09.04.2024

Exploring the Hidden Life of Cell Organelles with Yamuna Krishnan

🎙️ On this episode of the Nanion Podcast, we had the pleasure of speaking with Prof. Yamuna Krishnan, a world-leading expert in bionanotechnology, working at the interface of chemistry and cell biology.

Together with Yamuna, we dive deep into the heart of cellular function, exploring how the chemistry within cell organelles is reshaping our understanding of health and disease. From measuring elusive ion concentrations inside organelles to discovering the “hidden life” of ion channels and transporters within cells, Yamuna Krishnan’s insights are transforming the future of bionanotechnology and therapeutic development.

🌟 Episode Highlights:

  • The challenges and breakthroughs in studying intracellular ion homeostasis.
  • The innovative DNA nanodevices making invisible cellular processes visible.
  • The unexpected discoveries around organelle membrane potential.
  • The surprising activity of plasma membrane ion channels inside organelles.
  • The intriguing role of TMEM165 in lysosomal calcium entry.
  • The two transformative companies that leverage organelle-targeting nanotechnology for the early diagnosis and treatment of neurodegenerative diseases and cancer.

Whether you’re a scientist, a student, or simply someone fascinated by the cutting edge of biology, this episode offers rich insights into the pioneering research that’s setting the stage for new discoveries and therapies.

🎧 Listen now to be inspired by the possibilities that lie at the intersection of chemistry, biology, and technology.

 

📖 Prefer reading? Check out the full transcript below.

 

Hello everyone, and welcome to Nanion Podcast where we talk about all things ion channels, transporters, nanopores and more. My name is Artem Kondratskyi, and I will be your host today. Now, in today’s episode, we’re going to explore the world of cell organelles. You know, intracellular ion homeostasis and the role of ion channels within cells have emerged as a hot topic and have gained a lot of interest due to their close connections with human diseases.

However, studying intracellular ion homeostasis and the function of ion channels in organelles presents significant challenges, leaving many questions unanswered until recently. For instance, how do we measure the concentration of specific ions inside particular organelles? What is the membrane potential of different organelles, like lysosomes, recycling endosomes, or Golgi vesicles? Or, do plasma membrane ion channels function solely at the cell surface, or do they play a bigger role within the cell?

If you’re curious about the answers to these questions and want to learn more about cell organelles, then get ready to enjoy today’s episode of Nanion Podcast.

Joining us is Prof. Yamuna Krishnan, a world-leading expert in bionanotechnology, working at the interface of chemistry and cell biology. Currently, Yamuna is a professor of chemistry at the University of Chicago where she also leads a research group exploiting DNA-based nanodevices to study the chemical composition of cell organelles. Also, Yamuna is a co-founder and a CSO of two companies, called Esya and MacroLogic, that utilize her organelle-targeting nanotechnology for early diagnosis and treatment of neurodegenerative diseases. For her groundbreaking studies, Yamuna Krishnan has been awarded multiple awards, and was on Cell’s 40 under 40 list of scientists shaping current and future trends in biology. Thank you so much for joining us today, Yamuna. I’m looking forward to learning from you. How are you doing?

 

Thank you so much. I’m excited to be on this podcast and it’s wonderful to engage with the ion channel community.

 

Great. So, let’s start off our conversation with you setting up the context of your work for our listeners, telling us about the significance of your work. Basically, why cell organelles? What are the problems you are trying to solve with your research on cell organelles, and what are the challenges?

 

Yes, so organelles as you know, are sort of communicating networks of subcellular compartments and each subcellular compartment has a distinct biological function, because proteins and lipids are confined within organelle lumens and these lumens have a microenvironment that is tuned to promote the specific biochemistry that occurs inside that kind of organelles.

And these biochemistries are vital to producing, modifying, transporting or degrading biomolecules in and around the cell. And the biochemistries themselves are highly varied, because in the ER you have protein synthesis; in the mitochondria – ATP synthesis; proteolysis is happening in the lysosome; glycosylation is happening in the Golgi. So if you’re looking at organelles, they can be viewed essentially as a group of nanoscale biochemical reactors, that must constantly maintain a unique luminal ionic environment, while surrounded by the same bath of cytosol. They all share the same bath. And while the ionic milieu of the cytosol and nucleoplasm was fairly well defined, when our lab started in 2005, little was known about the ionic milieu inside organelles. And because of the paucity of chemical information on these micro spaces, we developed a technology to measure ions inside organelles. And it turned out that DNA nanodevices proved to be the most enabling scaffold with which to achieve this goal.

And you also mentioned something about the challenges right? You know, it’s interesting to consider the nature of the challenge by just considering the timescales in terms of unveiling the chemistry inside an organelle called the lysosome.

So, the first ever ion that was measured inside an organelle was actually done by Élie Metchnikoff, when in about 1896, he measured the pH of the lysosome. And he found that it was highly acidic. And thereafter, the second and third ions that were measured inside the lysosome were chloride and calcium, and that was done nearly 130 years later, by us, using DNA nanodevices. And it’s interesting to consider why there is a 130 year gap between measuring the first ion and the second ion. And why is it that every subsequent ion that was measured inside the lysosome was done like every two or three years? And this is because there are actually four major but completely separate technical challenges that needed to be overcome in order to measure ions inside organelles.

And what are these technical challenges?

The first challenge is actually a measurement problem. And it’s a chemical and an analytical challenge. And actually, what we’re doing is measurement versus detection. And that is significantly more challenging, because detection is qualitative, whereas measurement is quantitative. So you have to create ratiometric sensors, you have to validate them, you have to develop methods to calibrate them inside a living system, and then measure the absolute level of ions.

The second problem is a biological challenge of access with specificity. To solve this problem, you have to localize your probes inside a 20 attoliter volume of an organelle that is present inside a cell that is present inside a living organism. On top of that, your probe is externally introduced, and it has to contend with the various biological barriers presented by the anatomy of the living organism. And then it has to access the lumen of your organelle with precision and cell type specificity. So that’s not easy.

The third problem is the classic Blackbox problem. You have to design a reporter to measure ion levels, but you have no prior knowledge of what these ion levels are. And typically, a sensor can measure like within 1.5 to two orders of magnitude surrounding the binding constant of any dye. But for ions like calcium, you know it can range between nanomolar in the cytosol to millimolar in the extracellular site. That’s five orders of magnitude.

And even if all of these obstacles are overcome, final challenge is actually the acidity problem, because most organisms are acidic, and that acidity interferes with every kind of ion detection chemistry.

And so because of these four challenges, the luminal chemistry of organelles remained kind of unstudied, and unmapped, and unused in our understanding of health and disease.

And it looks like you’ve managed to overcome these challenges with your DNA-based nanodevices. So, for those who are not familiar with your work, could you explain in simple terms what these DNA-based nanodevices actually are and how they work.

So these DNA nanodevices, they’re just sort of nanoscale assemblies that are formed from synthetic DNA oligos that have artificial function engineered into them. Our DNA nanodevices are used to quantitate ions and second messengers and everything inside organelles that are present inside living cells and living organisms. And the reason we use DNA is because it’s a modular scaffold, so we can put independent functionalities onto each strand. So our devices will typically have three or four DNA strands, and each DNA strand will carry a specific motif or module that has a specific function.

So for example, one strand will carry an ion sensitive dye, or let’s call it a detection module. The second step will carry a normalizing dye for ratiometry, that together with the first strand can allow it to measure, so together they form the measuring module. And the third strand can be also integrated into that assembly, and you can make it bear some kind of a trafficking module, like an endocytic ligand, and then the whole assembly can backpack onto the trafficking protein and reach the lumen of the target organelle.

And so together, these modules allow us to measure ion concentrations, with accuracies that were previously unattainable in subcellular locations that were previously inaccessible.

So, you mentioned three strands. Why three? Is this the limit, or is it theoretically possible to create a nanodevice with five or ten strands, for example?

Yes, yes, it’s indeed possible, but always we have to ask ourselves, what is the function that you need? And does adding another strand really allow you to answer a question that you couldn’t with a smaller set of strands? And if there is a question like that, then for sure, we should increase the complexity of the device. Every time you put more DNA strands your structure becomes more complex and in biology, and in any technology, the simplest and the most basic kind of design is what you want for robustness of function. The more moving parts you have, the more things can go wrong.

So yes, in fact, there are DNA nanodevices that have hundreds of strands. These are megalith, highly exquisite architectures that people have made in vitro, and you see them on the covers of Nature and Science. But putting them inside a living system is a different ballgame. You have to be very careful when you’re dealing with a live cell. If you hit it with that much DNA from outside, the first thing that’s going to happen is that that cell is going to die. So you want to keep whatever you’re using to probe a cell with as minimum functionality, molecular weight, and everything that gives you the function you want. So our design principle is a very sort of Japanese design principle which is very minimalist. We don’t go for Rococo architectures. You know, even as a designer, you need to have your style and our style is basically: what do we need to do to keep the cell in as unperturbed state as possible?

I really like how you’ve described it, Yamuna. The Japanese design principles and your style as a designer… I like that analogy. So, from what I’ve understood, your style is minimalist, and you create nanodevices with three strands: one equipped with a detection module, another with a normalizing dye for ratiometric measurements, and the third carrying the trafficking module, which is crucial for delivering the entire nanodevice into the organelle of your choice. So, which organelles have you studied with your DNA probes so far?

So far, I think the kinds of organelles we’ve managed to probe are nearly any type of endosome, like the early endosomes, late endosomes, recycling endosomes, lysosomes, phagosomes, and also the trans Golgi network. And of course, you can always interrogate the plasma membrane.

So one of the reasons why we managed to go to the immediately access all of these endosomal networks, is because the targeting works by engaging receptor mediated endocytosis. So, most of the time, the moment you find a receptor, you can latch on to that receptor, which then takes the device into your organelle of interest. And if you’re tapping into the endocytic system, you can immediately access all of these specific organelles. And because there’s a very interesting way by which, through the endosomal network, you can access the trans Golgi network, via the retromer, that’s why we’ve also been able to access the trans Golgi network. But we are now thinking about how do we access organelles like the endoplasmic reticulum, or the mitochondria? These are not going to be easy problems to solve, but we’re thinking about it and still trying to develop ways by which we can get there.

And how do you manage to differentiate between different types of endosomes? For instance, if you want to study recycling endosomes or lysosomes without receiving any unwanted signals from early or late endosomes, how do you manage to selectively target your probes to a specific type of endosome?

You know, we would have the same accuracy in time and space as any endocytic tracer. So if you want to look for stable localization in recycling endosomes, it will initially label the early endosome, but then finally, because the receptor you’re using is highly enriched in recycling endosomes, you can very stably be looking at recycling endosomes for a long period of time. But initially, right at the beginning, it will transit through the early endosomes, But if you wait for some time, 95% of all your marked organelles will be recycling endosomes. There’ll be 5% or something which are likely to be early endosomes but in biology, that’s how molecular markers work.

For the lysosome, that’s great because it’s the endpoint, everything just goes there. But there you have to contend with things like stability. Because the lysosome is a highly degradative organelle, it just eats everything. So when you’re probing the lysosome specifically, you have to actually look for the stability of the DNA nanodevice. And so we’ve had a whole range of papers that show exactly the stability of these DNA nanodevices in every single, very popularly used systems. And these devices typically have a half life of eight hours inside the lysosome, but it takes roughly an hour for your devices to reach a lysosome. So within the first hours, you can basically make a measurement and leave. So if you’re interested in probing for a very, very long time, you might want to change out your DNA device. It’s not like GFP, where you can look at it for a very, very long time, but you have a window a time window within which you can get useful information.

So, we were discussing the trafficking part and the delivery of these DNA-based probes into specific organelles. Now, let’s move on to the detection part. What types of chemicals, ions, or processes can you detect or measure with your DNA-based nanoprobes?

So, initially, we had only done pH for a while and then we found out that you can really use any chemistry that has been used in the past to detect analytes in solution. You can take that chemistry, stick it on a DNA strand and measure that chemical. So we’ve thereby developed reporters for ions and for reactive species, like nitric oxide. We’ve even mapped enzymatic cleavage in real time within organelles. And in fact, very recently, we also measured absolute membrane potential of organelles. And it’s just as simple as taking an existing chemistry that is well known, well accepted, integrating that and then converting qualitative chemistry into quantitation and spatial resolution.

When you mention having developed reporters for ions, which specific ions do you mean? Which ions have you measured with your nanodevices?

I had been very interested in how organelle insides are communicating with the outside in the cytosol. And so we’ve developed reporters for ions like protons, chloride, calcium, sodium, potassium. And together, what we learned was that when we started making these measurements inside organelles, we found that there were very high transmembrane ion gradients across organelle membranes. And that immediately raised the question: should we also be measuring membrane potential? And that’s why we made this membrane potential reporter. And so one can envisage that you can use this to measure ions that are trace elements, like copper, manganese and things like that, but they are present in such trace quantities that the concentration of your DNA nanodevice is enough that you perturb the system, because you’ve taken away the few ions that are there to measure. And all of the ions that I talked to you about they’re all in millimolar concentration. Our sensors’ maximum concentration is micromolar, so the amount of signal they take is enough that we get an idea of the measure, but you haven’t biologically perturbed the system.

Since you’ve measured all these different ions, I’m curious about the results. How are ions distributed inside the cell? We know that, for example, there is 100 nM of calcium in the cytosol and 1 mM in the ER. Mitochondria and lysosomes have also been reported to contain calcium in the micromolar to hundreds of micromolar range. How about other ions and organelles?

I’m afraid that this can’t be answered in a soundbite, because each organelle has its own characteristic level of each of the major ions. And that is what contributes to the membrane potential of the organelle. And these values vary greatly depending on the type of organelle. Except for protons, the values of ions across the plasma membrane broadly will represent the two extremes that an ion level can assume. This is what you talked about, when mentioned calcium concentrations across the plasma membrane. In organelles, you’ll get the values somewhere in between the cytosolic and extracellular.

So let’s talk sodium for example. Extracellular sodium is 140 millimolar as you know, and intracellular sodium ranges between say five to 10 millimolar. Organelles of the endo-lysosomal pathway would be like, say 70 millimolar in the early endosomes, 50 millimolar in the late endosomes and 40 millimolar and the lysosomes. And we are yet to quantitate sodium in the recycling endosomes and the trans Golgi network, but it’s likely to be somewhere between five millimolar and 140. And that said, when we measured sodium, which is very unique compared to every other ion, this particular ion shows a very wide distribution in every organelle type that we looked at, and so something very interesting is going on with sodium inside organelles.

It’s clear for sodium. And, what about chloride ions, for example?

For chloride, the distribution of values is fairly narrow, but chloride is actually increasing as we go down the endo-lysosomal pathway. It increases from the early endosomes, is little higher in the late endosomes and by the time you reach the lysosome, it can be about 110 to 120 millimolar chloride. In fact, lysosome is a highly chloride rich organelle – this is what we found. It has I think 10 to 20% higher chloride inside the lysosome than the extracellular site. So it’s interesting to see how the lysosome is somehow enriching for chloride. And I feel that that’s because if you’re looking at all the ions except for sodium, they all are increasing, as we’re going down the endo-lysosomal pathway. Calcium is increasing, protons are increasing. We don’t know what the value of potassium is. But you can see that there’s only one anion that has to somehow be there in that bubble, to counter all of the other cations. And maybe that’s why chloride is so high in lysosomes.

You’ve mentioned the DNA voltage sensor and the organellar membrane potential a few times. Have you measured the membrane potential of different organelles?

Yes, we have. So, one of the most interesting things we found was when we probed organelles like the recycling endosomes and the trans Golgi network, with Voltaire, which is a membrane potential reporter. So, people had previously stated, without much evidence, that the recycling endosomes and trans Golgi network don’t have membrane potential. And so when we measured it, we actually found that they had membrane potential and pretty high membrane potential. The recycling endosome has the membrane potential, which is very similar to the plasma membrane. And the trans Golgi network has a membrane potential that’s as high as the lysosome. We found organelles like the early endosome had membrane potential higher than the lysosome. No one even considered whether these early endosomes had a membrane potential. So suddenly Voltaire transformed the idea of membranes of organelles from one that were like an inert plastic bag that acted as a surface, where adapter proteins will dock onto the organelle and drag the organelles around the cell etc., so changed from that to one where these membranes are fizzing with ions, continuously communicating their luminal status with the outside world.  They are active participants in their destiny.

Could you provide some specific values of the membrane potential of different organelles?

For sure. So, as you know, lysosome membrane potential has been measured, but it’s not like one size fits all. Lysosomes is some cells have plus 100 mV. I mean, it’s plus because the sign is inverted, because the membrane topology is inverted. And you’re looking at it as lumen positive. So, it’s plus 100 mV in some cells, but if you do this measurement in all different other kinds of cell types, it can range. So, plus 40 mV we’ve seen in some cells. In RAW macrophages, it’s plus 20 mV. The Golgi has pretty high membrane potential. It’s usually like 110 to120 mV, much less spread than in lysosomes. The early endosome has 135 mV, the late endosome – plus 40 millivolts, the recycling endosomes – plus 70 mV. And all this tells you that you could have channels opening there.

Yes, exactly. This immediately makes me think of voltage-gated ion channels, which could potentially be active in organelles. And in one of your recent papers, you demonstrated that HERG channels can be active in organelles. A very interesting study where you talk about the intracellular activity of plasma membrane ion channels, which is quite surprising. You know, this paper actually reminded me of my interview with Henry Colecraft (which I did for the Ion Channel Library) where he said that biophysicists are very good at measuring the activity of the channels once they arrive on the cell surface… But the reality is that most of the lifetime of the channel is spent inside the cell. Henry called it the hidden life of an ion channel. And they are now working to develop tools that will allow for spying on ion channels throughout their lifecycle, not just when they are on the cell surface. So, to me, your paper on HERG channels in organelles was like looking at the hidden life of the HERG channel inside the cell. Could you elaborate on this interesting finding and also comment on the potential physiological importance of such intracellular activity?

First, thank you for giving me a title for my next talk. I think I’m going to call it “The hidden life of ion channels”. That is so interesting. Yes, it was a long-held tenet that cell surface potassium channels are active only at the plasma membrane. And, we wanted to test whether cell surface channel could also work in an organelle. If you look at the lifecycle of a potassium channel in the cell, it’s born in the ER, it gets fully glycosylated, fully competent, fully functional when it reaches the trans Golgi network. Then it’s packaged into secretory vesicles, and then sent off to the plasma membrane and there it gets inserted in the plasma membrane. Then, as the cell continues its regular job of endocytosis, bits of plasma membrane containing these channels are sort of inadvertently scooped up into regular endosomes. And the cell has to save these good and properly functioning channels, and sort it, and send it back to the plasma membrane because it spent so much energy to make an ion channel. These are complex machines, and you don’t want to simply take it and then send it off to the lysosome to degrade, if it’s still a well functioning channel. So it’s then sent back to the plasma membrane via the recycling endosome and at some point when it’s damaged these channels are sent off to the lysosomes for degradation. And so if you asked me how was an early endosome supposed to realize that this potassium channel is not damaged, and that it has to send it to the recycling endosome, so that it could be put back on the cell surface? Or if you ask, how does early endosome senses damage in the organelle and send it off to the lysosomal pathway for degradation? Could it be say channel activity in the organelle? So that’s why it’s started getting us to think about cell surface ion channel activity in organelles.

Oh, this is very interesting. So, do you think the organellar activity of cell surface ion channels could, for example, be important for the cell to determine when to send the channel to the lysosome for degradation or when to send it back to the plasma membrane?

There are a whole host of potential functions. So, we know that a cell surface ion channel can work inside an organelle. What is the endogenous function of that? Nature must be using it for something. And I’ve been thinking about what it might be, and so for example (this is all speculations at this point) I think that channel activity might act as some kind of an endocytic clock or an exocytic clock. You know, the pH and the membrane potential can actually change the channel open probabilities. And that, in turn can dictate how long an organelle can stay in its specific state, before maturing to the next step on the secretory pathway, or the endocytic pathway. And I’m thinking that, if the channel doesn’t open I see the organelle in the same state, but once it opens, it can change the membrane potential, that can cause key other transmembrane proteins on the organelle to change their conformation, and recruit other players. You know, I have a hard time believing that proteins like these Rabs, and these SNAREs, they somehow stochastically dock on an organelle to mediate organelle contact, or transport or fusion. The organelle itself must be ready and willing to engage the Rabs and the SNAREs. But it’s still early days, and we allowed to dream of possibilities. So I think that the heart of an organelle deciding to function is likely a channel on the organelle.

Endocytic clock, exocytic clock. That’s a very exciting hypothesis. Now, moving from cell-surface channels to intracellular channels, your most recent article on the mechanism of lysosomal calcium entry has made quite a splash on social media. You’ve shown that TMEM165 acts as a proton-activated Ca2+ importer in lysosomes. This was quite unexpected given the known role of TMEM165 as a calcium/proton exchanger in the Golgi. Also, given the increasing interest in the lysosomal channel TMEM175 (for its role in Parkinson’s disease), discovery of the TMEM165 as another important lysosomal actor looks even more exciting. Please, tell me more about this intriguing TMEM165 story.

So it was actually quite a surprise to us too that it turned out to be not a calcium-proton exchanger, but instead what it is, is that it is a proton-activated calcium importer. And I found myself sort of half wishing that it had been a CAX (calcium-proton exchanger), because then our paper would have been a cleaner story, it would have satisfied the reviewers and it would have happily walked through and be published in Science. Instead, because of its non canonical electrophysiological characteristics, the reviewers noted that this cannot be a CAX because there is no change in the reversal potential. Even though it needs a transmembrane pH gradient, and it needs transmembrane calcium gradient in order to have a calcium current, there is no change in the reversal potential. So it is not a CAX never mind what others have said earlier. So you know, we actually are not ion channel people by training, so looking at the previous papers, we thought okay, let’s try and see if we can prove these reviewers wrong. We will use our DNA nanodevices and map calcium and pH in lysosomes when the protein is in action. And then we can show that it’s actually doing a calcium and a proton exchange. Surprise of surprises, the reviewers were right. We were wrong, and the previous group was wrong in thinking of it as a CAX. But regardless, what we then found was that it turned out to be an even more interesting protein than everyone thought. This is now the first calcium import mechanism in lysosomes, and it’s now in another journal that I really like, called Science Advances. And I’m very grateful that the reviewers actually appreciated that breakthrough for lysosome biology. But I want to say that this protein is very interesting even for electrophysiologists as well as structural biologists. And I hope people will come forward to help us solve the structure and try to understand: how is it that acidity on the luminal side makes this protein start to import calcium? Whatever you do with the gradient, it can never send calcium out the other way. That is why it’s not a channel. It will only take one way that is coming inside the lysosome.

So if you look at what was known previously, about calcium entry into the lysosomes, lysosome has to have a way to replenish its stores. So there were two theories proposed. One was this calcium-proton exchange. The other was, and it’s a beautiful story, that lysosome comes very close to the ER and then the ER releases some calcium in the vicinity. You have organelle contact and then suddenly in that synapse between a lysosome and the ER you probably have high calcium over there and then somehow calcium enters the lysosome. But you could think of TMEM 165 being that molecule. The moment I have high calcium in the cytosol, either when the cell is overall excited or if the lysosome needs to replenish and comes close to the ER and then the ER releases calcium in that space, then it can act. That’s how I’m thinking about it.

However, exactly how it’s working. We’ll only be able to understand when we have some kind of a structure or mutagenesis studies. For example, TMEM 175 was found to be a potassium channel. But there are competing papers that have said that on the lysosome you have another factor that is associating with it, that silences that channel, potassium channel activity, and it’s actually a proton activated proton channel. So, we address this and other people also have addressed TMEM 165 function by putting it on the cell surface. It’s a foreign membrane for the TMEM 165. And we don’t know what might be the modulators of it in a native membrane, which is why I think Nanion’s SURFE2R technology is very exciting, because it allows you to probe ion channel function in its native membrane state.

Yes, indeed, the SURFE2R technology could be really instrumental in such studies, especially when it comes to electrogenic transporters. Thank you for mentioning it. And I think it’s a good opportunity for me to remind our listeners that this podcast is brought to you by Nanion Technologies, a global provider of high-performance electrophysiology instrumentation for life sciences. We specialize in automated patch clamp, membrane biophysics, and cell analytics instruments. If you’re curious about how our technologies could advance your research, please visit our website at www.nanion.de. We’ll be happy to help you accelerate your research.

Now, let’s jump back into our conversation. So, what’s next for Yamuna Krishnan’s Lab? What are the next steps in your research? Are there other ions or organelles you plan to study using your DNA-based nanodevices?

You know, when I first started, Artem, I just wanted to understand what are the levels of ions in these organelles, just so that I can see what is driving the different biochemistries. But now I’ve become very interested in understanding the ionic determinants of organellar membrane potential, because it’s a way by which organelles are communicating to the outside. And if you ask me immediately what we’re doing, we’re trying to get maps for organelles we can’t access at the moment, like the mitochondria and the endoplasmic reticulum. It’s all very blue sky at the moment. We have some interesting new results. But I have to try and remain very calm and Zen about it all because I know my students and postdocs are as eager as I am to do it, but I have to give them the space they need to crack the problems they picked. But I think I’m very, very excited about new organelles like the ER and the mitochondria, but also very interested to understand what are the ionic determinants of the membrane potential of an organelle membrane? We know that for the plasma membrane. Organelle membrane is completely a black box, because we didn’t think they have membrane potential. But now we actually know quite a bit, and it would be great if we can build a model of that.

Sounds like the future holds some exciting discoveries from your lab. I’m looking forward to it. Now, my final question is about two companies, Esya and Macrologic, where you are the co-founder and CSO. Could you share what these companies focus on and how they relate to your scientific work?

Yes, you know, I never thought that I would ever have any companies. I just wanted to do fundamental research. In fact, the organization that I joined back in India was called the Tata Institute of Fundamental Research. I didn’t want to have anything to do with industry. I just wanted to advance the edge of knowledge.

But I think when you do something very fundamental, it will at some point have some application. So since we were able to map ions in organelles, you can think of two applications immediately for an organelle specific delivery technology. One could be in diagnostics, and the other could be potentially in therapeutics.

So we have a diagnostics company and that company is called Esya, and it uses our nanodevices to profile ion levels inside lysosomes. So this has now come as a very nice platform for early diagnosis of neurodegenerative diseases. And I think the first product is for very early detection of Alzheimer’s disease.

The second company is Macrologic. This is a company that uses our patented organelle delivery technology to deliver therapeutics to lysosomes that are present inside tumor-associated macrophages. So we found that our devices go straight to macrophages, and specifically to lysosomes. And this is especially true for tumor-associated macrophages. So in the environment, if you have tumor-associated macrophages, this is the one place where it gets completely enriched. And so we’re trying to use that now to turn cold tumors hot. As you know, cold tumors are tumors that don’t respond to immunotherapy. But now if you change the macrophages ability to present antigens, you can now get T cells to come there. So these are the two aspects of the technology. Second has nothing to do with ion channels, though.

Well, I guess we can never completely rule out the involvement of ion channels… But, anyway, it sounds really exciting, even though it’s not about ion channels, as you said.

I would always say it’s not about ion channels yet.

Putting it that way makes it sound even more exciting and intriguing. Thanks a lot.

So, it was great talking to you today, Yamuna. Thank you for sharing your insights with me and our listeners. I wish you all the best and look forward to exciting new discoveries from your lab.

Thank you so much. It was great talking to you