Episode Transcript
[00:00:00] Speaker A: Foreign.
[00:00:14] Speaker B: Welcome to Base by Bass, the papercast that brings genomics to you wherever you are. Thanks for listening and don't forget to follow and rate us in your podcast app.
So for today's deep dive, we're getting into a question of really extreme biological physics.
[00:00:30] Speaker A: It's all about speed.
[00:00:31] Speaker B: Exactly how does a cell pull off these huge, ultra fast structural changes? I mean, think about it. A cell needs to secrete a massive protein, one that's critical for signaling. How fast can it do that without using the usual pathway, the standard ER.
[00:00:48] Speaker A: And Golgi assembly line?
[00:00:49] Speaker B: Right, so when scientists first tried to watch this unconventional protein secretion, they set it up in a lab dish.
[00:00:56] Speaker A: A cell free system.
[00:00:57] Speaker B: A cell free system, maybe a giant vesicle. And the whole maneuver, the protein literally punching a hole to get out, takes.
[00:01:04] Speaker A: Several minutes, which, you know, sounds reasonable in a lab, but that was the foundational puzzle.
[00:01:09] Speaker B: Why is that?
[00:01:10] Speaker A: Because when they looked at the exact same process, but in a living cell with some really sophisticated microscopy, the results were just, well, they were mind boggling. The entire translocation event, the protein opening, the membrane, slipping through, and the membrane sealing back up, took on average just 200 milliseconds.
[00:01:29] Speaker B: 200 milliseconds? That's. That's not just a little faster?
[00:01:33] Speaker A: Not even close.
[00:01:33] Speaker B: We're talking about a process that takes minutes in a dish happening in less than a quarter of a second. In a cell, the acceleration is on the order of, what, tens of thousands of times faster?
[00:01:44] Speaker A: It's the difference between driving across the country and blinking. I mean, that huge gap in speed told researchers that something fundamental, something biophysically critical was missing from their simplified lab model.
[00:01:56] Speaker B: So the cell isn't just letting it happen.
[00:01:58] Speaker A: No, it's actively driving it. With extreme prejudice.
[00:02:01] Speaker B: So if all the protein parts were the same in both systems, what is the living cell doing to get this, this incredible speed?
[00:02:08] Speaker A: And that is the mission for this deep dive. We're looking at new research that points to one of the key governing factors, the strict, precise, asymmetric distribution of lipids in the plasma membrane.
[00:02:20] Speaker B: So it's not just about what parts you have, but where you put them.
[00:02:23] Speaker A: That's it. Where you place them dictates the speed limit.
[00:02:26] Speaker B: Today we're celebrating the really elegant work of the team led by Manpreet Kaur, Fabio Lollicado and Walter Nickel at the Heidelberg University Biochemistry Center.
[00:02:34] Speaker A: Yeah, their study is a huge advance. It moves our understanding from just, you know, listing the proteins involved to actually dissecting the complex physics behind the speed and efficiency that really defines life.
[00:02:46] Speaker B: Okay, so let's set the stage. The process we're talking about is called unconventional protein secretion UPS type I.
[00:02:53] Speaker A: And the poster child for this mechanism is a protein called Fibroblast Growth Factor 2.
[00:03:00] Speaker B: And FGF2 is a huge deal. It's involved in everything from wound healing to. And this is really important, driving tumor progression and a lot of cancers.
[00:03:09] Speaker A: Correct. And because it's so vital for rapid signaling, it. It just can't wait for that standard Ergolgi assembly line. It takes a shortcut.
[00:03:17] Speaker B: It bypasses the whole system and goes directly across the plasma membrane.
[00:03:20] Speaker A: Exactly. Straight out to get its job done. Yeah.
[00:03:23] Speaker B: And there's this critical sequence of events that has to happen on the inside of the cell membrane to make that possible. Can you walk us through that?
[00:03:29] Speaker A: Sure. It's a very ordered cascade. So inside the cell, FGF2 first bumps.
[00:03:35] Speaker B: Into the sodium potassium pump, the now K ATPase.
[00:03:39] Speaker A: Right. That acts as its first receptor. This then brings in something called Teck kinase, which modifies the FGF2. But the final absolute key to the whole process is a specific lipid.
[00:03:49] Speaker B: Okay.
[00:03:50] Speaker A: It's called phosphatidyl inositol 4.4-5 bisphosphate.
[00:03:54] Speaker B: Let's just call it PI 405p.
[00:03:56] Speaker A: Yes.
[00:03:56] Speaker B: And PI 4.4.5p is a powerhouse on that inner membrane leaflet. It's known for causing huge changes to cell structure. What does it do to FGF2?
[00:04:05] Speaker A: So when FGF2 finds a cluster of pi405p, that interaction triggers a massive change.
It forces the FGF2 proteins to stick together, to oligomerize. To oligomerize? Yes, into these little clusters of at least four molecules. And this pileup of protein and lipid puts a tremendous amount of local stress on the membrane.
[00:04:24] Speaker B: It sounds like the cell is intentionally creating a weak point.
[00:04:26] Speaker A: That's a perfect way to put it. The stress forces the stable bilayer to change shape, to transform into a high curvature structure. Specifically a toroidal lipidic pore.
[00:04:37] Speaker B: Which is like a tiny self healing donut hole in the membrane.
[00:04:41] Speaker A: Exactly. FGF2 slips through that temporary pore. Once it's outside, it's immediately grabbed and disassembled by an external receptor called glipicon.
[00:04:49] Speaker B: 1, or GPC1, making it ready to go and send its signals.
[00:04:53] Speaker A: Right.
[00:04:53] Speaker B: So we know the machinery, but the problem, as you said, is that speed gap. The lab membranes, the guvs, they have lipids spread out evenly, symmetrically.
[00:05:01] Speaker A: But a living cell phikes tooth and Nail to maintain asymmetry, it keeps Pi4.5 pursue almost exclusively on the inner leaflet facing the cytoplasm.
[00:05:12] Speaker B: And that asymmetry was the prime suspect for the missing speed factor.
[00:05:16] Speaker A: Right, so the researcher's hypothesis was that this asymmetric distribution, unlike asymmetric 1, actively lowers the huge energy hill you have to climb to force that bore open.
[00:05:25] Speaker B: But to test that they couldn't just use regular guv's, they had to braid their own asymmetric ones. Which I imagine is a huge technical challenge. How'd they do it?
[00:05:32] Speaker A: It was a very clever enzymatic approach. They didn't start with pi45hero. They started with liposomes that contained its precursor pi4p.
[00:05:41] Speaker B: And why was that important?
[00:05:43] Speaker A: Because pi4p doesn't bind FGF2. So it gave them a clean slate. It let them control exactly where and when the actual binding lipid appeared.
[00:05:53] Speaker B: Ah, very smart. It isolates the variable.
[00:05:56] Speaker A: Precisely. So then they added the enzyme that does the final conversion. A couple kinase called PIP5K1C, ATP and magnesium. But only to the solution on the outside of the liposomes.
[00:06:07] Speaker B: So the enzyme couldn't get inside.
[00:06:09] Speaker A: Correct. So it could only convert the PI 4p on the outer leaflet into PI 4.5p. And just like that, they generated a membrane where the high affinity lipid was stuck on the outside.
[00:06:18] Speaker B: They basically reversed the cell's natural setup in a totally controlled way.
[00:06:22] Speaker A: That's brilliant. But how do you prove it? How do you know the lipids are actually stuck on one side?
[00:06:26] Speaker B: They came up with this really elegant validation test. They figured if the PI 4.5p is really stuck on the outside, what happens if we break the vesicles apart and let them reform?
[00:06:34] Speaker A: The lipids would all mix up. Exactly. They'd randomize and you'd end up with a symmetric vesicle with the PI 450 diluted across both sides.
[00:06:42] Speaker B: And did that work?
[00:06:43] Speaker A: It worked perfectly. When they tested the new symmetric vesicles, the binding affinity for their test protein dropped significantly, almost in half.
[00:06:52] Speaker B: Which confirms their starting material really was asymmetric.
[00:06:55] Speaker A: It confirmed they had total control over the membrane's architecture. So.
[00:06:59] Speaker B: So with that figured out, they could move on to the functional tests. What were their main experimental setups?
[00:07:04] Speaker A: They used two main systems. First, the in vitro GUV system, where they just measured how long it took for a fluorescent dye to leak into.
[00:07:12] Speaker B: The vesicle, a sign that a pore has opened.
[00:07:14] Speaker A: A direct signal of pore formation. And the second test, which is maybe even more relevant was something they called the acute wave cellular assay.
[00:07:22] Speaker B: The acute wave assay. What's that?
[00:07:24] Speaker A: They did this in living cells. Chok1 cells. First they get the cells to make a bunch of FGF2. Then at time zero, they do a really harsh heparin wash to strip off.
[00:07:34] Speaker B: Any FGF2 that's already on the outside.
[00:07:37] Speaker A: Exactly. It gives them a clean surface. Then they just watch with a confocal microscope to see how fast new FGF2 reappears. That speed is the acute wave of secretion.
[00:07:48] Speaker B: Okay, so let's get to the payoff. Did changing the lipid distribution in the dish actually bridge that massive time gap?
[00:07:55] Speaker A: It gave really strong direct evidence. So, in these symmetric guvs, where PI 425p euro was on both sides, the average time to form a pore was about 95 minutes.
[00:08:05] Speaker B: Okay, an hour and a half.
[00:08:06] Speaker A: Now, in their asymmetric guv's, where the lipid was concentrated on one side.
[00:08:10] Speaker B: Yeah.
[00:08:10] Speaker A: That average time dropped to just 45 minutes.
[00:08:13] Speaker B: 45 minutes from 95.
[00:08:15] Speaker A: Yep.
[00:08:15] Speaker B: That's more than twice as fast. Just by rearranging the same pieces.
[00:08:18] Speaker A: Exactly. It shows that asymmetry isn't just a passive feature. It's an active kinetic regulator. It's physically lowering the energy barrier needed to make that pore.
[00:08:27] Speaker B: In one crucial detail. They checked the total efficiency.
[00:08:30] Speaker A: They did, and that's key. The total percentage of guv's that eventually formed a pore was the same in both cases.
[00:08:37] Speaker B: So it's not that symmetry makes it.
[00:08:39] Speaker A: Impossible, it just makes it incredibly slow. It proves asymmetry controls the speed, not the fundamental possibility. And the cell is all about speed.
[00:08:48] Speaker B: Okay, so let's take this back to the living cell. What happened in that acute wave assay when they deliberately broke the cell's natural asymmetry?
[00:08:55] Speaker A: This is where it gets really interesting. They flooded the outside of the cells with extra PI 4.5p, forcing it onto the outer leaflet. And the result was a profound inhibition.
[00:09:06] Speaker B: It blocked secretion almost completely.
[00:09:09] Speaker A: At the 30 minute mark, there was basically no acute wave of FGF2 reappearing on the surface compared to the control cells.
[00:09:15] Speaker B: So just having the lipid around isn't enough? It's like location is everything. But was this specific to pi4.5p, or would any lipid thrown on the outside cause a problem?
[00:09:24] Speaker A: Great question. They tested that when they added other acidic lipids that don't bind to FGS2, like phosphatidylserin or the precursor pi4p.
[00:09:33] Speaker B: Nothing happened.
[00:09:34] Speaker A: Absolutely nothing. No inhibitory effect. The block was specific to lipids that can actually interact with FGF2.
[00:09:40] Speaker B: What about other lipids that do bind FGF2?
[00:09:42] Speaker A: They found that other phosphonositides that are known to bind FGF2, like pi3.4p and pi345p, also blocks secretion when added externally.
[00:09:53] Speaker B: Which suggests the problem is neutralizing that very specific inner leaflet environment that FGF2 needs to cluster.
[00:10:00] Speaker A: Exactly. And the most compelling evidence comes from the recovery.
[00:10:03] Speaker B: The cell fixes itself.
[00:10:04] Speaker A: It does. They watched that external PI 445p over time, and after about 40 minutes, it started to disappear from the surface. The cell was actively removing it. And? And by the 60 minute mark, secretion.
[00:10:15] Speaker B: Was back to normal.
[00:10:16] Speaker A: Secretion efficiency was fully recovered. It's the smoking gun. Lose asymmetry, you block the function. Re establish asymmetry, you restore it.
[00:10:24] Speaker B: Let's zoom back in on the physics then, because this is the real discovery. How does this arrangement create so much speed? What is it about pi4.5p that lowers that energy barrier?
[00:10:35] Speaker A: It comes down to its shape. PI 4.5p is what biophysicists call a non bilayer lipid.
[00:10:40] Speaker B: Meaning it doesn't like to lie flat.
[00:10:42] Speaker A: Exactly. Unlike most lipids that form nice flat sheets, its geometry, it has a bulky head group at a specific charge. It actually prefers to be in high curvature structures like the inside of a pore.
[00:10:53] Speaker B: So it's already predisposed to bending the membrane.
[00:10:56] Speaker A: It is. So now imagine FGF2 clustering on the inner leaflet. It forcibly recruits about 20 of these non bilayer lipids into one tiny spot.
[00:11:05] Speaker B: And because they're all stuck on that.
[00:11:07] Speaker A: One inner side, it creates this immense local tension. It's like having a coiled spring pushing inward on just one side of a thin sheet of paper.
[00:11:14] Speaker B: That tension, plus the electrochemical gradient from having no PI 4 or 5 po on the other side just destabilizes the whole area.
[00:11:22] Speaker A: It compromises the bilayer's integrity to a point where forming that high curvature toroidal pore is actually the path of least resistance. It relieves the stress so the cell.
[00:11:31] Speaker B: Doesn'T have to spend a ton of energy forcing the pore open. The asymmetry has already primed the bomb. That explains the 200 millisecond timescale.
[00:11:39] Speaker A: It's a metabolically expensive system to maintain, for sure, but it enables this incredible speed when it's a matter of life or death, like in an immune response or cancer signaling.
[00:11:49] Speaker B: The paper also suggests there's a second rule for the asymmetry on the outside.
[00:11:53] Speaker A: Of the cell, right? It's also crucial for what happens after the protein gets out. Remember GPC1, the receptor on the outside that has to catch the FGF2?
The thinking is that if the outer leaflet were suddenly covered in PI, it would physically or electrochemically get in the way of GPC1 doing its job.
[00:12:12] Speaker B: So by keeping the outer surface clean of PI 4.5p, the cell ensures the capture machinery is ready and waiting to grab the protein the instant it emerges.
[00:12:21] Speaker A: Absolutely. The asymmetry solves two problems at once. It helps open the door, and it makes sure someone is there to greet the guest on the other side.
[00:12:28] Speaker B: And this all connects back to why cells spend so much energy, so much ATP, on enzymes like flippases and floppases.
[00:12:34] Speaker A: It does.
This study provides a beautiful functional reason for that massive energy investment.
It establishes pi405pouo asymmetry as a defined, validated and actively maintained biophysical switch. It's not just structure, it's a control mechanism.
[00:12:51] Speaker B: So, bringing it all together, what's the big take home message here?
[00:12:55] Speaker A: The ultra fast life critical secretion of proteins like FGF2 depends entirely on the precise non symmetric arrangement of pi4tu. You have to keep it on the inner leaflet to build up the tension needed for that rapid pore formation.
[00:13:07] Speaker B: Without that asymmetry, the process slows down by orders of magnitude.
[00:13:11] Speaker A: It's just a fantastic example of how a simple biophysical rule where you put your lipids can exert such profound control over a complex cellular process.
[00:13:19] Speaker B: And it leaves you with a pretty big question.
[00:13:21] Speaker A: It really does. If FGF2 secretion, which is so critical in things like aggressive cancer, is this finely tuned by lipid asymmetry, what other essential high speed cellular events, maybe something like synaptic zessical release or even viral entry, relying on the very same biophysical switch that cells fight so hard to maintain.
[00:13:39] Speaker B: This episode was based on an Open Access article under the CCBY 4.0 license. You can find a direct link to the paper and the license in our episode description if you enjoyed this, follow or subscribe in your podcast app and leave a five star rating. If you'd like to support our work, use the donation link in the description now. Stay with us for an original track created especially for this episode and inspired by the article you've just heard about. Thanks for listening and join us next time as we explore more science. Base by base.
[00:14:29] Speaker C: Light gathers under quiet skin Singles pull the shadows in, pressurizing where the shadow shapes align A hidden doorway Starts to shine in the places no one sees Pulse moves through the boundaries what was closed begins to bend Making room to cross again Land where the mem Rain opens wide Forces flow from either side Pattern twist to let it through Every barrier breaking new in the moment Tension feels face Something finds it forward for you.
Crowded spaces start to shift Tiny currents start to lift Clusters forming in a narrow line Searching for a path defined if the threshold learns to yield Every secret is revealed all the pressure turns to light Guiding shapes into the night.
Brain opens wide Forces flow from either side Paths twist to let it through Every barrier breaking new in the moment Tension fades Something finds it forward Way.
