 MAP kinase pathway and the transcription factor ATF-7. These data show that the immunostimulatory activity of RPW-24 is required for its efficacy and define a novel C. elegans-based strategy to identify compounds with activity against antibiotic-resistant bacterial pathogens.

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RPW-24 is toxic to C. elegans.(A) Wild-type C. elegans leave a lawn of E. coli containing 70 µM RPW-24 more so than DMSO after 16 hours of exposure. (B) The average percentage of wild-type C. elegans that were off a lawn of E. coli supplemented with either DMSO or 70 µM RPW-24 after 16 hours of exposure from three plates per condition [as in (A)] is presented with error bars representing SEM. P = 0.002 for the comparison of the two conditions. (C) RPW-24 shortens the lifespan of wild-type C. elegans in a dose-dependent manner. Data at each time point are the average of three plates per strain, each with approximately 50 animals per plate (sample sizes are given in Table S2). (D) RPW-24 slows development of wild-type C. elegans. Developmental stage was accessed in 50 animals per treatment group and presented as the percentage of the population that was at the indicated development stage after 65 hours of incubation at 20°C.
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pgen-1002733-g006: RPW-24 is toxic to C. elegans.(A) Wild-type C. elegans leave a lawn of E. coli containing 70 µM RPW-24 more so than DMSO after 16 hours of exposure. (B) The average percentage of wild-type C. elegans that were off a lawn of E. coli supplemented with either DMSO or 70 µM RPW-24 after 16 hours of exposure from three plates per condition [as in (A)] is presented with error bars representing SEM. P = 0.002 for the comparison of the two conditions. (C) RPW-24 shortens the lifespan of wild-type C. elegans in a dose-dependent manner. Data at each time point are the average of three plates per strain, each with approximately 50 animals per plate (sample sizes are given in Table S2). (D) RPW-24 slows development of wild-type C. elegans. Developmental stage was accessed in 50 animals per treatment group and presented as the percentage of the population that was at the indicated development stage after 65 hours of incubation at 20°C.

Mentions: To determine if RPW-24 adversely affects wild-type nematodes growing in the absence of pathogen, we first used a behavioral assay designed to study the aversion response of C. elegans to xenobiotic toxins [22]. The addition of some poisons to the center of small lawns of non-pathogenic E. coli causes C. elegans animals to leave the lawn, presumably to minimize toxin exposure. Interestingly, we observed a significant aversion response to 70 µM RPW-24 (Figure 6A and 6B). After 16 hours, 51% of the nematodes had left the lawn containing RPW-24, whereas only 7% of animals left a control lawn (P = 0.002; Figure 6B). Next, we conducted a lifespan assay on nematode growth media supplemented with either DMSO or varying concentrations of RPW-24 and found that RPW-24 shortened C. elegans lifespan in a dose-dependent manner (Figure 6C), with 70 µM RPW-24 resulting in a 24% reduction in median lifespan. Interestingly, we observed lifespan shortening only at compound concentrations that rescued C. elegans from P. aeruginosa infection [7, 35 and 70 µM, but not 0.7 µM]. We also found that 70 µM RPW-24 slowed the development of animals when they were exposed at the first larval stage (L1)(Figure 6D).


Stimulation of host immune defenses by a small molecule protects C. elegans from bacterial infection.

Pukkila-Worley R, Feinbaum R, Kirienko NV, Larkins-Ford J, Conery AL, Ausubel FM - PLoS Genet. (2012)

RPW-24 is toxic to C. elegans.(A) Wild-type C. elegans leave a lawn of E. coli containing 70 µM RPW-24 more so than DMSO after 16 hours of exposure. (B) The average percentage of wild-type C. elegans that were off a lawn of E. coli supplemented with either DMSO or 70 µM RPW-24 after 16 hours of exposure from three plates per condition [as in (A)] is presented with error bars representing SEM. P = 0.002 for the comparison of the two conditions. (C) RPW-24 shortens the lifespan of wild-type C. elegans in a dose-dependent manner. Data at each time point are the average of three plates per strain, each with approximately 50 animals per plate (sample sizes are given in Table S2). (D) RPW-24 slows development of wild-type C. elegans. Developmental stage was accessed in 50 animals per treatment group and presented as the percentage of the population that was at the indicated development stage after 65 hours of incubation at 20°C.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3375230&req=5

pgen-1002733-g006: RPW-24 is toxic to C. elegans.(A) Wild-type C. elegans leave a lawn of E. coli containing 70 µM RPW-24 more so than DMSO after 16 hours of exposure. (B) The average percentage of wild-type C. elegans that were off a lawn of E. coli supplemented with either DMSO or 70 µM RPW-24 after 16 hours of exposure from three plates per condition [as in (A)] is presented with error bars representing SEM. P = 0.002 for the comparison of the two conditions. (C) RPW-24 shortens the lifespan of wild-type C. elegans in a dose-dependent manner. Data at each time point are the average of three plates per strain, each with approximately 50 animals per plate (sample sizes are given in Table S2). (D) RPW-24 slows development of wild-type C. elegans. Developmental stage was accessed in 50 animals per treatment group and presented as the percentage of the population that was at the indicated development stage after 65 hours of incubation at 20°C.
Mentions: To determine if RPW-24 adversely affects wild-type nematodes growing in the absence of pathogen, we first used a behavioral assay designed to study the aversion response of C. elegans to xenobiotic toxins [22]. The addition of some poisons to the center of small lawns of non-pathogenic E. coli causes C. elegans animals to leave the lawn, presumably to minimize toxin exposure. Interestingly, we observed a significant aversion response to 70 µM RPW-24 (Figure 6A and 6B). After 16 hours, 51% of the nematodes had left the lawn containing RPW-24, whereas only 7% of animals left a control lawn (P = 0.002; Figure 6B). Next, we conducted a lifespan assay on nematode growth media supplemented with either DMSO or varying concentrations of RPW-24 and found that RPW-24 shortened C. elegans lifespan in a dose-dependent manner (Figure 6C), with 70 µM RPW-24 resulting in a 24% reduction in median lifespan. Interestingly, we observed lifespan shortening only at compound concentrations that rescued C. elegans from P. aeruginosa infection [7, 35 and 70 µM, but not 0.7 µM]. We also found that 70 µM RPW-24 slowed the development of animals when they were exposed at the first larval stage (L1)(Figure 6D).

Bottom Line: We previously used a bacterial infection assay in C. elegans to identify 119 compounds that affect host-microbe interactions among 37,214 tested.Using transcriptome profiling, epistasis pathway analyses with C. elegans mutants, and an RNAi screen, we show that RPW-24 promotes resistance to Pseudomonas aeruginosa infection by inducing the transcription of a remarkably small number of C. elegans genes (∼1.3% of all genes) in a manner that partially depends on the evolutionarily-conserved p38 MAP kinase pathway and the transcription factor ATF-7.These data show that the immunostimulatory activity of RPW-24 is required for its efficacy and define a novel C. elegans-based strategy to identify compounds with activity against antibiotic-resistant bacterial pathogens.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America.

ABSTRACT
The nematode Caenorhabditis elegans offers currently untapped potential for carrying out high-throughput, live-animal screens of low molecular weight compound libraries to identify molecules that target a variety of cellular processes. We previously used a bacterial infection assay in C. elegans to identify 119 compounds that affect host-microbe interactions among 37,214 tested. Here we show that one of these small molecules, RPW-24, protects C. elegans from bacterial infection by stimulating the host immune response of the nematode. Using transcriptome profiling, epistasis pathway analyses with C. elegans mutants, and an RNAi screen, we show that RPW-24 promotes resistance to Pseudomonas aeruginosa infection by inducing the transcription of a remarkably small number of C. elegans genes (∼1.3% of all genes) in a manner that partially depends on the evolutionarily-conserved p38 MAP kinase pathway and the transcription factor ATF-7. These data show that the immunostimulatory activity of RPW-24 is required for its efficacy and define a novel C. elegans-based strategy to identify compounds with activity against antibiotic-resistant bacterial pathogens.

Show MeSH
Related in: MedlinePlus