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Non-apoptotic role of BID in inflammation and innate immunity

Abstract

Innate immunity is a fundamental defence response that depends on evolutionarily conserved pattern recognition receptors for sensing infections or danger signals1,2. Nucleotide-binding and oligomerization domain (NOD) proteins are cytosolic pattern-recognition receptors of paramount importance in the intestine, and their dysregulation is associated with inflammatory bowel disease3,4. They sense peptidoglycans from commensal microorganisms and pathogens and coordinate signalling events that culminate in the induction of inflammation and anti-microbial responses2. However, the signalling mechanisms involved in this process are not fully understood. Here, using genome-wide RNA interference, we identify candidate genes that modulate the NOD1 inflammatory response in intestinal epithelial cells. Our results reveal a significant crosstalk between innate immunity and apoptosis and identify BID, a BCL2 family protein, as a critical component of the inflammatory response. Colonocytes depleted of BID or macrophages from Bid−/− mice are markedly defective in cytokine production in response to NOD activation. Furthermore, Bid−/− mice are unresponsive to local or systemic exposure to NOD agonists or their protective effect in experimental colitis. Mechanistically, BID interacts with NOD1, NOD2 and the IκB kinase (IKK) complex, impacting NF-κB and extracellular signal-regulated kinase (ERK) signalling. Our results define a novel role of BID in inflammation and immunity independent of its apoptotic function, furthering the mounting evidence of evolutionary conservation between the mechanisms of apoptosis and immunity.

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Figure 1: Genome-wide RNA interference screen for genes regulating NOD1 signalling.
Figure 2: BID is required for NOD1 and NOD2 signalling.
Figure 3: BID regulates NOD signalling independently of its apoptotic determinants by linking NOD proteins to the IKK complex.
Figure 4: Bid −/− mice exhibit a blunted inflammatory response after NOD stimulation in vivo and are not protected from DSS colitis by MDP.

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Acknowledgements

We thank A. Strasser for providing Bid−/− mice, G. Shore for BID antibodies and the McGill University high throughput/high content screening facility. We also thank D. Zhai for BID purification. This work was supported by grants from the Canadian Institutes for Health Research (CIHR-MOP 82801) and the Burroughs Wellcome Fund to M.S. M.S. is a Canadian Institutes for Health Research New Investigator. G.Y. is supported by a PDF-Fellowship from the McGill University Health Center. C.P.D. is supported by a fellowship grant from the SASS Foundation for Medical Research.

Author information

Authors and Affiliations

Authors

Contributions

G.Y. and M.S. designed the research. G.Y., K.D. and M.S. performed the screen. G.Y. and M.S. analysed the data. G.Y. performed most experiments. R.G.C. performed Ni/NTA pull-down assay; P.F., R.G.C., C.P.D., D.R.G. and J.C.R. contributed new reagents/analytical tools. G.Y. and M.S. wrote the paper.

Corresponding author

Correspondence to Maya Saleh.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Figures

The file contains Supplementary Figures 1-5 with legends. Supplementary Tables 1-13 were omitted when this paper first appeared online, but added on 18 May 2011. (PDF 334 kb)

Supplementary Table 1

This table shows the 225 positive regulators of NOD1 signaling identified in the NOD1 primary screen. The genes are arranged in alphabetical order. The siRNA sequences are shown in columns F and G. The Z-scores are listed in column H. (XLS 80 kb)

Supplementary Table 2

This table shows the 198 negative regulators of NOD1 signaling identified in the NOD1 primary screen. The genes are arranged in alphabetical order. The siRNA sequences are shown in columns F and G. The Z-scores are listed in column H. (XLS 71 kb)

Supplementary Table 3

This table shows the gene list comparison between “Screen 1” and “the validation screen” and identifies 200 common genes. The genes are arranged in alphabetical order. The Z-scores are listed in columns F and G. (XLS 69 kb)

Supplementary Table 4

This table shows the gene list comparison between “Screen 1” and “the TNF secondary screen” and identifies 114 common genes. The genes are arranged in alphabetical order. The Z-scores are listed in columns F and G. (XLS 51 kb)

Supplementary Table 5.

This table shows the gene list comparison between “the validation screen” and “the TNF secondary screen” identifies and 60 common genes. The genes are arranged in alphabetical order. The Z-scores are listed in columns F and G. (XLS 41 kb)

Supplementary Table 6

This table shows the classification of the NOD1 siRNA screen hits into biological processes. Genes identified from the NOD1 siRNA primary screen were classified into biological processes using the PANTHER classification system and according to GO ontology. The biological processes of positive regulators of NOD1 signaling are presented. Genes with unassigned annotations are included in the tables as ‘unclassified’. (XLS 103 kb)

Supplementary Table 7

This table shows the classification of the NOD1 siRNA screen hits into biological processes. Genes identified from the NOD1 siRNA primary screen were classified into biological processes using the PANTHER classification system and according to GO ontology. The biological processes of negative regulators of NOD1 signaling are presented. Genes with unassigned annotations are included in the tables as ‘unclassified’. (XLS 90 kb)

Supplementary Table 8

This table shows the classification of the NOD1 siRNA screen hits into molecular functions. Genes identified from the NOD1 siRNA primary screen were classified into molecular functions using the PANTHER classification system and according to GO ontology. The molecular functions of positive (regulators of NOD1 signaling are presented. Genes with unassigned annotations are included in the tables as ‘unclassified’. (XLS 71 kb)

Supplementary Table 9

This table shows the classification of the NOD1 siRNA screen hits into molecular functions. Genes identified from the NOD1 siRNA primary screen were classified into molecular functions using the PANTHER classification system and according to GO ontology. The molecular functions of negative (regulators of NOD1 signaling are presented. Genes with unassigned annotations are included in the tables as ‘unclassified’. (XLS 63 kb)

Supplementary Table 10

This table shows the validated ‘hits’ identified as enriched in immune tissues/cells. Genes that show expression enrichment in immune cells and tissues determined by Wilcoxon rank-sum test (p<0.05) are presented. (XLS 23 kb)

Supplementary Table 11

This table shows the validated ‘hits’ identified as enriched in neuronal tissues. Genes that show expression enrichment in neuronal tissues determined by Wilcoxon rank-sum test (p<0.05) are presented. (XLS 20 kb)

Supplementary Table 12

In this table the 80 tissues in the Novartis GNF1H microarray dataset are listed. Tissues clustered in duplicate are ordered from left to right on the heat map. (XLS 28 kb)

Supplementary Table 13

This table shows a list of the primers used for cloning, mutagenesis and quantitative real-time PCR experiments. (XLS 26 kb)

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Yeretssian, G., Correa, R., Doiron, K. et al. Non-apoptotic role of BID in inflammation and innate immunity. Nature 474, 96–99 (2011). https://doi.org/10.1038/nature09982

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