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Uracil-induced signaling pathways for DUOX-dependent gut immunity.

Lee KA, Kim B, You H, Lee WJ - Fly (Austin) (2015)

Bottom Line: We have recently found that uracil stimulates Hedgehog signaling, which in turn upregulates cadherin99C (Cad99C) expression in enterocytes.These observations illustrate the complexity of signaling cascades in uracil-induced signaling pathways.Moreover, we will provide a brief discussion on two different models for uracil recognition and uracil-induced DUOX activation in Drosophila enterocytes.

View Article: PubMed Central - PubMed

Affiliation: a School of Biological Science, Seoul National University and National Creative Research Initiative Center for Hologenomics, Seoul National University , Seoul , South Korea.

ABSTRACT
Intestinal dual oxidase (DUOX) activation is the first line of host defense against enteric infection in Drosophila. DUOX enzymatic activity is mainly controlled by phospholipase C-β (PLCβ)-dependent calcium mobilization, whereas DUOX gene expression is mainly controlled by the MEKK1-p38 mitogen-activated protein kinase pathway. Furthermore, bacterial-derived uracil molecules act as ligands for DUOX activation. However, our current understanding of uracil-induced signal transduction pathways remain incomplete. We have recently found that uracil stimulates Hedgehog signaling, which in turn upregulates cadherin99C (Cad99C) expression in enterocytes. Cad99C molecules, along with PLCβ and protein kinase C, induce the formation of signaling endosomes that facilitate intracellular calcium mobilization for DUOX activity. These observations illustrate the complexity of signaling cascades in uracil-induced signaling pathways. Here, we further demonstrated the role of lipid raft formation and calmodulin-dependent protein kinase-II on endosome formation and calcium mobilization, respectively. Moreover, we will provide a brief discussion on two different models for uracil recognition and uracil-induced DUOX activation in Drosophila enterocytes.

No MeSH data available.


Related in: MedlinePlus

CaMKII is a uracil-induced kinase required for host survival against enteric infection. (A) Identification of the protein kinase genes upregulated by uracil treatment. The details of mRNA-sequencing data were described previously.11 Color bar, gradient of log2-fold-changes at 2 or 16 h following uracil ingestion. Sixteen KD animals indicated by bold letters were subjected to gut infection experiments. (B) CaMKII is required for host survival against gut infection. The host survival rate following gut infection with E. carotovora is shown. A log rank analysis on the Kaplan-Meier data showed a significant difference in survival between control flies and CaMKII-KD flies. (C) CaMKII is induced following uracil ingestion. The expression levels of CaMKII were analyzed in anterior midguts obtained from adult flies following uracil ingestion (20 nM for 16 h). Target gene expression in the untreated control flies was taken arbitrarily as 1, and the results were shown as the relative levels of expression. Comparisons of 2 samples were made by Student's t-test; the values were expressed as the mean ± SEM (*P < 0.05) of at least 3 independent experiments.
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f0002: CaMKII is a uracil-induced kinase required for host survival against enteric infection. (A) Identification of the protein kinase genes upregulated by uracil treatment. The details of mRNA-sequencing data were described previously.11 Color bar, gradient of log2-fold-changes at 2 or 16 h following uracil ingestion. Sixteen KD animals indicated by bold letters were subjected to gut infection experiments. (B) CaMKII is required for host survival against gut infection. The host survival rate following gut infection with E. carotovora is shown. A log rank analysis on the Kaplan-Meier data showed a significant difference in survival between control flies and CaMKII-KD flies. (C) CaMKII is induced following uracil ingestion. The expression levels of CaMKII were analyzed in anterior midguts obtained from adult flies following uracil ingestion (20 nM for 16 h). Target gene expression in the untreated control flies was taken arbitrarily as 1, and the results were shown as the relative levels of expression. Comparisons of 2 samples were made by Student's t-test; the values were expressed as the mean ± SEM (*P < 0.05) of at least 3 independent experiments.

Mentions: To fill the knowledge gap in the uracil-induced intracellular signal transduction pathway, we focused on the protein kinases that play pivotal roles in ligand-induced intracellular signal transduction. Among the uracil-induced transcriptome, 35 kinases were found to be inducible (Fig. 2A). To see whether these kinases are involved in DUOX-dependent host defense, we examined the survival rates of knockdown (KD) animals for different uracil-induced kinases following pathogen infection because animals having impaired uracil-induced DUOX-activating ability (e.g., flies having impaired Hh signaling or impaired calcium signaling) show high rates of host lethality following enteric infection.11 Among 35 uracil-induced kinases, we used KD animals for 16 kinases that were subjected to enteric infection (Fig. 2A). Our mini-screening revealed that only one KD animal having reduced the expression of CaMKII was susceptible to gut infection (Fig. 2B), suggesting that CaMKII, along with previously identified PKCs, is the protein kinase involved in DUOX activation.11 By using quantitative PCR analyses, we could also confirm that uracil stimulation was able to induce the expression of CaMKII in the midgut (Fig. 2C).Figure 2.


Uracil-induced signaling pathways for DUOX-dependent gut immunity.

Lee KA, Kim B, You H, Lee WJ - Fly (Austin) (2015)

CaMKII is a uracil-induced kinase required for host survival against enteric infection. (A) Identification of the protein kinase genes upregulated by uracil treatment. The details of mRNA-sequencing data were described previously.11 Color bar, gradient of log2-fold-changes at 2 or 16 h following uracil ingestion. Sixteen KD animals indicated by bold letters were subjected to gut infection experiments. (B) CaMKII is required for host survival against gut infection. The host survival rate following gut infection with E. carotovora is shown. A log rank analysis on the Kaplan-Meier data showed a significant difference in survival between control flies and CaMKII-KD flies. (C) CaMKII is induced following uracil ingestion. The expression levels of CaMKII were analyzed in anterior midguts obtained from adult flies following uracil ingestion (20 nM for 16 h). Target gene expression in the untreated control flies was taken arbitrarily as 1, and the results were shown as the relative levels of expression. Comparisons of 2 samples were made by Student's t-test; the values were expressed as the mean ± SEM (*P < 0.05) of at least 3 independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4862428&req=5

f0002: CaMKII is a uracil-induced kinase required for host survival against enteric infection. (A) Identification of the protein kinase genes upregulated by uracil treatment. The details of mRNA-sequencing data were described previously.11 Color bar, gradient of log2-fold-changes at 2 or 16 h following uracil ingestion. Sixteen KD animals indicated by bold letters were subjected to gut infection experiments. (B) CaMKII is required for host survival against gut infection. The host survival rate following gut infection with E. carotovora is shown. A log rank analysis on the Kaplan-Meier data showed a significant difference in survival between control flies and CaMKII-KD flies. (C) CaMKII is induced following uracil ingestion. The expression levels of CaMKII were analyzed in anterior midguts obtained from adult flies following uracil ingestion (20 nM for 16 h). Target gene expression in the untreated control flies was taken arbitrarily as 1, and the results were shown as the relative levels of expression. Comparisons of 2 samples were made by Student's t-test; the values were expressed as the mean ± SEM (*P < 0.05) of at least 3 independent experiments.
Mentions: To fill the knowledge gap in the uracil-induced intracellular signal transduction pathway, we focused on the protein kinases that play pivotal roles in ligand-induced intracellular signal transduction. Among the uracil-induced transcriptome, 35 kinases were found to be inducible (Fig. 2A). To see whether these kinases are involved in DUOX-dependent host defense, we examined the survival rates of knockdown (KD) animals for different uracil-induced kinases following pathogen infection because animals having impaired uracil-induced DUOX-activating ability (e.g., flies having impaired Hh signaling or impaired calcium signaling) show high rates of host lethality following enteric infection.11 Among 35 uracil-induced kinases, we used KD animals for 16 kinases that were subjected to enteric infection (Fig. 2A). Our mini-screening revealed that only one KD animal having reduced the expression of CaMKII was susceptible to gut infection (Fig. 2B), suggesting that CaMKII, along with previously identified PKCs, is the protein kinase involved in DUOX activation.11 By using quantitative PCR analyses, we could also confirm that uracil stimulation was able to induce the expression of CaMKII in the midgut (Fig. 2C).Figure 2.

Bottom Line: We have recently found that uracil stimulates Hedgehog signaling, which in turn upregulates cadherin99C (Cad99C) expression in enterocytes.These observations illustrate the complexity of signaling cascades in uracil-induced signaling pathways.Moreover, we will provide a brief discussion on two different models for uracil recognition and uracil-induced DUOX activation in Drosophila enterocytes.

View Article: PubMed Central - PubMed

Affiliation: a School of Biological Science, Seoul National University and National Creative Research Initiative Center for Hologenomics, Seoul National University , Seoul , South Korea.

ABSTRACT
Intestinal dual oxidase (DUOX) activation is the first line of host defense against enteric infection in Drosophila. DUOX enzymatic activity is mainly controlled by phospholipase C-β (PLCβ)-dependent calcium mobilization, whereas DUOX gene expression is mainly controlled by the MEKK1-p38 mitogen-activated protein kinase pathway. Furthermore, bacterial-derived uracil molecules act as ligands for DUOX activation. However, our current understanding of uracil-induced signal transduction pathways remain incomplete. We have recently found that uracil stimulates Hedgehog signaling, which in turn upregulates cadherin99C (Cad99C) expression in enterocytes. Cad99C molecules, along with PLCβ and protein kinase C, induce the formation of signaling endosomes that facilitate intracellular calcium mobilization for DUOX activity. These observations illustrate the complexity of signaling cascades in uracil-induced signaling pathways. Here, we further demonstrated the role of lipid raft formation and calmodulin-dependent protein kinase-II on endosome formation and calcium mobilization, respectively. Moreover, we will provide a brief discussion on two different models for uracil recognition and uracil-induced DUOX activation in Drosophila enterocytes.

No MeSH data available.


Related in: MedlinePlus