This week, I'm going to take a break from vaccines and do some innate immunity. Today's topic: the provocatively named "Inflammasome." This Nature Review from last month focused on inflammasomes and anti-viral immunity, but I think the inflammasome itself needs its own post.
A breakthrough in our understanding of the mechanisms that control the activation of inflammatory caspases came from the identification and characterization of the inflammasome, a large (~700 kDa) multiprotein complex that recruits inflammatory caspases and triggers their activation. Inflammasomes are often defined by their constituent PRR family member, which functions as a scaffold protein to bring caspase-1 molecules together and mediate proximity-induced auto-activation of caspase-1.
Note: the TL;DR version below
When immune cells like macrophages encounter bacteria and viruses they have a few typical responses, and one of the most important responses is the production of pro-inflammatory cytokines. Cytokines are signaling proteins of the immune system; they are secreted by cells to communicate information to other cells. Pro-inflammatory cytokines then, are cytokines that induce inflammation. There are a lot of pro-inflammatory cytokines produced by macrophages, with names like tumor necrosis factor (TNF), interleukin 1 and interleukin 6 (IL-1 and IL-6). If a macrophage sees a bacterium, it will make TNF, IL-1 and IL-6 (along with many others). If it sees viruses, it will make all the same cytokines (and some others). But in order to study how this works, scientists started to use purified ligands to make things easier. If I want to study how a particular receptor (like TLR4) works, I don't want to use whole bacteria - they have a ton of different molecules that might activate tons of different receptors, and I wouldn't be able to tell what TLR4 itself was doing.
So instead I use purified lipo-poly saccharide (LPS), which is a molecule that uniquely activates TLR4 (also called a (ligand"). But when scientists first started using these ligands, they noticed something weird. If they measured what the cells secreted, they found TNF and IL-6, but no IL-1. When they looked inside the cells, on the other hand, there was plenty of IL-1. So there was no problem with the production of IL-1, it just wasn't getting out of the cell.
This is where I need to talk about caspases. Caspases are a large family of proteases - which means they cut other proteins. Some proteases are fairly non-specific, they'll chew up lots of different proteins at lots of different places. Caspases are much more specific; they are very particular about what proteins they cut, and where those cuts are made. It turns out that when IL-1 is made, it's made in a immature form called pro-IL-1 that can't be secreted until it's cleaved in half by caspase-1. Once this happens, IL-1 can make its way out of the cell and do its thing. But there's always some caspase-1 in the cell, so why isn't pro-IL-1 cleaved immediately?
In general, caspases do very dangerous things. Inflammation triggered by IL-1 can be very destructive, so it needs to be kept in check (more on that later), and other caspases are involved in the signaling cascade that causes cell suicide (apoptosis). This means that caspases need to be tightly regulated, so caspases are first made in an inactive form: pro-caspase. Just like IL-1, in order to be fully active, a caspase must be cut by another protease (usually another caspase).
This is where the inflammasome comes in. It's just a fancy name given to a large complex of proteins whose function is to activate caspase-1 and allow inflammation. Inflammasome assembly is initiated by a pattern recognition receptor (PRR) binding to its ligand. This allows the PRR to recruit a protein called ASC, which then recruits a bunch of pro-caspase-1 molecules, forming a wheel-like structure (where caspase-1 forms the spokes). When a lot of caspase-1 is forced into close proximity by the inflammasome, they are able to cleave each other, activating the whole complex (as soon as one gets cleaved and activated, it will immediately cleave and activate all of its neighbors). Once the complex is activated, it can go on to cleave any pro-IL-1 in the cell, and allow it to be secreted.
[Adapted from doi:10.1038/nri2296]
TL;DR version: PRR assembles inflammasome --> inflammasome activates caspase-1 --> caspase-1 activates IL-1 --> IL-1 gets secreted.
You may be wondering, "why is any of this necessary?" It's hard to definitively answer "why" questions in biology, but one idea is the danger hypothesis. Our bodies are swarming with bacteria, fungi and viruses, but most of them won't end up doing any harm. The pattern-recognition receptors of the innate immune system can't distinguish between the good guys and the bad guys. The inflammasome is triggered by signals of danger (DNA in the cytoplasm and membrane damage for instance), so if a cell sees a bacteria, and there's some signal that something is wrong, you want to induce inflammation, but if there's only bacteria, but no danger, or there's danger but no pathogen, inflammation can be destructive.
The inflammasome was only discovered in 2002, so there's a lot we don't know, but it's a fascinating and complex field. Take a look at that first paragraph and see if it makes sense now. If it still doesn't, please let me know, I'm still working on how to explain complex topics like this. Too much detail? Too little?
Kanneganti, T. (2010). Central roles of NLRs and inflammasomes in viral infection Nature Reviews Immunology, 10 (10), 688-698 DOI: 10.1038/nri2851
I think the explanation was very good. One thing remaining open to me though, is why the LPS binding to TLR4 (a PRR) does not activate the Inflammasome, while you are saying: "Inflammasome assembly is initiated by a pattern recognition receptor (PRR) binding to its ligand."
Additionally as far as I know LPS does induce IL1 secretion - at least in experiments with whole blood or PBMCs.
Anyways, I think it was a good explanation!
Great post - and a really good explanation. I must admit I'd never heard of the inflammasome before - not sure whether it's a more recent idea or just something too specific to be addressed during my brief immunology course.
btw ... i'm currently looking for new posts for the MolBio carnival coming out on December 6th and I would love to include one of your posts, if you have no objections. You can find out more information, (and submit a post!), here: http://molbiocarnival.blogspot.com/
@tiffany - There are particular PRRs (AIM2 and NALP3 for instance) that activate the inflammasome, but not all PRRs do. TLR's definitely don't, at least not directly. Which leads to your second question (from the review):
The discovery that NLRP3 can detect a wide range of divergent PAMPs â such as bacterial LPS, peptidoglycan and bacterial or viral nucleic acids â as well as endogenous danger signals (DAMPs), such as monosodium urate (MSU), calcium pyrophosphate dihydrate and ATP, has led to the hypothesis that NLRP3 might sense a common downstream effect caused by these molecules, rather than the PAMPs or DAMPs themselves. Reactive oxygen species (ROS) that are generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase have been proposed as second messengers that activate the NLRP3 inflammasome
It sounds like some of the damaging things released by TLR activation could feed back and activate the inflammasome, but it's definitely not the LPS itself.
@Lab Rat - Inflammasome was described first in 2002, but it didn't start showing up in immunology textbooks until later. When I took immunology as an undergrad in 2004, we didn't hear about it either. I got an earful about it in my first year graduate immunology classes, but that's because Jurg Tschopp (the guy who discovered it) came and gave us a guest lecture.
As for the Carnival, I saw that announcement, but I don't know which of my posts really count as mol bio... I have some ideas though, I'll try to get back to you soon (last few weeks have just been crazy)
Hi Kevin -
Reactive oxygen species (ROS) that are generated by nicotinamide adenine dinucleotide phosphate (NADPH) oxidase have been proposed as second messengers that activate the NLRP3 inflammasome
Now that is a pretty interesting statement. I've hit the edge of my biology understanding, would this mean that any oxidative stress could participate in activation of NLRP3? Or only ROS generated specifically through NADPH? Can you help?
I've got some other thoughts on this post that I'm not able to articulate clearly yet, but this was a nice write up.
@pD - presumably, ROS from any source could activate the inflammasome, but at the same time, ROS doesn't always activate the inflammasome. There may be other levels of regulation involved, it might depend on the cell type, and it might be completely wrong. I don't want to pretend I'm an expert on this, so what follows is paraphrasing the review...
Inhibiting ROS in some cells inhibits inflammasome activation, and NALP3 has been shown to interact with a protein known to be a sensor for ROS. However, certain stimuli that activate the NALP3 inflammasome work just fine in cells lacking NADPH oxidase, and it's been shown that increased levels of ROS can actually inhibit the inflammasome.
However, most reports indicate that TLR signalling or ROS alone is insufficient for NLRP3 inflammasome activation and for the production of significant amounts of IL-1Î² and/or IL-18. So, a two-signal hypothesis for inflammasome activation has emerged in which production of pro-IL-1Î² and pro-IL-18 is regulated independently of caspase-1 activation; only when signals for both pro-IL-1Î² and pro-IL-18 production and caspase-1 activation are received are significant amounts of active IL-1Î² and IL-18 produced.
The literature for brand new fields is often contradictory and confusing. But as labs continue to study, and continue to argue their points, eventually a consensus is usually reached. Given another decade, I bet we'll have this thing figured out :-).
[wikipedia: List_of_prizes_for_evidence_of_the_paranormal (I removed the actual hyperlink, but the folks at wikipedia caught and expunged his edits almost immediately - KB)]
HOW NOSTRADAMUS WON ALL THE PARANORMAL PRIZES!
[wikipedia: Nostradamus (same deal)]
THE HIGH PRICE OF REVOLUTION
[youtube link deleted - KB]
Hi Kevin - was reading at the Digital Cuttlefish's and wanted to congratulate you on your first Mabussing (Mabus-ing? I'm not sure how one writes it).
Oh, and I'm enjoying your blog; very glad you joined SB. I love the stuff on mucosal immunity, oral vaccines and VLPs, very cool.
Hey look, I just got another one! Glad you're enjoying reading. Any and all feedback is welcome.
Immunology is getting really strange!
The explanation was good.
lightbulb went on, Thanks Kevin