Science Signaling Podcast: 15 January 2013

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Science Signaling  15 Jan 2013:
Vol. 6, Issue 258, pp. pc2
DOI: 10.1126/scisignal.2003894


This Podcast features an interview with Gabriel Núñez, senior author of a Research Resource published in the 15 January 2013 issue of Science Signaling. A group led by Núñez has performed a screen to identify regulators of the NOD2 and NF-κB signaling pathways. Signaling through NOD2 and NF-κB induces normal inflammatory responses, but mutations that alter the activity of NOD2 have been associated with inflammatory disorders such as Blau syndrome and Crohn's disease. The results of this screen help shed light on the genetic basis of inflammatory disease and offer potential targets for understanding and treating such disorders.

(Length: 16 min; file size: 9.4 MB; file format: mp3; location: http://podcasts.aaas.org/science_signaling/ScienceSignaling_130115.mp3)

Technical Details

Length: 16 min

File size: 9.4 MB

File Format: mp3

RSS Feed: http://stke.sciencemag.org/rss/podcast.xml

Listen to Podcast: http://podcasts.aaas.org/science_signaling/ScienceSignaling_130115.mp3

Educational Details

Learning Resource Type: Audio

Context: High school upper division 11-12, undergraduate lower division 13-14, undergraduate upper division 15-16, graduate, professional, general public and informal education

Intended Users: Teacher, learner

Intended Educational Use: Learn, teach

Discipline: Bioinformatics, genomics and proteomics; cell biology; genetics and heredity, human biology, immunology, molecular biology

Keywords: Science Signaling, Blau syndrome, Crohn's disease, inflammation, inflammatory disease, NF-kappaB, NOD2


Host – Annalisa VanHookWelcome to the Science Signaling Podcast for January 15th, 2013. I’m Annalisa VanHook, and today I’ll be speaking with Gabriel Núñez about a paper that appears in this week’s issue of Science Signaling on the identification of regulators of an inflammatory signaling pathway (1).

Inflammation is an important weapon in the immune system's arsenal of defenses against pathogens. The innate immune system triggers the inflammatory response when a tissue is damaged or infected, and this acute inflammation helps the body get rid of the pathogen, and it stimulates healing. Normally, the inflammatory response is terminated when the threat is resolved. But if the inflammatory response is not terminated—or if a tissue is constantly subjected to conditions that cause inflammation—chronic inflammation results, and this actually causes tissue damage rather than promote healing. Chronic inflammation can cause a wide range of diseases, from asthma and rheumatoid arthritis to atherosclerosis, inflammatory bowel disease, and even cancer.

This week's issue of Science Signaling focuses on the molecular signals that regulate inflammation. One of the articles in this issue describes a screen to identify regulators of the NOD2 and NF-κB signaling pathways. Signaling through NOD2 and NF-κB induces normal inflammatory responses, but mutations that alter the activity of NOD2 have been associated with inflammatory disorders such as Blau syndrome and Crohn's disease. A group led by Gabriel Núñez has performed a screen to identify regulators of this signaling network, and their results help shed light on the genetic basis of inflammatory diseases like Crohn's. Dr. Núñez spoke to me from the University of Michigan in Ann Arbor.

Interviewer – Annalisa VanHookBefore we address the work in this paper specifically, I want to talk a little bit about inflammation and disease in general. How does uncontrolled inflammation or prolonged inflammation cause disease?

Interviewee – Gabriel NúñezSo, many inflammatory molecules—they are made for controlling microbes, infectious microbes—also have a downside to them, [which] is that they can cause tissue damage. So, typically, under low levels of infection, there is not much damage in the tissues, but when there is overwhelming infection, [this] can lead to overproduction of certain molecules, like TNF-α, which can damage the tissues. So, in some diseases, some of these same pathways are aberrantly activated and they are overproducing molecules, like TNF-α, that can lead to local tissue damage. And this is the case, for example, of Crohn’s disease, a disease that we are going to be discussing later on today in more detail.

Interviewer – Annalisa VanHookIn this study, you were specifically looking for regulators of NOD2 and NF-κB signaling during inflammation. How does signaling through NOD2 and through NF-κB promote inflammation?

Interviewee – Gabriel NúñezSo, NOD2 acts as an intercellular receptor that is activated by muramyl dipeptide [(MDP)]. This is a moiety present in the cell wall of practically all bacteria. So, upon stimulation of NOD2, NOD2 activates NF-κB, which is a transcriptional factor that induces a large number of proinflammatory genes.

Interviewer – Annalisa VanHookAfter the inflammatory response has performed its function of helping to get rid of a pathogen or to stimulate healing, it has to be stopped or else it can cause tissue damage. How is inflammation that’s induced by NOD2 or NF-κB terminated?

Interviewee – Gabriel NúñezSo, there are two mechanisms that can occur. One of them is [that] NF-κB signaling itself can induce genes, which are functioning in a negative feedback loop. For example, in the case of NOD2, we found that two genes, A20 and also IκBα, which is a factor that sequesters NF-κB in the cytoplasm, are induced upon activation. So, these all act as negative regulators, so these affect in a negative manner the induction of inflammation. Another mechanism is that the ligand itself—the initial stimulus—this also can be reduced in amount. So, that also, of course, will have an impact in the strength of the signaling pathway itself.

Interviewer – Annalisa VanHookSo you mentioned Crohn’s disease earlier as a disease that can result from defects in inflammation. How does dysregulation of NF-κB signaling that’s induced by NOD2 cause Crohn’s disease?

Interviewee – Gabriel NúñezThis is a very complicated question, but this is a very important question. So, we know that mutations in NOD2 are loss-of-function mutations. This means that they make proteins, NOD2 proteins, which are impaired in the recognition of MDP, the microbial ligand which activates NOD2. So, we don’t fully understand how the impaired function of NOD2 mutations trigger Crohn’s disease. One possibility, which is supported by experimental evidence, is that NOD2 acts to prevent the local invasion in the intestine of some commensal bacteria. These are the bacteria that normally live in our guts. And in the presence of NOD2 loss-of-function mutations, these bacteria may accumulate locally and trigger secondarily the aberrant activation of increased NF-κB because of the other pathways which will be also triggered by this bacteria. So, under normal conditions, NOD2 will prevent this local invasion, so there will not be activation—abnormal activation. But in the presence of NOD2 mutations, in which NOD2 bacterial recognition is impaired, then this lead to increase of certain bacterial populations, which now will aberrantly activate the other signaling pathways which are involved in bacterial recognition, for example, like Toll-like receptors, which is another major pathway, which is activated by bacteria.

Interviewer – Annalisa VanHookAnd it’s that extra immune response in the gut that causes the symptoms of Crohn’s disease.

Interviewee – Gabriel NúñezYes. Because there’s more inflammation, it leads to local production of molecules there like TNF-α, which cause local tissue damage. And this is why many patients benefit from TNF blockade, because this is a molecule that is very important for microbial elimination but also—it's like a two-edged sword—also it can induce tissue damage.

Interviewer – Annalisa VanHookSo, in this study, you did a screen. What was the set-up for that screen you did? How did you identify genes that encode regulators of NOD2 and NF-κB signaling?

Interviewee – Gabriel NúñezSo, we first engineer[ed] a reporter cell line that, upon addition of the NOD2 agonist, muramyl dipeptide, induces the activation of NF-κB, which is revealed by a luminescence readout. So then, we inhibited over 18,000 genes using a methodology called siRNAi, which basically inhibit[s] the expression of genes. And then we basically could identify genes that promote or inhibit NOD2 signaling. So, in the case of positive regulator, the inhibition of the RNA will result in lower signaling, so that will say there is a positive regulator of NOD2. While in some cases, inhibition of the gene induces enhanced signaling, so we can say that is a negative regulator of NOD2 signaling.

Interviewer – Annalisa VanHookWhat sorts of proteins did you identify as regulating this network?

Interviewee – Gabriel NúñezSo, because it was a genome-wide screen, we identified many genes which are involved in different steps of NF-κB activation. For example, some of the genes were the transcriptional factor itself, NF-κB—for example, RelA—and also all the critical components for NOD2, specifically RIPK2. We also identified another set of genes which are important for proteasome activation, which is a very important pathway in regulating NF-κB. Other genes were genes which are important for nuclear pore components. So, many of the NF-κB components have to travel to the nucleus, so inhibiting some of these nuclear pore components will interfere with NF-κB pathway itself. There were other genes that were for regulatory components of NF-κB. And, so there were several types of genes which are involved in NF-κB activation that were revealed by this screen.

Interviewer – Annalisa VanHookDid you identify some factors that were previously unknown to regulate activation of NF-κB signaling by NOD2?

Interviewee – Gabriel NúñezYes. We identified some factors that were previously unknown—or they were known, but they were not previously implicated in the NF-κB signaling pathway.

Interviewer – Annalisa VanHookAnd what were some of those factors?

Interviewee – Gabriel NúñezWell, some of these factors—some of them we don’t understand how they function, so more work needs to be done. But I think this work provide now opportunities for many investigators around the world to follow up these leads which we identified. And so, I think this is one of the major factors in the work that this will allow now scientists to follow up some of these factors that we have found to be important for the NOD2 signaling, and they can be followed up by in future studies.

Interviewer – Annalisa VanHookHow do you think that your results can be used to help understand or to treat inflammatory diseases that are caused by dysregulation of the NOD2–NF-κB signaling axis?

Interviewee – Gabriel NúñezSo, one possibility is that the screen revealed many genes that we don’t quite understand the function. Importantly, some of these genes that we found to be important in the NOD2 signaling pathway have been linked to increase in susceptibility or protection from Crohn’s diseases by genome-wide association studies, but there was no really known function for these genes. So, some of the genes we identified involved in the NOD2 pathway [are] actually associated with Crohn’s disease. So, this data suggest that perhaps many patients which they have normal NOD2 alleles may have mutations in other genes in the NOD2 pathway, which have been revealed by our studies.

Interviewer – Annalisa VanHookCrohn’s disease specifically affects the intestine, but NOD2 signaling is important in immune system function throughout the body. So, are any of these factors that you identified as regulating NF-κB signaling downstream of NOD2 that are potentially important for Crohn’s disease, could the fact that those molecules might be used in other contexts complicate the possibility of maybe being able to target those molecules for Crohn’s disease therapies?

Interviewee – Gabriel NúñezYeah, this is a common problem with targeting inflammatory pathways. Typically, inflammatory pathways are there to prevent pathogen invasion. So always when you develop strategies to inhibit inflammatory pathways, you always have a chance to also interfere with microbial eradication, so you could have problems with increased infection or increased invasion of certain pathogens. And this is—it’s a balance that have to be follow[ed] to be effective to reduce inflammation but at the same time not overdoing this, that will be increase the chances for infection of these patients being treated. So, this has always been a problem in strategies to diminish tissue damage and diseases which are inflammatory. There has to be a balance because these pathways are there not to cause disease; they are there primarily to protect us from bacterial invasion and viral infection.

Interviewer – Annalisa VanHookHow about inflammatory diseases other than Crohn’s? Are there other inflammatory diseases in which NOD2 signaling has been implicated?

Interviewee – Gabriel NúñezThere have been another different type of mutations; they are actually gain-of-function mutations. So these are, they are syndromes caused by single–amino acid changes in the NOD2 protein, which cause increased activation of the NOD2 proteins. So, these NOD2 become constitutively active. So, the mutation is activated in the absence of the microbial ligand, so these patients have symptoms of rheumatoid syndromes like arthritis, problems with inflammation in the eyes, and other symptoms in the skin that mimic infection by a microbe but it’s actually, there’s no infection. It just reflects the fact that these mutations are constitutively active. So these patients are rare syndromes, typically children which affected by these, familial syndromes. They’re high penetrance, meaning that when you have the mutation, you will have the disease. [This is] very different from Crohn’s disease, in which environmental factors—namely the bacteria in the intestines—also play a very critical role. So, yes, this is the opposite type; it’s a gain-of-function mutation versus loss-of-function mutations in Crohn’s disease.

Interviewer – Annalisa VanHookThank you, Dr. Núñez, for speaking with me.

Interviewee – Gabriel NúñezYou are welcome.

Host – Annalisa VanHookThat was Gabriel Núñez, senior author of a paper that appears in the current issue of Science Signaling. That paper is by Warner and colleagues, and it’s titled “A Genome-Wide siRNA Screen Reveals Positive and Negative Regulators of the NOD2 and NF-κB Signaling Pathways” (1).

You can read that paper, as well as other articles on the molecular regulation of inflammation, in this week's issue of Science Signaling. This special Focus Issue also includes research on the role of a metalloprotease in skin inflammation (2), a Perspective on inflammation and multiple sclerosis (3), and an Editorial Guide by editor John Foley (4), who highlights these new articles and related articles from our archives.


And that wraps up this Science Signaling Podcast. If you have any questions or suggestions, please write to us at sciencesignalingeditors{at}aaas.org. This show is a production of Science Signaling and of AAAS—Advancing Science, Serving Society. I'm Annalisa VanHook, and on behalf of Science Signaling and its publisher, the American Association for the Advancement of Science, thanks for listening.


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