Scientist Stories: Roy Parker, RNA & Abnormalities in Human Diseases

Morning. I do that every morning when I teach class too, and I stand there and wait for the response. So y'all were really good to come back quickly. This morning we're gonna have a session to start on tau and tangles, and it's my great pleasure to introduce first Roy Parker, who is a distinguished professor in biochemistry at CU Boulder and also happens to have the office right next door to mine. And he's going to be talking about the role of RNA in these aggregations and how they contribute to disease and what we can do about those.

Thank you. It's really great to be here. And I want to echo what Larry was saying about seeing people in person while you talk is so much more pleasing than just talking to a screen. So I want to thank— Larry and Meredith and Larry for inviting me. I've come to this symposium many times over the years. And it's really a privilege to be able to talk at it. What I want to do today is actually show my slides. OK, terrific. For those of you who don't know me, I'm historically an RNA biologist and have studied many different aspects of how RNAs are made and used in our cells. And I want to tell you two stories about that today that connect to disease. One, I want to talk about how some RNAs are degraded and how that allows us to have insights into how we might treat some various stem cell diseases. And then in the second part of my talk, I'm going to talk about RNA and its connection to neurodegenerative disease. This is a new area for me in my lab, but I think it's so important to all of us that we should probably start talking about it because Talking about science is how you develop ideas and how you get new feedback from the audience and pushes you in new directions. So I think that's an important part of science. All right, so to get started then, we're all familiar with the idea that you take DNA and you copy that into mRNAs and that mRNA is used to translate to make proteins. And how much mRNA you make then is really important because if you make more mRNA in the cell, you'll make more protein, and that changes the regulatory state of the cell. But the other part of that, of course, is that if you change the rate at which RNAs are destroyed, you'll have more RNAs at steady state, and you'll make more protein. So an important part of regulating the physiology of our cells is controlling the rate at which RNAs get degraded. And I'm going to summarize 100 person years in my lab in one slide. And the details of this slide don't matter other than the fact that a lot of really terrific people contributed to this. Over the years, my lab and other labs worked out the manners by which RNAs are degraded in human cells and how our cells control that process. And there's lots of interesting stories there that I'm not going to talk about today. But one of the things that we learned that I thought was interesting is it allowed us to understand some human diseases because of the principles we learned from studying RNA degradation. And one of the principles that became very important was the idea that cells clean up after themselves and they degrade defective RNAs. And so imagine you have a healthy RNA. It goes about its function. There's some kind of way to degrade it, but that's slower than function, so nothing happens. But if you have a defective RNA that slows down the function of that molecule, then that competing destruction pathway is now faster, and the RNA is destroyed, and it got rid of this defective molecule. And the analogy I often use to think about this, for those of you who aren't molecular biologists, this is like being on the freeway and your car breaks down. And our society wants to deal with that. We want to get that car out of there. So we call a tow truck and it removes it to let traffic keep going, to let the function keep occurring. Now let's imagine, you know, sometimes when your car breaks down, you wanna fix it. You get a flat tire, if you have a few minutes, you can change that tire and be back on the road again. But consider if you lived in a city that had unlimited and predatory tow trucks. So your car breaks, you get a flat tire, And before you can even get the spare out of the trunk, a tow truck has arrived, hooked you up, towed it away, taken it to the car crusher, and your car is destroyed. So that leads to destruction of potentially a functional machine. And this is exactly what happens in a class of human diseases. And so these are what we call hyperdegradation diseases, where instead of the normal binding of RNAs and proteins together to make functional machines, what happens is that gets slowed down. The cell could still deal with that if it had enough time, but the competing RNA degradation now degrades the RNA, and now there's no chance for that RNA to function. And this leads to the loss of RNAs, and that can lead to some human diseases, such as spinal muscular atrophy falls in this family. But the one I want to talk about is a disease called dyskeratosis congenita. This is a disease where your telomerase RNA actually gets degraded too fast. And this is important because telomerase— this is a complex in all of our cells, well, particularly in all of our stem cells, which adds ends to our DNA molecules when they divide, when they get replicated. And this is important because it solves the end replication problem of DNA. Replication. And it's important in cells that divide frequently. So this is like your bone marrow, your stem cells, your epithelial cells in your lung. And so people who have defects in telomerase, the most critical thing that happens, they end up with bone marrow failure. They run out— their stem cells in their bone marrow run out of the telomeres, and they just can't make enough cells anymore. They also have skin disease and other issues. And there's no way to treat this. I'm going to turn on my timer because I don't think the timer up here is working. And I have no idea how long I'm going to be talking. Maybe I have unlimited time. OK, so we got interested in this. And Sid Shukla, who was a PhD student in my lab a few years ago, had the idea, OK, if we understand how the telomerase RNA is degraded, we could block that degradation, and then we could treat the disease. And this was a collaboration with Jens Schmidt in Tom Cech's lab. Who's across the hall from me. And Sid basically set out to ask this question, what happens to the RNA in these individuals? And to make a long story short, what he learned is that, again, there's these competing pathways. In normal people, these proteins bind— oop, I don't have a pointer, do I? But in normal people, these proteins bind to the end of the RNA, and they protect it. From this degradation pathway. But what happens in people who have mutations, let's say in one of these particular proteins called dyskerin, now the protein can't bind. And this enzyme comes in called PAPD5 and adds a tag. And that tag marks the RNA for destruction. And so again, to draw an analogy, you can think about this just like someone going through the woods, a logger. Tagging trees for the loggers to cut them down. And then once that tag is there, it recruits a degradation machine. The RNA is degraded. And you get bone marrow failure and skin conditions and also some pulmonary fibrosis often occurs. All right, so what can we do about that? Well, it turned out that many companies had already made drugs which block this enzyme. And these are because this enzyme is part of hepatitis B's lifestyle. So several biotech companies had made inhibitors of this compound. And so we simply said, well, can we take these preexisting drugs and rescue the production of the RNA and therefore rescue these stem cell problems? And that actually works quite well. We've worked with Luis Batista at Washington University in St. Louis. Luis actually is an expert in blood differentiation and culture. And we can take normal human embryo stem cells, differentiate them into the plethora of different kind of blood cells that our bodies make. But if you have a mutation which causes dyskeratosis congenita and causes loss of the telomerase RNA, that no longer works. The cells poop out. And then if we add this drug, we can then rescue this. And so this is really pleasing. And then also another group recently has shown that you can do a similar thing in a mouse model of this disease. So we've gone from trying to understand how RNA degradation works to what we think is an approach ready to go into people. And there's a couple biotech companies now that are trying to start that process. All right, so my lab continues to work on diseases like this. We have a similar story with a neutropenia type of disease, and we're also studying the opposite situation occurs in our cells where RNAs last too long. And because of that, that leads to disease, which drives some forms of muscular dystrophy and ALS. If anybody's interested, we can talk more about that later. All right, so a few years ago then, I had a midlife crisis, probably because I'm getting older. And I decided I should start working on neurodegenerative disease. And there's really three reasons. But not because I forgot anything yet. But this is a big deal, and I'll talk about why. It's also, to me, a really interesting problem because we don't understand the biology. And if we don't understand the biology, we can't really end up with treatments. And there's also a really interesting connection to RNA, which I'll tell you about, that I think is important in the biology and may lead us to insights into how we might to treat this. All right, so this is a big problem. James talked about this a little bit yesterday. Alzheimer's alone, which is about half the dementia cases in our country, is the 6th leading cause of death. And so if you double that, there's really about 12 million cases a year from neurodegeneration disease. It's extremely expensive to treat because these patients actually often need supportive care for a long time. And it's also emotionally devastating for any of you who've had family members or been caregivers in this situation. It's a really hard thing to deal with these diseases. So this is a big problem, and it's something that we really need as a society to find solutions to because we're getting better at cancer and heart disease. And so we're pushing people towards neurodegenerative diseases as we all age. Our society is getting older. This is a demographic from 2050 where 12.5% of our population will be over 75 years old. And so we need to start figuring out solutions to this. Now I'm going to tell you what I think the crux of the issues are here. And these are my opinions. And I want to acknowledge that neurodegenerative diseases are awfully complex. And I'm going to simplify that, and I apologize that, but I think to try to get across the key points one needs to do that. It involves neuroinflammation, the immune system, different cell types in the brain, neurons, microglia. Your cardiovascular health plays a role in your susceptibility to neurodegenerative diseases, and many genetic and environmental inputs. So keep that caveat in mind that it really is a complicated problem. So the first key point is that neurons die. And that all these neurodegenerative diseases leads to neurons to dying. We generally don't make new neurons in our brain. So enough neurons die and you lose function. It's really that straightforward. So what we have to do is figure out why neurons die and figure out ways to keep them around. It looks like neurons die largely because of the buildup of aberrant protein aggregates. And so these are cases where you have proteins which are forming clumps in cells. And then that leads to those cells dying, particularly neurons. And that leads to pathology. And I want to point out that there's an idea in the field that different diseases are primarily driven by different protein aggregates, such as Alzheimer's is driven by A-beta and a protein tau that we'll talk about today. Parkinson's being driven more by alpha-synuclein. And I think one of the caveats we all have to keep in mind is that really there's often copathology. And it's not one protein in one disease. People who have alpha-synuclein and get Parkinson's often have tau aggregates as well. So it's actually a complicated interlapping network of these different interactions. Because these proteins form these aberrant clusters and clumps, you can think of these as a class of what people call protein folding diseases. We all are familiar with the idea of proteins folding up into their proper structure. We're going to hear a talk about how we can predict that later today. And what happens in neurodegenerative disease is that those proteins don't form their normal structure. They form an aberrant structure, much like this chair at Robbins which she tried to build from IKEA. But it's worse, right? It's actually much worse than this because these misfolded proteins don't just misfold. They spread. And this is by what people call a prion mechanism. So imagine you have a whole bunch of proteins which are in their normal shape. One of them forms an aberrant structure. And that aberrant structure then interacts with normal proteins and converts them into the aberrant structure and then propagates. And so you end up propagating and propagating until you end up with these fibers or aggregates, which then cause neurons to die. You can think of this very much like mad cow disease. Mad cow is also a prion where you ingest a prion seed, and then that propagates in your brain and causes neurons to die. This is a similar phenomenon, but instead of you eating the seed, the seed occurs stochastically in our brains at some rate. An analogy here is that similar kinds of things can happen in human cultures. A politician can enter an abnormal state. If that triggers other politicians to enter that abnormal state, you can propagate toxic ideas in our cultures as well. [LAUGHTER] So yet there's one more layer here. That prions can spread between neurons. So it's not just that this occurs within a cell and that it starts propagating aberrant structures. It's that when that neuron dies, it releases those seeds, and those seeds can be taken up by other neurons and start the whole process over again. And so it's really this prion spreading or prion-like spreading between neurons that leads to the degenerative process that we see in neurodegenerative diseases. And if you look at all these different proteins, which play a role in neurodegenerative disease and others as well that aren't on here, they all spread— can be demonstrated to spread by this prion-like mechanism where they form a structure, convert other molecules into that structure, and then spread between cells. Now, there's lots of ideas. And I think one of the exciting things right now in the biotech arena is lots of different efforts to try to intervene with these prion-like spread phenomena. You can try to reduce the amount of the protein, try to figure out ways to get rid of these aggregates when they form in cells, target their toxicity, sequester the extracellular forms with antibodies so they can't be taken up by other neurons, block the uptake of those seeds by understanding their receptors. And I think Dave is going to talk about some ideas about how you might even disassemble these structures when they form. I just want to mention one that I find particularly innovative. This is an approach called genus therapy. It's driven by a woman, Dr. Tsai at MIT. This is a really interesting observation she made that if you use light or sound to induce gamma oscillations in your brain— these are 40-hertz oscillations roughly— you can actually reduce protein aggregates. This works in mice. They just did a small phase 2 trial and saw some benefit in people with mild cognitive impairment. That's actually a light on my nightstand that cycles at 40 hertz. So it can't hurt at this stage. And I have some familial connections to these kinds of diseases. Anyway, but if you're interested, there's a really interesting set of papers on this that I recommend you look at. Where to get it? Yeah. All right, now there's lots of different proteins that do this. The big dog in the room is tau. Tau forms aggregates in at least 25 different neurodegenerative diseases. If you take all the cases of dementia and you separate them not by what they're called, Alzheimer's versus PSP, but you separate them by what protein is the predominant aggregate that forms, 80% of the cases are driven by tau. And so this occurs in things like CTE, the repetitive head trauma that athletes get. That's really a tau-driven pathology. It occurs in some infections. 1 in 500 or 1,000 people who get measles develops a tauopathy about 10 years later. If you know people who don't get their kids vaccinated because of measles because they say it's not a big deal, tell them there's a 1 in 1,000 chance that your kid 10 years from now is going to come down with a fatal tauopathy. And then Alzheimer's disease is also driven by tau. We heard James talk about the A-beta hypothesis and A-beta plaques. And I think the best understanding now really is that A-beta plaques are important in Alzheimer's, but what they do is they contribute to tau getting initiated down this pathway of forming aggregates and propagating and spreading. And that tau, at least I think the best current hypothesis, is really what leads to neuronal death. Let's hope that we're right on this one. So that actually leads to a key question. How do beta amyloid plaques promote tau aggregates? And we have some ideas about that. All right, so my lab has decided to get interested in studying tau. And there's a lot of really important questions. What really are these aggregates? How and where do they form in cells? How do cells control that process or prevent it? And what's the basis of toxicity? We still don't understand why neurons die in these cases. And then there's also clinical questions. If you can actually understand that, can you reduce the toxicity, reduce their formation, or reduce their spread? And we got started in this really because I study RNA biology. Why am I doing this? And it's always intrigued me that these proteins that aggregate, many of them are really good RNA binding proteins. TDP-43, which forms aggregates in about 90% of people with ALS, is a terrific RNA binding protein. And tau is actually a really good RNA binding protein, although most people don't think of it that way. So we got interested in this and decided to try to do some experiments. And this is really driven by a— a terrific MD-PhD student, Evan Lester, who's now back in the clinic. And he actually asked the question, really, are there RNAs in these tau aggregates? And could they play some role in what these assemblies are? And he did this because another part of my lab works at the formation of RNA aggregates in cells. RNA, like protein, can form aberrant structures in cells. And we thought there might be some overlap between those two kinds of— ideas. So how does he do this, right? Because as much as we'd like to study the interior of the brains of our colleagues, we're not able to do that. So I just want to talk about 3 different models that we use to understand some of the data. And the first thing that you do is you use mouse models. And these are mice which are either normal and don't make tau aggregates and stay fine, Or you have mouse models, which are genetically engineered, often because of a mutation in tau itself that causes pathology, that they develop tau aggregates. And so you can study this in the mouse. And then you can also then, because these behave like prions, we can then take those mouse brains and transfer an extract from them into cell line models in the lab. And these are cell lines which express tau. And if you do that, you can see that an extract from a normal mouse brain doesn't do anything. But if you take a brain extract from a mouse that has a tau pathology, it will start the whole process over again, and you'll start getting these tau aggregates. And then of course, mice and cells are not people. So the third thing that we have to do is always go back into people. And we do this typically by looking at postmortem tissues from patients. And here we've been helped tremendously by Stan Prusner at UCSF and Nadine Bacher at the Barrow Neurological Institute. They both have pretty big access to brain banks where we can then take samples and look at them to see if what we learned actually is true in humans. All right, so Evan learned, I think, two important things. One is that tau aggregates actually have a lot of RNA in them. So this is just a picture showing a tau aggregate that he purified from a mouse brain. And if you stain it in green for tau and in red for RNA, you can see that whole complex is full of RNA. For the aficionados in the room, the primary RNAs in here are snRNAs and snoRNAs, RNAs involved in RNA processing typically in the nucleus. He also found a lot of RNA binding proteins that were in there. And in particular, one I want to highlight today just some interesting observations is this protein SRM2. You can see that's the red protein in the middle there. And typically, it's found in the nucleus. This controls the splicing of RNAs in our cells. And in the case of when you get tau aggregates, you can see that that protein moves and overlaps with the green tau aggregates in the cytoplasm. Now, again, is this true in people? So we go back. Because now we found this protein that we think is really important in tau pathology for reasons I'll tell you about in a second. But is this occurring in people as well? This is actually taking a sample from a patient who had cortical basal degeneration. It's a form of tauopathy. And you can see in the top part that the SRM2 protein, which is red, overlaps with the blue, which is the nucleus of the cell.. And that's where it's supposed to be. That's a normal person. But in the bottom, you can see those green tau fibers are forming. And what you see is that the red protein leaves the nucleus, the blue area, and overlaps with the tau protein. And so we see the same thing in Alzheimer's brains and also some other forms of tauopathies. So this occurs in humans as well as in our model systems. And that tells us it's worth studying because if it doesn't occur patients, then we're not so excited about it. So why do we care? I think we care about this for two reasons. One is the idea that this mislocalization of a protein might explain part of the toxicity. And we know this protein regulates a process called splicing in the brain. And it needs to be in the nucleus to do that. And when it moves out to the cytoplasm to interact with these tau aggregates, It causes splicing changes. And if you actually do RNA sequencing on patients that have had tau pathologies, you can find lots of changes in splicing. And whether that's really relevant to the disease, I think, requires more work, but it's an interesting correlation. The other reason we care about it is we think it's going to tell us how tau aggregates form in cells. And I'm going to put forth a specific hypothesis because we know the answer. But the hypothesis is going to be, could tau aggregates grow off a certain type of what's called an RNA protein granule, which is going to be a little blob in the cell containing RNAs and proteins that contain this SRM2 protein? And this is important because if we know where and how tau aggregates grow in cells, maybe we could inhibit that process and change their toxicity. All right, so to look at this, what we're going to do is I'm going to show you a movie. And in this movie, I'm going to show you the green is the tau protein. And when it forms the protein aggregates, it turns green. We've engineered to only do it when it aggregates. And the red is this SRRM2 protein. And I'm going to play this movie. And what you're going to see is normally the red protein's in the nucleus. Once in a while, it moves out to the cytoplasm, forms that structure. And then look, the green stuff grows off the red assembly. So I'm going to show that again. So this is what we think that is in cartoon form. So on the top there is the cartoon of this structure in the cytoplasm containing the SRM2 protein. We think this is related to a structure called mitotic interchromatin granule, a very unstudied structure. There's only 2 papers of it in the whole universe that we can find. Which is both good and bad. And that we think that's what's seeding these. And I'll show you the movie again so you can kind of see it. But there, things are going along. The red thing forms. And then the green tau aggregates grow off of that. This is not a rare occurrence. We observe this in about 50% of the cells that we see in this movie that make tau aggregates. The other 50%, the tau aggregate forms. We don't know if there's a small seed there or if it's forming by a different manner. And then the cell dies here, as you see. It's not a very pretty sight. All right, so there's two big questions we're trying to answer now in my lab. The first is, does this have anything to do with disease? Because that's in a cell line model. But we're really interested because of, I think, a couple different observations. First, there's an old paper— old meaning 4 years ago, which from a group in Japan who showed that in postmortem neurons from Alzheimer's disease patients, they see these cytosolic SRM2 blobs, assemblies. And they didn't look at tau because they didn't have any reason to. But that suggests their interpretation was that something about A-beta was causing this movement of SRM2 to these cytoplasmic assemblies. The other thing that we've looked at, Swathi Reddy, a student in the lab, she got interested in this idea of how the relationship between neuroinflammation and tau, because neuroinflammation clearly can contribute to the formation of these tau fibers. And so she actually has been looking in neurons that we derive from iPSC cells. And you can see that a normal neuron on the left, the protein is largely in the nucleus where it's supposed to be.. If we add this prostaglandin PJD2, this is a prostaglandin which can reach 10 micromolar during severe neuroinflammation events. You can see that the protein moves out to the cytoplasm and forms these small foci. And in some case, some cells, it's really extreme like the one on the right. So this suggests just there's some connection of neuroinflammation to these potential assemblies in the cytosol. And maybe they play a role in that process of tau aggregation. So the other big question that my lab's working on now is what's really going on? What is this biochemical property of these structures? What's relevant about that assembly that forms? Can we alter that to change tau aggregation, either to limit it? What's really the role of RNA in those aggregates? And can we, from this understanding, learn anything about why neurons die? And I realize we haven't solved the whole problem here. But this is where we've gotten in a few years, and we hope to make progress here. All right, so I just want to thank you all for listening. I have no idea what time we're at. But I also want to thank my lab. My lab is a terrific group of people, and they're really the ones who do all the work. And this is our building, the JSCB building, which is a terrific place with the Biochemistry Institute, the BioFrontiers Institute, and the Biochemistry Department. And also the funding from NIH and Howard Hughes. Thank you.

All right. I'm going to start with a question from online. It simply says, what is the delay?

What is the delay?

Is that an online, or is he talking about you?

I'll interpret that in a way that I can answer. Exactly. So a really important issue is the delay. Why do these diseases take so long? Because they do take a long time. From stuff I didn't talk about, people think that A-beta plaques start forming 20 to 30 years before you get symptoms. And tau can form several years before you start getting the first symptom, which is usually mild cognitive inhibition. And so why does it take so long to go from the initial events to the phenotype? And I think it's two things. I think one, this is a slow process by which one neuron dies and another, then another, then another, and it spreads. And so if you lose a neuron here or there, that's not so bad. Many people have strokes. That kills neurons, but you don't end up with a neurodegenerative disease from a stroke. You end up with loss of local function, and then you're done. So I think the delay is this progressive spreading. And I also think that our brains are really overwired. We can lose some neurons and still be pretty functional, but there's a point where once you start to lose enough neurons, you lose function. And I think that's what— it's the combination of the spreading and also the robustness of our brain and how many neurons we have to lose before we get a phenotype. Questions? Being the precocious individual I am, at 51, I already have fairly significant cognitive impairment associated with post-acute COVID-19 syndrome, a.k.a. long COVID. So unfortunately, I did not understand a word you just said. But my question is, where do I get one of those gamma wave lights? So you can buy them on Amazon. And I would recommend you look at the— there's a bioRxiv preprint of their small phase 2 trial. There's a phase 3 trial that's in process now. I think the best improvement— is actually people who have both the light and the sound stimulation because they do a 40-hertz sound and a 40-hertz light. And if you're exposed to both of those, both the animals and the people showed the biggest improvement. So if you're worried, get on Amazon. I've been doing this for a couple of months. I don't think I've changed. Yeah, I mean, and I would pay attention to it because this is a really innovative way of thinking about the problem. And if it makes a difference, it could be a game changer. But it could also fall flat on its face too at this stage. Yeah, Gene. Roy, thanks for the great talk. With respect to that lovely film that you showed, in other domains of biology, that activation would be associated with a sensor or a pioneer factor. So what do you think is going on there? What is this aggregate trying to do? Yeah, so that aggregate we think formed because of imbalances in nucleocytoplasmic transport. But the tau starts growing off it. It's not a new event of tau. We've actually provided a seed from outside there. So it's not that this starts new events, it just allows them to grow much, much faster. So I don't know if that answers your question, but we don't actually think that that structure has a role during mitosis when the nuclear envelope breaks down. But it's not really important. It just happens once in a while in a non-mitotic cell. It's kind of an accident, we think. Yeah, anybody?

Go ahead, John. Oh, okay. I really like the interesting thing with the snoRNAs and small RNAs, and they're super abundant. And I'm wondering, have you thought, could there be some sort of clock in this delay of these RNAs? I know they're post-mitotic, but they still could be being made and processed do they go out to the cytoplasm and sort of seed things? Or do you really think it's like how your model is where they grab them from the nucleus?

Because they're not normally out in the cytoplasm. Yeah, I think that— I don't think we have a good understanding of how those RNAs end up in those structures. And you have to remember, most of this is occurring in a neuron. And neurons are post-mitotic cells. So they don't— many of our cells will divide. The nucleus breaks down. And things in the nucleus can access the cytosol. But neurons don't do that. They're post-mitotic. So how RNAs get out of the nucleus into these aggregates in the cytosol is not clear to me. I think it could be a leakage event. And there's some evidence that— as neurons start to get tired, they start to get damage to their nuclear envelope and they start to leak. I don't think— I don't know. That could be a clock. That could be a clock, right. But I think— and to be honest, I'm very new to this field. I think, but I don't know, that if you look over time, it's not that it takes 30 years to start losing neurons. It takes 30 years to lose enough neurons to have a phenotype. So I think it's a neuron loss phenomenon.

We had a question on the other side. Yes, hi. It's a very interesting story. So as you know, tau, it's mostly the microtubule binding proteins and certain areas within that that seems to be like the most involved in the aggregates. I was wondering if there's any modeling that suggests that this RNA interact with this particular sticky part of the aggregates, which is known to polymerize.

The tau protein has—

Now with cryo-EM, they have a lot of good idea of how this THF.

Right. So there's a— I'm going to back up so that everybody can hopefully get the question. So tau is this pretty big protein. It has a chunk of it in the middle, which is the part of it that forms these structures, these what are called parathilical fibers or these aggregates or fibers. There's a lot of different names for them. But it's kind of this one chunk in the middle. That chunk— also will bind to microtubules. And it's thought that tau plays an important role in regulating microtubule dynamics, although the data for that in organisms is actually quite weak. So what I think the question is, is what's really the relationship between microtubule binding and these alternative fates of tau forming these structures? Pretty good evidence from human mutations which cause disease that if the tau binds to— is less able to bind to microtubules, it's more prone to forming these aberrant structures. And I think of this as like a competition, that microtubules keep the tau in a healthy state. And if they fall off that, they can enter this other state. I will say, when we look at tau molecules in cells, we see tau in the nucleus quite frequently. And this has been reports in the literature as well. And I think that tau plays a role in the nucleus as well. It's just not well understood yet. And it's not just a microtubule-binding protein. That's my interpretation so far.

All right. So I have a question from online. How much of the spreading is independent of cell death? And how much is for sure dependent on cell death?

I don't know. I don't know the answer to that question. I think there's certainly models and evidence that things can go outside of cells. But whether seeds can also pass through what are called nanotubes or through extracellular vesicles, I think there's also some data that suggests that's true. So I think I don't have a good answer, which is the relationship between those. Yes. Hi.

So I'm a physicist, not biologist. So I'm very interested in this 40 Hz oscillation thing that you talked about. That sounds like magic to me, although I want to say, I mean, one possibility is my understanding that this affects protein folding.

Is that what you're— Yeah. OK.

So I can imagine that that means that you could even model this in a protein folding system by putting a perturbative oscillator or something like that. But also, the other thing that it drew my attention to is that the natural— sorry, not the natural world. The world already oscillates at 60 hertz. This room right now, if we took a high-speed camera and we filmed it, we are now all being exposed to a very noticeable flashing light we've been exposed to our whole life. So I'm wondering if it affects protein folding, does it also have the potential that other frequencies cause harm and that we haven't noticed that before?

Nobody ever pieced it together. I can't speak with authority to those questions. I mentioned that because I think it's really fascinating. It's not my immediate expertise. I do think that their data in mice suggests that what it's doing is helping— it's doing something to trigger the clearance of aggregated proteins. And the way that they got into this initially is that they observed when they were studying mice that were genetically altered to have Alzheimer's kinds of diseases, that they observed that those mice didn't have any gamma oscillations or had greatly reduced gamma oscillations. We all have gamma oscillations all the time, but those mice were way down. And then they looked at people who had ADD, saw the same thing. And they said, well, maybe we can correct this. And it was really remarkable that it had an impact. I don't think it's protein folding per se, but that's a really interesting idea. Maybe we can talk about this at coffee, but this gets to a bigger issue, I think, that we don't understand very well as scientists, is how do these patterns of neuronal activity regulate our physiology? And I don't think we have really good answers to that. I certainly don't. But I think if we could understand that, we might be able to do some very interesting things.

So— Last question in the back.

So in looking at the movie, a question occurs as to whether or not SRM2 moving out of the nucleus, is that active or passive? And then related to that, is there a difference between the SRM2 that stays in versus that that goes out? Those are good questions. So first, we think it gets out by an active mRNA— RNA— not RNA— an active export pathway. We don't know which one. And then we think, but don't know, but we have some preliminary data that it actually has unique phosphorylations when it's out in the cytosol. And we're trying to figure out what that kinase is because if we could trap that back in the nucleus, maybe we could change some things.

So yeah. There's one more question in the corner over there if we can get to it.

Jeannie.

Jeannie.

Jeannie, we're going to give you—

Jeannie, we're going to bring you a mic. Thank you. How does this apply to different types of—

I mean, I'm thinking of Lewy body, for instance. Yeah. There's different parts to that question. So Lewy bodies are aggregates that are primarily made up of alpha-synuclein. And these are primarily seen in Parkinson's. But they also form in other diseases as well, just like tau aggregates can form in Parkinson's. So in terms of how does this apply, I think there's— in terms of— Lewy bodies are the only thing that we don't find RNA in. So there's no RNA in Lewy bodies for whatever reason. So they're different than tau aggregates, and they're different from TDP-43 aggregates. So I think there's some differences there. In terms of, I think, understanding how these fibrils form, I guess your question raises an interesting question. If we looked at the formation of— Lewy body aggregates, would they form on this structure as well? We have not done that experiment. But we have not pushed this into understanding of Lewy bodies. But that's an interesting suggestion.

Another question?

Clark?

I'm wondering about the connection between neuroplasticity in the brain and why we don't know that— and neurons dying. What's the connection there?

I don't really have any expertise to answer that question, to be honest. The obvious prediction would be that as you lose neurons, your plasticity and your ability to rewire and to form new connections will go down because you have less neurons to work with. But I don't have any real expertise there.

All right, I think we have one last question in the front, and then we'll need to move forward.

Hi. Having lived in England in the '80s during the mad cow scare, having not eaten hamburger and the like, what is the— the impression I have, and I haven't looked for quite some time with Jakob-Creutzfeldt disease, is the timeline. When I was in the UK, people were actually developing and dying from this disease. It wasn't taking 5, 10, or 10, 20, 30 years. What is the difference? What drives that forward at least appear to be a faster rate than in this case with some of these other illnesses, folding diseases?

Yeah, that's a really interesting question. You know, the timeline of mad cow disease, you're absolutely right, it was relatively short. But yet in the New Guinea natives, when Jakob Creutzfeldt, it actually tended to be longer. But it's still driven by this protein called the prion protein, PrP, the original prion form that was described. So I don't have a good answer to that. That's a really interesting question. Why in one context do you get pathology and disease in a few years, and in another case, it takes 20. If we can understand that, if you can change 20 to 40, that makes a big difference in these diseases, right? Because now you go from an onset of 80 to 100, and it becomes less of an issue for society.

Thank you, Roy. That was fantastic.