In 30 years I witnessed this progression, in 30 years from now, what story will you tell? If you think big it is going to happen.
Transcript
Matthew Kose-Dunn: Hello and welcome to Science Off Camera. I’m Dr. Matthew Kose-Dunn from Teledyne Photometrics, part of the Teledyne Imaging group. In this podcast, I’ll be speaking with imaging specialists and industry leaders in scientific imaging from around the world about what they do, the advances they have made, and the cool imaging setups they have in their labs. In this episode of Science Off Camera, I’m speaking with Dr. Seamus Holden, Sir Henry Dale Fellow and group leader at the Centre for Bacterial Cell Biology at the University of Newcastle in the UK, I’ll be speaking to Seamus about science, his life in research, and how he’s adapting to change in his world.
Seamus Holden: My name is Seamus Holden, I’m a Wellcome Trust Henry Dale Fellow, and we work in my lab, there’s eight of us. We have two postdocs, a bunch of students, and myself, and we work between physics and microbiology, hopefully trying to do do work where we learn something about both disciplines, right? But essentially, what we’re interested in is how bacteria grow and divide. Bacteria are surrounded by this peptidoglycan cell wall, right? That’s their coat of armour and provides their protection against osmotic insults and environmental insults and it gives them their shape. And it’s: how do you build that structure? It’s this giant sugar peptide macromolecule, this big mesh. Coming to this as physicists, you’ve got proteins, which are responsible for building the structure, which are say, order of nanometers size. The cell wall, the bacterium, is like a micron scale. So you’ve got a thousandfold scale difference. And yet, bacteria build these highly regular shapes, they’re way more regular than eukaryotic cells, these beautiful structures. I mean, we tend to study rods but you get spirals, spheres, all sorts of diversity and shape is really important for bacterial survival, right? But the scale differences like between a human builder and the tallest building in the world. And yet these proteins spontaneously self assemble to build highly regular structures without some, you know, fancy architect and foreman keeping an eye on them. So that’s just a fascinating problem.
The best way to look at that sort of thing is through microscopy. Live cell high-resolution bacterial microscopy is like microscopy on hard mode. Bacteria are tiny and super photosensitive, not that they’re necessarily any more photosensitive than eukaryotes, but because their volume is so small then all of the fancy techniques that you could use for eukaryotes like light-sheet, still illuminates the whole bug, whereas with eukaryotic cells, you can illuminate a thin slice. So phototoxicity and photobleaching are much worse even for like TIRF type approaches. In a bacterium TIRF is going to illuminate like a sixth of the cell, in eukaryotes, you’ve got a thin slice, and then a really thick cell. So actually, you’re just illuminating like a tiny, tiny, tiny fraction of your current cell. No matter which way you illuminate a cell, your light dose is always way higher. This means you need to use reasonably smart tricks to not just fry the bugs, and actually be able to look at them for extended periods. Sensitive cameras, for a start, are really important, sensitive, high dynamic range cameras. That’s why we’ve had a lot of fun with the Photometrics BSI, we did some testing on that for you guys a while back, we got a little application note. Because they’re like, turbo EMCCD, plus higher dynamic range. That’s been very nice. But denoising is the thing. I still love super-resolution microscopy. That’s where like I started and we’ve still got projects on that and it’s super exciting. But I think the real frontier is high resolution, live cell dynamic single molecule with a single filament, and for that, I think denoising is just the really exciting frontier. Because anytime you try and do super-resolution you need a higher light dose, so you can’t image for as long, so your dynamics aren’t sensitive. So what we really try and do is clever mobilisation strategies, we have things where we stand the bacteria on their heads so we can look at the division plane better, and develop the method to solve the impossible problem. Normally you view bacteria side-on during division. These are really quite nice images and they’re denoised, and it’s quite sensitive. But if you want to image cell division, right, you’ve got a rod-shaped cell, and you’re viewing it side on and you can do 3D imaging approaches, but that’s more phototoxicity. It’s lower resolution, right? By having the bacteria standing up, we’re only imaging a small cross-section of it, and we’ve learned quite a lot about division. But there’s still a hell of a lot more to figure out.
I think the real frontier is high resolution, live cell dynamic single molecule, and for that, denoising is really exciting.
Matt: So the aim of this research is to disrupt bacterial cell division?
Seamus: Yes, yes, exactly. So there are drugs, we’ve actually been looking at the mode of action of PC19 a lot, there are specific drugs that target the bacterial cytoskeleton and cell division, right. They’re great antimicrobial targets, a derivative of one of them actually completed phase one trials as an anti Staph oral compound. These are like serious compounds, serious drug candidates. If you can better understand the mode of action, it’s really important for drug approval, but also if you know the mode of action you can understand why and how likely resistance is to evolve. That’s kind of the justification for really delving into the physical mechanisms of this process.
Matt: So how did you get to this stage? Personally, what kind of path led you here?
Seamus: Pure physics to like, biophysics methods geek? At the start of it, I know Richard Dawkins is a controversial figure but his early science writing is just brilliant, I didn’t even do biology A level but I was studying physics at university one of my biologist friends lent me ‘The Blind Watchmaker’, and it just blew my mind. It’s been game over ever since. So I got into biophysics, I did my PhD when STORM, PALM, super-resolution microscopy, all of that was exploding. So I spent my PhD doing image analysis approaches for super-resolution microscopy and that was really fun. We had this STORM algorithm which was basically adopting astronomy approaches, with astronomers you’ve got your stars, a crowded stellar field, overlapping stars, and you need to measure their brightness, which allows you to measure the Hubble Space constant, which gives you the age the universe. But it’s about measuring the brightness of overlapping spots. You want to find the centers of multiple overlapping spots very effectively so you can go faster, we did the first one of them and it was great, it’s been really powerful for live cell super-resolution microscopy approaches, and that was proper methods geekery.
I did my PhD with Achilles Kapanidis in Oxford, and actually, Oxford biophysics has always been a real centre of bacterial biophysics. I got hooked on a project where we’re using STORM/PALM to look at RNA polymerase in E. coli and the spatial organisation thereof. I was just hooked, bacteria are like the simplest living organisms, right? So they’re very attractive if you have this grand goal of maybe totally understanding an organism. They’re hard to image, which is fun. I mean, it’s annoying, but it’s fun. Right. So then, yeah, I’ve just been studying bacterial biophysics ever since.
How do you practically do science? That’s quite an interesting question. It’s quite an interesting challenge.
Matt: It’s always good to see this physics-educated approach to biology, which is what the future is moving more towards now. These multi-disciplinary approaches are getting engineers and physicists to have their fresh new perspective on biological problems, and as a biophysicist yourself, you can see it has made a lot of progress.
Seamus: I always push back on that a little bit. Right? The thing is in biology, details matter. And in physics, in pure physics, actually, the whole point is like: what details can you throw away? That approach is still really useful in biology, but details really matter in biology. I think, for truly interdisciplinary approaches to be successful (either through collaboration or through transdisciplinarity), you need to combine the quantitative aspects of physics, knowing what you throw away, and a deep understanding of the biological problem. I think that’s really critical. Otherwise, you just end up with a surface approach.
When I set up my lab, I actually made this really rationalised choice, right, which was to move to the Centre for Bacterial Cell Biology and Microbiology Institute and kind of try and go native. So my colleagues are some of the leading experts on bacterial cell wall biochemistry, bacterial cell envelope genetics, this sort of thing. We now have this deep understanding of how bacteria grow. The alternative way of doing it is long-term collaboration between two groups, but where you really learn. Where the biologists learn how to be quantitative image data scientists, and the physicists learn the biological problem. Either way, I think I think we really need to step up sustained collaboration or occupation of that overlapping space. How do you practically do science? That’s quite an interesting question. It’s quite an interesting challenge.
Matt: It’s a good comment on the field. We had a moment during my PhD where our lab flooded due to building works, and they moved us into the maths and computing building, just because it had some free desk space. But I learned so much about that part of science and the theoretical and practical statistics. And you start to speak to some of these more tame statisticians, they look at your data and say ‘Oh, this is great data, you know, of course, to be relevant, you need to do this about 1000 more times.’ I’ve done it three more times. How’s that? So it’s, we’re gonna have to compromise? Because these are long-term experiments run on live organisms? Well, ideally, you’d have a couple million more data points. So you come to an understanding in the middle, somewhere
Seamus: Setting up that sort of dialogue is just wonderful, right?
Matt: Definitely! So for you, what part of the scientific process of research is your favourite, the aspect you enjoy the most?
Seamus: I think it’s that moment of discovery, right? Where it kind of all comes together and particularly to say we’ve worked really hard to develop this new approach to answer a question, and then you get the biological answer. Right. And you see something no one’s ever seen before. Like when I was sitting on the microscope on the one warm day of the year in Newcastle, we first got this microhold approach working, and I saw FtsZ wiggling around these bacterial cells. FtsZ treadmills in live bacteria, no one had seen that before, it turns out a bunch of us made this discovery simultaneously, but no one had ever seen that before. And I was like, yeah, that thrill of biological discovery is pretty unbeatable, right? But to get there, you get this fun kind of side benefit that you get to develop a load of exciting new tech as well. Geeking out on the tech is almost as much fun.
If we are enormously unproductive for the next few months, but my team’s mental and physical health is okay, then I’m fine with that.
Matt: Would you say the imaging systems are the most important part of the lab? If somebody broke into your lab, which one piece of equipment would you want to know was definitely still there?
Seamus: Our systems are all bespoke, custom super-resolution microscope, single-molecule tracking scope, and we’re in the middle of building another one. We’ve built the second one entirely in CAD, so we’ve got this complete 3D design that we’re going to share for open-source purposes. If you build it from scratch yourself, then it’s more hackable, say we needed things like ring TIRF, instead of negotiating and writing a grant, we could just build it in a couple of weeks, right? So much blood sweat and tears has gone into these microscopes, they’re the precious thing. Going in and shutting off the microscope incubator on Friday, when we were doing the lab shut down. That was less lighthearted. I don’t think that incubator has been off because we leave it on over Christmas, so it hasn’t been switched off for years… So we have these beautiful custom super resolution microscopes, we share the designs on GitHub for an open-source incubator, or focus shifter. I mean, we do our bit, but this is a wonderful movement going on in microscopy at the moment, that people are sharing their designs. If you do that, then you avoid rinsing, like six months of your time reinventing the wheel.
Matt: If you could have any other piece of equipment, would it just be extras for the microscopes? Are there other accessories and things that would make the research easier?
Seamus: The thing that’s exciting, where the tech development is going for us, is combining high resolution microscopy with microfluidics and microfabrication type approaches. For us it’s about putting it all together. So the nanofabrication for the fancy mobilisation strategies, the high resolution imaging, the microfluidics, possibly then zooming in and doing super resolution on top of it, maybe in a correlative fashion. Microfluidics offers single cell perturbation on top of that, so that that’s the next level, I think. But all of this hinges on really, really robust live cell microscopy for extended periods. So you got to just tickle them with light, suddenly, you need the microscopist, you need the biologist, you need the, what now we have to call data scientists, but we would have just called statistician or something in the olden days, right? You need you need all of them talking to each other and working together and funded.
Matt: Definitely the future is in as you said, more direct collaboration, more long term collaboration, be serious about living with and opening labs with people you’re willing to collaborate with, from different fields.
Seamus: To be fair to them, the funders are starting to listen to this sort of idea, the BBSRC had their ‘Physics Of Life’ call, Wellcome have their collaborative awards, that becomes the interesting challenge, how you sustain this large scale.
Matt: What is it you miss most about the lab and your time in the lab, other than the people and your colleagues?
Seamus: I’ve been talking to people about this a lot, there’s so much, we’ve got so much exciting stuff going on right now. When you take away our microscope or whatever, we kind of grind to a halt, right? Because we’re an experimental lab. If we are enormously unproductive for the next few months, but my team’s mental and physical health is okay, then I’m fine with that.
Matt: Do you try and keep the excitement and momentum going? Or is just about maintaining?
Seamus: I don’t know, we’re kind of feeling our way. I’m really conscious that I don’t want to push anyone too hard. If that passion for science is what gets you through then great. But if you need to just step back and like watch Voltron or something then that’s fine too. Strange times.
Matt: It’ll be interesting to see how things look once we come out the other end of this, if things move more remote for people who can, or if people just go on as if nothing happened?
Seamus: Well, I tell you what, if scientists end up travelling less out of all this, I think that from an environmental perspective, and also as someone with small kids, it’ll be right. I love hanging out with my science friends all across the world, but I think if we switch to more remote ways and reduce our travel and carbon footprint, at least that would be some sort of positive.
Matt: We’re all a lot more used to the AV equipment and being remote, but still being interactive. It’s amazing what difference just having the camera on makes.
Seamus: It’s been a real crash course, that’s for sure. We’re keeping seminar series and we have a weekly super-lab in our building, we’re keeping that going. From the British perspective, what some of the postdocs are trying to figure out is how do you keep like the equivalent of the virtual pub after the seminar series? Because that’s quite a critical part of the community! So those are certain technical issues to still be resolved. How do you virtualize the pub? I’m not sure we’ll do that properly, anytime soon.
Matt: Everyone sharing a pint over webcam?
Seamus: Well, that yeah, that’s nice. That seems to be working a bit as well, you know. That keeps our kind of community cohesion going, which is nice.
Matt: It’s almost cruel that the weather is now really picking up and is quite nice. But we’re all confined to our homes.
Seamus: Yeah. I mean, we’re super fortunate. We have a lovely garden, and that that eeps the kids reasonably happy, I mean, they’re holding up really well.
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