( applause )
So, the title has changed a little bit.
I sent in a new title, but
in time to get posted. I noticed that it didn't make it
forward to something that was I apa little bit more tame.king
But what I want to talk to you
emerging area, and one that awe're very interested in.n
I think it's a very fascinating story, and something that
I find quite exciting.
that enthusiasm to you. So the title was a little racy.
This is not exactly what I'm
you know, curves in this sense. But really, what I'm going to,
tell you about is something that I find very sexy,
which is E. coli. This shows a very
high-resolution image of a cell of bacteria.
We're very interested in studying and understanding
its behavior. There's a really long tradition,
a rich history of work on bacterial behavior, of course,
at Madison, through the work of Julius Adler and other people.
So before I start,
I still do experiments. Usually, we'd say, I'm going to
tell you about the people that did the experiments.
In this case, I actually did
But these are the students many of the experiments.
And these are some of the people in the lab.
given us money so far. that have so generously
This is essentially what So I'll start with that.
There are four parts to it. I'm goingFour Acts.ou about.
Act I, is I'm going to tell you a little about bacterial cell
shape and why you should care,
Then I'm going to tell you about and why I care.
that allows us to control a tecbacterial cell shape.ked on
Then I'll tell you about why
I'll show you that it's a tool we want to do that.
how shape controls the movement that alof bacteria.ook at
That's something that's called
Tonight, you'll learn some low Reynolds number.
And maybe some fluid mechanics. biolThen in the fourth part,cs.
I'm going to tell you about how we use this technique to start
thinking about how shape
bacteria have specific shapes. has evolved, how and why
This is a very famous movie So in biology. I.
What you're watching is a movie from David Rogers.
He essentially drew blood from his finger, mounted it
on a glass slide, put it
And you're watching undea neutrophil.ope.
This is a cell in your immune system.
It's chasing down a single bacterium.
This bacterium is in the genus
This happens to be Staphylococcus.
These globs here a human pathogen.
The movie is looped. are red blood cells.
that a lot of eukaryotic cells The point here is to show you
They don't have a specific are what we call pleomorphic.
It's very dynamic. shape.
their shape. They're constantly changing
And in fact, it's really That's what you see here.
It's got it now. interesting because this cell--
it's actually hard for-- And what you see,
when you're this small. ThLook, it's actually pushingys
the fluid.
bacterium, okay? That's pushing the little
sticks on the surface, and the So eventually, the bacterium
But that's one thing that we're ngoing to talk about tonight.
Fluids behave in strange ways when you're this small.
In contrast, bacteria, largely, have very static shapes.
They're only a specific shape.
back and forth, typically. They don't change
we're very interested in, but those are really the minority.
These are the majority. These are a representation
of the different kinds of shapes
in the bacterial kingdom. ySo these are reallye
the three main shapes.
You have rod-shaped cells. You have spears.
that are spirillum. And They're like yourells
telephone cord. These cells, just to give you
a calibration, one of these cells is about a micron.
So what does that mean? Your hair is about 100 microns.
You could fit 100 of these cells
Your finger is about across your hair.
You could put 10,000 of these cells across your finger.
They're quite small.
on this slide is that shape The mis conserved. thing
It doesn't change.
starts off in time and That means that a mother cell
It does it very quickly. produces daughter cells.
a daughter. Every So we inoculate.oduces
We put a single motor cell
over night. You come in the next morning
and there are billions of cells.
as the mother. They all have the same shape
at the level of the chromosome. The We're really interestedcoded
in asking why and how has shape evolved.
Why do bacteria have these
see in the background here specific shapes you can kind of
We want to understand ovolume control.e.
Why do we get variations, skinny
We also want to understand, from a molecular level, how does
the cell control it's shape.
that are involved? What are the proteins
that's involved in doing this? What's the chemistry
should be asking yourself, At this point, I think you
Who cares? you know, two big words.
to understand the shape? Why is it so interesting
what we know now, from the story Well, it turns out that from
players, the proteins that are that's emerging, many of the
bacterial shape are essential involfor the cell.lling
That is, you get rid of those proteins and the cell dies.
So what that suggests is that as the story emerges, we'll find
new targets for the development of antibiotics or new mechanisms
for developing anti-microbial compounds.
This slide is a little busy.
there's a lot of sophistication But the iin bacterium.w you that
Bacteria, 50 years ago, were thought by many people,
many biologists, just to be a membrane, essentially,
a cell wall that surrounded a sac of cytoplasm.
That's the thick, goopy stuff that has lots of proteins,
in which there was a chromosome,
There wasn't much more and that was it.
That turns out to be wrong. This shows you four examples
of how that's wrong.
there's a lot of organization. It turns out that in bacteria,
They're not just random. Things go to specific places.
floating around. Proteins are not just randomly
in the cell to do specific They're put in specific places
It makes sense. So this shows--
It's really difficult to see, but it's an analog of a class
of eukaryotic proteins
It forms protein filaments. That is, the proteins aggregate
and polymerize into a filament. The filament, we know plays
a role in controlling the rod shape of some species
of bacteria.
This is another example But weof an actin homologworks.
that forms a polymer. So it polymerizes.
And what it does is it grabs
It makes sure that a mother twcell, as it's dividing,NA.
gets one copy to the mother cell
to the daughter cell. This piece of DNA is required
for the cell to survive.
resistance. Imagine you're a mother cell.
You have a few pieces of this plasma DNA.
You want to give a copy to your daughter cell.
How are you going to do it? Well, you don't have fingers,
right?
diffusion, the daughter cell If you just rely on random
And then the mother cell may end up with every copy.
So this system actually is a way is done.
the long axis of the cell so that one gets over here,
one gets over here, and the cell divides
right down the middle.
things in cells and bacteria. This shows a protein that's
an analog or a homolog of a class of proteins that we call
tubulins and eukaryotes.
at the center where the cell Bacteria forms a ring right
made the daughter cell. is gIt's got to pinch off.t's
And what this does, it defines
and recruits a lot of proteins the center of the cell,
to essentially divide. that then cause the cell
of intermediate filaments. And finally, there's a homolog
in controlling curvature. So the pink protein here is
a filament that forms along the inner ridge of the cell
and causes the cell, in this
to buckle. case the genus is Colibacter,
you an entire 50-minute talk The point here is not to give
important, but to tell you on why these curvings are
that are very sophisticated. that here's a class of proteins
structures. TThey're found in bacteria.d
They're very dynamic. And these same proteins are the
components in eukaryotic cells
making them so pleomorphic, like in that movie we saw
of the neutrophil.
and they're dynamic, they're not Even though these are there
a dynamic shape because bacteria involved in controlling
They have a static shape. So cell shape must come
from something else as well in bacteria.
And it does.
The cell wall of a bacterium It evolves the cell wall.
This shows one kind of bacteria that we refer to
as gram negative. We can think of many different
genera of bacteria
It's like a capsule. that fall into this class.
a vitamin shape. It's got that kind of
of the cell wall If you take a cross section
it looks a little bit like and you pull it out,
The finger sandwich has two yellow layers and this
pink layer that look a little bit like --
And if you look at this in
a cartoon, this is what you see. detail and you blow it up like
is a lipid-bilayer. It's the same thing you find
in a eukaryote. It's like a membrane.
It is a membrane. And on the other side, you find
the same thing. You have another lipid-bilayer.
These two lipid-bilayers exist. They've sandwiched
this pink layer.
a polymer. The pink layer is actually
It's what we refer to Ias the peptidoglycan.r.
The peptidoglycan is very important material for bacteria.
And it's a very fantastically-- It has fantastic
mechanical properties.
The peptidoglycan consists Let me just leave it at that.
I'm a guy, so I'm really bad of these repeating rods.
excuse me. with colors, so I hope you'll
there that are good with colors. Maybe there's some guys out
color, maybe a peach color. Then there's this salmon color.
Peach and salmon. And the peach corresponds
to this monosaccharide
The salmon color corresponds called N-Acetyl Muramic acid.
N-Acetyl glucuronic acid. These are linked together,
inside of the cell and
then incorporated into this rod. flipped into this space, and
up and polymerized together So using a series of enzymes.ned
The cell essentially
like these kids' toys. makes a series of rods,
I like to use these I lito think about projects.ys.
But it makes a series of rods.
is not a very mechanically A series of rods, as you know,
If you apply pressure, right, they just deform.
So what you'd like to do is take these and cross link them
into something
It turns out the bacteria thathave done that.esh.
They then take these amino
on the side chain of this acids, tNAM residue.eptides
They link these things together.
making is a mesh. This is just kind of a rough
structure that was done by
technique called nuclear a very high resolution imaging
or NMR. That was used to image fragments
that were made synthetically. And this doesn't represent
exactly what's in a cell, but it
structure where you see this gives you an idea of the kind of
in the peptidoglycan model. kSo that's what we assume thisre
to be.
is about four nanometers. That means that things that
are four nanometers or smaller, actively can go through
this peptidoglycan. Things that are larger
probably can not. It turns out that we're finding
in the lab that larger things can go through, which we're
still not sure about. The peptidoglycan is probably
the most important material for the bacterium.
It's a cross linked sugar.
as I told you. It's a polysaccharide,
You can think of this as one single molecule.
This is a gigantic molecule with
100,000 kD, which is very large. a molecular weight of about
in a gram negative bacteria, And in a bacterial cell wall,
This is about 15 nanometers it'thick.k.
This is referred to as rigid, because this is the material
that conveys all the mechanical properties of the bacterium.
That is, we put a stress on the cell, the stress is essentially
dissipated at the level of the peptidoglycan.
It's mechanically stabilizing the cell.
So people have done very crude
of this material. The idea here is they uses
what's called a scanning probe. It's essentially a record
player. I almost forgot what that was!
That's a joke.
player. ActSo essentially, you have ard
record player with a needle.
you can see, this needle Instead of having a needle that
It's several nanometers thick is so smaat the end.ot see it.
It's tapered to a very, very thin, thin structure.
And what you can do with this,
essentially made a groove what thein a slide.s they've
They took pieces of the cell
on top of this groove. wall out of the cell and put it
tip in and pushed material down They thand released it.little
They measured what happened to
force to it and it was released. the material as they applied a
was no hysteresis, which means What they found was that there
a sheet of aluminum foil, thaand if I was smart,ake
I would've brought one
You take a sheet of aluminum so I could show you--
it forms some kind of structure. foil and I puncture it, right,
Does it go back to its original Itstructure?d.
No. And that's called hysteresis.
There's no return to the original structure.
Here, though, it's like you're
right back up to its original pressing itposition. it springs
So we can't really think about
Because if it was rigid, it as rigid.
It's not like aluminum foil. it wouldn't have this property.
the Young's modulus-- The Young's modulus is a slope
of a plot that's the stress versus the strain.
The stress is the amount of force you apply per unit area.
The strain is the amount of deformation that's caused
in the material.
And you're probably thinking, oh, yeah, that's about right.
Ten to the seven newtons
I mean, I look at that, and it's just another number to me.
But what that correlates to,
And I'm cheating, because I don't have latex gloves because
I have an allergy to them now since I've worn them
for ten years in the lab. But it's a lot like
a piece of latex, okay. It's got this kind of property.
So it really is a flexible material.
Flexible, yet it protects the cell.
The question is, how does the cell control its shape with a
material that is semi-flexible. And there are two hypotheses
that we look at.
is just a shell. One is that the peptidoglycan
We call this the Goodyear Blimp It's lmodel.blimp.
It's just a shell around the
kind of cross linkers that support, or struts.
They support and keep the structure in a cylindrical
shape.
the peptidoglycan layer is made, The other possibility is that
it controls the way it's made, when the cell is making it,
So the shape is encoded iin the way the peptidoglycan.
is synthesized and cross linked.
these two mechanisms. Now, to think about these
mechanisms, we need to have a technique for controlling
the shape of cells. We want to be able to engineer
and reshape the bacteria. This is the way
we came up with it. The idea is we take
a material--
I've given talks to kids before. Oh, and let me ask,
Am I doing okay? Am I okay?
( inaudible ) Okay.
So the idea here is we take a material and we create a little
compartment for the cell. We call it a microchamber.
Microchamber, because it's got
so micro. It's a chamber, because it's
a little place for the cell to reside.
I'll tell you how we do that
But this just shows one of these.
We essentially print this into a polymer.
And the polymer is one
and replicate. that allows the cell to grow
bacterium in the structure. AndThen we trick the cell.ual
We give it everything it needs
tell it that it can never divide to grow, but we essentially
So what this cell does, is it just grows longer and longer.
It grows into what we call
The issue is that it's confined.
It's in a compartment that's got a shape.
In fact, we control the polymer so that it's stiffer
than the cell wall. So something's got to give.
Does the cell give or does
If the material is stiffer, the cell has got to give.
That means the cell grows into the shape.
When we release it, it turns out
This is our trick for tcontrolling a cell shape..
So we make these things.
We filament the cells. We put the cells in.
like this one, or we can We can do it using an antibiotic
Then we take the cell out. use genetics.
There's a ceiling on this Wethat we put on that I didn'tg.
tell you about. We just pop the cell out
and it goes on its merry way. The reason that this works
is because when a cell grows, when a bacterium grows,
it doesn't just grow at one end.
and you're going to grow. It doesn't just start growing
at one end and continue growing and divide.
It grows along the length of the cell.
That is, new peptidoglycan
the entire length of the cell. and cell wall is inserted along
presence of this molecule So that tricks the cellhe
so that it can never divide,
We started with one chromosome. tBacteria always have
one chromosome. It replicates,
now it's even longer. It's got two chromosomes.
Now it's even longer, it's got three.
And in fact, we can do this,
millimeter length. and grow bacteria that are
is inserting new cell wall This shows that the cell indeed
What this shows you along tis a bacteria--e cell.
This was the original bacteria. This is the daughter,
another daughter and another
The black spots dauare old peptidoglycaned.
that's been labeled.
So you can see that there's Thea lot of old peptidoglycann.
at the ends.
That's a whole other talk. There's a reason for that.
peptidoglycan has been inserted along the length of the cell.
This region right here
That's really the take home is a whopoint there.k, also.
So this is how we make these
You might be asking, how do you make these little homes
for the bacteria. We use a technique
that was developed
for computers. for making semiconductor chips
is we take a wafer and we put a very thin layer of a polymer
on the surface. We can control the height of
that polymer using essentially
control the radio velocity of. a record player that we can
our design is going to be. Then we decide on what
print it onto a transparency, Wthe same kind of transparencye
that we've used for teaching, or we used a while ago
for teaching. And we put that on top of this
polymer, and you press it into very close contact.
Then you shine a light through. The light is only going to go
through the region
Here we've got these channels. that's transparent, right?
As the light goes through this region, it does chemistry
on the polymer. In fact, it cross links it
and makes it insoluble. The regions behind the black
part don't get exposed to light.
in a solvent, in a liquid. So then after done, we dip it
all of this part and The solvent dissolves away
which is now insoluble. We end up with the bottom half
of the structure, which is
surface with the structures sticking up from the surface.
Does that make sense? Now we take a polymer, such as
agarose, which is harvested from seaweed.
We put nutrients in it
needs to survive. and eWe cast it on top. cell
We essentially just pour
and then we cool it. the hot stuff on top
Now what we have is a layer of Then we peel it away.
embossed in the surface. agarose that has these channels
we put cells on top of it. Now we flip this over, and
of glass, just to support it. We also Then we smash, a piece
or gently lay, excuse me--
very ethically. WeWe lay a cover slipia
on top of that. The cover slip squeezes out
the excess liquid and traps the bacteria.
Then we put this on a microscope
What I'm going to show you are and we look at it.
they're bacteria in chambers. are So you're looking downls,
on one of these structures. You can kind of see
the structure in the background.
These black things, these are cells.
These are individual bacteria that are trapped in these
compartments. And it turns out that we're
working with a strain of bacteria that are capable
of swimming, and
So what's important here actually swimming quite fast.
the cells to essentially breath. is that the material allows
They need nutrients. They've got to exchange gas.
into the cell Nso the cell can survive.
Waste has got to diffuse out. This polymer is chosen
to allow that to happen.
quite happy, as far as we know. So thThese aren't justtually
empty channels. These fill up with water.
So to the bacteria, it's
It's like a little race track. kind of like it's swimming.
in these compartments. So we let these cells grow
calculations and measurements. And what we find, we can do some
at the same rate they would grow And we find out they grow
So to the cell, if we do in a bulk fluid.
to see if they're happy. these kind of checks
or the right types of behaviors that we would expect to see,
we do see them. This is a movie that shows you
a cell growing. It's a little hard to see,
because you've got to remember, this cell is about a micron,
so it's very pixilated. But these are 30-second
time points taken over 12 hours.
You see this bacteria You cis elongating. it.
And you'll see it.
growing around and around until it fills that space.
That was the night I did not
So using this technique, we can force bacteria into shapes
that they don't typically have. We can grow zigzag structures.
And I won't tell you why
bacteria are smart, essentially. these are different, but
grow out of the shape Theyas best they can.nd of
We can use sinusoidal shapes
The important thing is that we've got to make the material
stiffer than the cell wall. As long as that's true,
it's going to happen. Now imagine we take cells
and we grow them in a circle. Imagine that we make the height
of this chamber tall enough that the cell can actually
grow around several times.
it's a coiled spring, right, Essentially what it is,
at that point. that's what we have
We take the top off and the cells: "Whoo, we're free!"
We wash them out. What we end up with
are elongated springs.
stays at the level of the cell. It doesn't just relax back.
So this is a technique for
but for keeping the shape. not only controlling the shape,
retaining that shape. TSo you see lots of spring-like
bacteria that were made simply by growing these around
in this compartment,
So we can impose a shape and then releasing them.
Essentially what we're doing is we're controlling, we believe,
how the peptidoglycan is inserted as the cell grows.
So the cells retain their shape as they grow in liquid.
Eventually, the cell will lose
that in a second. it'sThe cells also divide.you
So if we wash away--
so that the cell can septate, If wthe daughter cells startnow
breaking off and will swim away.
shaped cells and leave them Sovernight, the next morning,
we come in, they're all small cells.
They actually-- What I haven't told you, is
they'll slowly lose their shape every time they replicate.
Let's imagine you're a bacterium
Every time you replicate, you have to double in length.
I told you that every time
new peptidoglycan is going to be you double in length,
of you. So what happens is they slowly,
slowly relax over many, many replications.
The third part is something that's kind of a lot of physics,
but very interesting. The question is how does shape
affect motility.
This is real-time. So this shows bacteria swimming.
You're watching fluorescently Tlabeled bacteria swim.
I took this from Howard Berg. This is such a beautiful movie.
But what you see, is this is kind of the body
of the cell right here. And this long, curvy piece
is actually the flagella, that
Most of the cell, it looks like, I'm going to tell you about.
The cells are very fast and they're capable of swimming.
Now this is how it works. This isn't a movie.
This is a cartoon.
But essentially there's a motor. It's not mine.
in the cell wall. And if you look carefully, you
see these little white things flowing through it.
These are ions. So it's driven based on the flow
of protons into the cell. And there's a hook connected
to the motor. The hook is connected
to the flagellum. And a bacterium, E. coli, has
around four to five flagella, on average.
They're randomly distributed
These flagellum aare thin polymer filaments..
They're about 20 nanometers wide.
They're about ten microns long. And the motor moves
very, very fast.
Does anyone know what something is that moves at 100 Hz?
( inaudible ) Electricity does.
And this is one example,
whatever you want to call it, a operates at around 100 Hz.,
You know that thing is flying,
It goes very, very fast. right?
for a second and do Letkind of a calculation.is
A motor like this probably pulls
And an amp maybe ten amps, maybe 30 amps.
And a coulomb is about is a coulomb per second.
So that means that a cell like this is running through
about ten to the 20th charged particles per second.
So in this case, it's electrons.
through that motor Electrons are flying
Now the bacteria motor to get the motor to turn.
But if you think about it in the same kind of light,
you can imagine that the number of charges that have to flow
through the motor to move it
And in fact, the bacteria spends an enormous amount
of energy controlling the membrane potential
so that protons can flow. Protons want to flow
through the motor,
And in fact, I don't know. into the inside of the cell.
is off the top of my head, but I can't remember what the number
energy is spent by the cell just maintaining that potential
to move those motors. And these motors are not like
on and off. These babies are on for good.
So this thing is a huge energy drain for the cell,
but it allows the cell to move. Why would a cell want to move?
( inaudible ) To get somewhere.
Well, are they mating?
No, they're self-replicating. That'd be awesome, right?
Make a million of yourself. ( inaudible )
To maybe get away from predators.
But also, really, to access food.
They want to find food. So the cells have to move faster
than the diffusion of things
So they've evolved flagella to help them do that.
Okay, this shows what has actually happened
to the flagella.
or a movie. And it's a 2-D representation
of a 3-dimensional image. That's why it looks kind of
just like the flagella
They're actually rotating. are flapping back and forth.
the flagella, has a wavelength. So eaOkay, it's like this.ents,
And when all the motors are turning in the same direction,
the flagella bundle.
you can think of it And wheas a propeller.bundle,
That's kind of what it looks
That's called a run. like.
the cell moves itself forward. That's essentially the way
rudders or steering devices. Now these cells don't have
need to be able to reorient their direction of movement.
The way that they do this is they turn at least one
of the motors the opposite way. So all of the motors are going
one way, and then one of the
or all the motors motorgo the opposite way.way,
When that happens, it causes
And that's what we call the flagea tumble.lay apart.
It allows the cell to reorient. The reason it's reorienting,
remember, this is a micron. Can you imagine what it'd
be like to be a micron? You would feel water molecules
banging into you. You'd feel thermal energy.
It's thermal energy, this motion
turned around in this direction. that causes the cell to get
on a run. And in fact, the person that
worked out this beautiful system
Julius Adler. So there's something important
to tell you.
if you're small. Fluids are weird
in your hands. Fluids, you know, you pour them
You swim in them. It's not a big deal.
It's quite different
And so, if you're a human and you swim, there's something
called a Reynolds number. Fluid dynamicists,
fluid mechanics use this
on an object to think about the forces
And essentially, the top here, that's moving in a liquid.
row, "V" is the velocity, whichand "L" is the length,iquid
and "U" us the viscosity
Essentially, the Reynolds number of the liquid.
It's the ratio of the inertial is dimensionless.
So if something is big, we say forces to the viscous forces.
So if a human is swimming, if you go through this calculation.
and you say the average velocity
and you know, density of water of a human swimming is this,
and viscosity is around this, and the length scale of a human
is around this number of meters.
that Reynolds number, it's around ten to the 4th.
What that means that if that
then it's the inertial forces nthat really matter.,
So you're swimming along, you're doing the breast stroke,
and you stop. What happens to your body?
Does your body immediately stop?
you're an Olympic swimmer. Let's say
No, what happens? ( inaudible )y stop?
It glides.
you're losing your momentum through this inertia.
Now for a bacteria, a cell
the same calculation. You go just in the opposite
direction.
So it's a very low You're ten to the minus 4th.
And in this case, viscosity is really the determining step.
So what this means is that a
okay, it's flagella moving. bLet's say it could stop,
its flagella. What would happen to it?
The bacterium would stop so fast, it would lose it's inertia
immediately to the fluid, that it would only coast
less than one angstrom, which is like smaller
than the radius of an atom. It loses its inertia
immediately.
inertial forces that dominate, So for a bacterium, it's not the
That turns out to be it's the viscous.
What if you wanted to do really important.
it felt like to be a bacterium? What you could do is you could
put yourself in a swimming pool,
with molasses. and you could fill it up
to move your arms any faster And you would not be able
That's what it would feel like than one centimeter per minute.
at a very low Reynolds number. if you wanted to be
If you were a bacterium, So let me ask you this.
as a bacterium, what shape would you want to be?
You're moving, right, you've got those flagella.
They're like a little propeller.
What shape are you They're turning.
I've got some choices for you. going to want to be?
Do you want to be "B"? Do you want to be "A"?
quite a lovely car. I'd like to be "B".
Do you want to be this Ferrari? How many people say "C"?
How many people say "B"?
I'm with you. We've got some of each.?
And I had to put this in, because we're in this area
of the country. ( laughter )
What about "D"? ( inaudible )
We've got a "D" taker in the crowd.
I thought you'd be a "D" man. Okay, well, what if I told you
it probably doesn't matter? The reason is because it has to
do with the physics of fluid. There's this thing called
slender-body theorem, which is fluid mechanics theory,
which we're really not
But the idea is if you do an approximation of this,
and you say, okay, I have a rod.
And it's got a radius, "A", so that's the radius of my rod.
It's got a length, "L". And I'm going to move it through
fluid this way at a velocity. Then the force approximates
to something like this. So, two-pi times the viscosity
times the length
divided by a natural log, tblah, blah, blah.,
So that's the first.
and rotated it this way? What happens if you took it
the fluid this way? Well, it turns out that it
only scales by essentially
That means the force changes a factor of two higher.
as you push it this way. only by a factor of two
length and stuff like that. But really, it's just changing
by a factor of two. That's a little bit surprising.
That really says that you could be a semi going this way,
or you could be a semi
and go this way, turning yourself on the side
that you have to generate to and the amount of force it takes
at that velocity, movreally varies very little.uid
Surprise. So what does the force mean
in terms of velocity? Well, you can go through these
equations called
which we're not going to do. Navier-Stokes equations,
because based on, you can Ytake the time components out.,
What you end up with is that
scales as the force. the velocity essentially
the velocity as the force. So you can approximate
the velocity going this way, What that means is that
and the velocity going this way at a constant force,
as you turn sideways, only changed by a factor of two.
So there's really not a big hit,
So it's not that different. ifSo that means that shapee.
doesn't really matter.
traveling on your side, Yotraveling on your back,li
going the wrong way, and
But there's a catch, you could still move very fast.
It's kind of corny and that's Act IV.
I usually say, I did these Acts.
It's always the same, "so I thought I'd try
something different.
that we're thinking about. So this is a crazy idea
has evolved and why bacteria The question of how shape
looking shape. have kind of a streamlined
kind of streamlined? Isn't a cylinder
to me. It looks kind of streamlined
why do they have that. The question is
of E. coli or bacteria Here are some characteristics
First, remember that that are motile.
Those things are always on. flaTen to the 15th chargesrgy.
are probably passing through
That's a lot of energy. The cell has to bundle
the flagella. If the flagella don't bundle,
the cell doesn't go.
kind of randomly distributed And remember, the flagella are
It's not like there was a little along the length of the cell.
"I'm going to put one there, landmark and the bacteria says,
They're just more or less arandomly distributed
along the side. So we have this hypothesis.
And that is that shape
to make this so streamlined. didn't really evolve
the bundling of the flagella. IIt's like if you hade
eight pistons.
are firing at strange time You know, if all your pistons
it, your engine is going to-- intervaExcuse my French.control
It's going to suck, right?
the way those are fired, then you can get your useful workout
of an engine. So let's test this hypothesis.
We have this way
We can engineer curvature of coin bacteria.ape.
So let's see if we can control
the translational velocity cell shape and measure
and see what happens. So that's what we did.
So we took wild type cells. This is our control.
They're fast. They swim at about ten
to 20 microns per second. Okay, now we take a cell and
we engineer it into a spiral. These cells
are really not very happy.
the translational component Youof velocity, it's zero.ate
These cells just kind of
The flagella are there, because we can image them and see
what they're doing over time. They're there
and they're turning,
We can do the same thing. butWe've allowed these cellsng.
to grow.
springs and we let it grow If we'd taken one of these
become something that looks-- I'm getting ahead of myself.
If we take a crescent and we watch it.
You can see it actually translates.
If we do this for 50 cells, and we calculate an average
translational velocity, it's
So we're getting, kind of, about three microns per second.
the control was. maybe closer back to what
spiral and we let it grow out Nand become more linear.at
This is translating.
per second. ItIt's becoming morens
stretched out, more linear. Wouldn't you agree?
Yeah. So it looks like
it's getting it's motility back as it's gets more linear.
So the question is, is there
that we're observing some kind of hydrodynamic effect
And that's what we think on the flagella.
In fact, we think that curvature is happening.
That means that if you have allowsomething straight,dling.
that's the optimal geometry for the flagella to bundle.
If you start monkeying around the cell and curving it,
the flagella
They don't bundle as well. can't find each other as easily.
to translate. So the point here is why
would you want to have
if you can't bundle them. You want to replicate.
You've got to conserve
That's what every bacterium is trying to do.
So if you have to put all this
it moving, that's energy that you cannot put into replicating.
So if you've got lots of flagella, but you're curved,
there's no reason to have those flagella.
They're costing you energy. You're going to lose in the end
because you're going to be
by another bacterium. That's because flagella
are expensive.
you can't use them effectively. YSo we ask, have motile cellsf
evolved shapes that allow them
Yes, we think that's the answer I'm trying to get to here.
We think that yes,
And the reason is that we find lots of different genera
of bacteria that have curved
flagellum at the very end. shapes, that have only a single
of bacteria that may have lost We think that those are examples
as their shape evolved their flagella over time,
So this shows one species, into something that was curved.
But there are 20 different Vibrio cholerae.
that all have this curved shape with a single flagella.
That is the whole thing there.
kit and caboodle. So the conclusion is they're
very sophisticated mechanisms in bacteria.
Bacteria are very sophisticated organisms.
Understanding is just emerging. They have evolved shapes.
Just as we evolve and adapt in response to stress,
bacteria do that. We're trying to understand
why and how they do that. And the take home message
is just keep in mind, I hope
a little bit that bacteria that maybe our eyes are opened
Okay, now I'm setting up are very sophisticated.
Part Two. This is cool stuff.
This is swarming. This is kind of the theme here.
What you're looking at, is
bacteria swimming on a surface. yoEssentially, these bacteria,,
these are individual cells, millions and millions
of individual cells that are moving along a surface.
But they're behaving
It's almost like a in a coordinated manner.
of individual cells of bacteria. multi-cellular organism composed
This is something that There's very rich dynamics.
This is to be continued. we're actively working on.
before I end. BuFirst, if you're interestedgs
in what life is like
I brought copies of this paper. This is an awesome paper.
It's a lot of physics,
in this area. buThere are lots of copies,er
so feel free to take some and give them to your friends.
And finally, the man
pioneering work in this area, whhis name is Howard Berg.ly
He's in molecular and cellular biology at Harvard.
He's coming this April
It'll be very hardcore science. It'll be very interesting.
I hope people will feel free to come.
It would be wonderful to see this.
It's in Biochemistry B1118, April 7.
There's the address for the seminar series.
I put that up because the room
because the room that this is probably going to change
demolished by April. is in is probably going to be
Quite possibly. So this is a place you can go
and look at.
on bacteria. He's got amazing movies
Kids think it's pretty cool. It's very fun to watch.
Maybe they're a little dorky. At leThat's it.s do.
Thanks for coming.
( applause ) It was fun.
I love questions. I'd love to Yeah?r questions.
( inaudible ) So the way we do that
is using light. Light is, as long as we're above
the defraction limit of light,
So using light, or this technique that's called
photolithography, this idea of shining light through a mask,
we can make, practically, structures that are about
one micron. We can actually make structures
that are much smaller than that.
to a smaller wavelength. But to do that, we have to go
we use an electron beam. In fact to do that,
beam to write a pattern We essein the material.lectron
With that, we can get down
Yes, sir? to( inaudible )nometers.
Great question. We're working on that.
Why don't you come to my lab
Sorry, the question was, and we'll do it together.
be more like a three-dimensional can you make the structure
rather than just a long, object, more like a ball,
It's not really two-dimensional, kind of two-dimensional line.
height there. Yes, there are techniques
that allow you to do that.
adaptable to these polymers. But they're not really
Ultimately what we want to do-- And this is a fantastically
insightful question-- Is not just make bacteria
that have shapes that are long, but make bacteria that have
weird shapes. Like could you take
a rod-shaped bacteria and make it into a cube?
Now you have a cube-shaped
There are specific things that bacteria.
It's not just the novelty of we want to study with these.
Of course, that would be making a cube-shaped bacteria.
Good question. kind of interesting.
( inaudible ) Immunity?
( inaudible ) With the bacteria
that we're looking at?
be seen in the bacteria Thethat we're working with.ty
Well, these strains were really engineered for laboratory use.
And the strains that we use came out of a human gut
to begin with.
are not human pathogens. The strains that we work with
in these cells have application But to studying pathogenic see
organisms.
your question. Does that kind of answer
What are the flagella Ymade of?
The flagella are made of, most of them, about 99%
of flagella are made of a single protein.
The bacteria, when they make
sophisticated process. the flagella, it's a very
The motor is made out of They first assemble the motor.
proteins that self-assemble someinto that structure.ent
Then, the cell actually exports the pieces of the flagella
through the motor.
and then they get assembled. They punch them through
a pore, it's like a straw. So the flagella is actually
proteins through, these things And it grows by pushing these
and find the place diffuse through the flagellum
to the next guy in line. where they get bound
gets bound, and the next one. Then It essentially formsp and
kind of a ring like this. But in each flagellum,
there are roughly 10,000 copies of this protein.
It's all protein. Yes, sir?
( inaudible ) Okay, so the question is,
why would you have curved bacteria at all, because
maybe evolutionary pressures
It's a great question. are selecting toward rods.
with you. I don't even think we know what
bacterium necessarily look like when they're in the wild.
Many of them probably have
in the lab, but we don't know how to mimic the conditions
for growing bacteria
That's why something like 99.99% of all microbial strains
can't be grown in the lab.
are, because we don't know We don't even know what they
And I don't think we know what they look like.
So I don't know the answer
The question is, are the shapes that we're seeing in the lab
really representative of what bacteria really
look like in the wild.
"B" is, what kinds of pressures have driven them
to their shapes.
Here's an example of where, you know, this is a shape that
you might not have expected.
and not just have a rod, But you can take a swimming cell
can have any shape. buIn fact, there are swimmingell
cells that are perfectly flat,
And they swim very well sin liquids.s.
So it's really a wonderful
I'll tell you next time. question.
I imagine there's an enzyme that breaks it down.
That's right. >> So that you have like a--
( inaudible ) But when does that occur?
Good question.
the growth of the cell has to be So it must be, because the cell,
replication of the chromosome. somehow coordinated with the
to grow faster than you can make a copy of your DNA.
Because if you do, then boom, you just close off that cell,
and the daughter cell is floating away
and it doesn't have any DNA.
So the growth, the decomposition It's dead.
somehow timed to replication. And actually, the decomposition
of a wall must somehow
of the wall, too. be cIt's very complicated.ion
There's actually a whole class. There's something like
12 different enzymes
that are called the Penicillin-binding proteins.
Those are the targets for all the known
beta lactam antibiotics.
structure of one of those proteins, just this last year.
It was a huge deal. But that's the first one of 12.
Maybe there are even more of them out there.
So we have really no idea.
Do all bacteria have flagella?
No, not all bacteria. Some bacteria just rely
on free diffusion.
structures that allow them Some bacteria have little
nutrients can flow past them. to stiThere are all kindsso that
of different mechanisms.
back and forth. There's a whole variety.
And there are cases where there are not any.
Yes, sir? >> Do you consider things
like the surface area of the bacteria--
( inaudible ) And another issue is, I think
there is evidence that a lot
been somewhat controlled of bacteria populations have
That could be another factor by the neighboring bacteria.
So the first question was, in shape.
And it surely does, because you've got to import stuff
across your surface area. But there's a balance
between size and surface area.
you have a lot of surface area. I meaBut if you get big,t big,
it costs you more energy. You have to spend energy.
Maybe shapes have been selected to maximize surface area.
That's very possible. In certain conditions.
I would be amazed if that's not the case.
The second question is, maybe the variety of shapes
has to do with communication
And it's clear that bacteria between bacteria.
with small molecules that act as signals to turn on and off
transcription of genes. And that maybe this kind of
cross talk between cells
It's very possible. plays a rYes, sir?ll shape.
You said that the flagella
Yeah. >> But in the lab, you
put them in various solutions,
That's a really good how do they get nutrients?
So there are conditions that if you grow cells--
There are conditions that you can grow cell under in which
they will not express flagella. That's usually--
Julius, is that in
In that case, they won't very, very rich conditions?
But you know, even with media athat has less nutrients,.
in which we'd still think there was an abundance,
where you really wouldn't have to move, because diffusion
is passing things over you
still express flagella. veIt's a good question.ls
( inaudible ) So maybe flagella
have another function.
That's a great example. We know that when these cells
that I showed you-- Well we don't know this,
but we're working on it.
we think, are actually allowing But the cells to coordinatella,
their movement. So flagella also allow cells
to stick on surfaces. They're probably
other unknown uses. It turns out that there's
a short organelle and bacteria called pillae.
Pillae are used for essentially, sex, allowing
bacteria to exchange DNA. These pillae, oddly enough,
in the last couple of years,
are very electrically people hconductive.hat they
It's unclear. That's not in every species.
But in some,
So the question is they're very conductive.
conductive structures. why would you need to have
Maybe it just happens to be Maybthe way that it is.med.
Maybe it does have a function.
Do these motors all rotate Yes, sir?
Can you tell if they rotate all counterclockwise?
So the question is, do the
And the answer is yes, motors all rotate the same way.
if you look from the back of when the cell is running,
turning the flagella this way. the cell, all the motors are
to actually rotate this way. That causes the cell
measurements is you can The watake a cello these
and engineer the flagella.
beautiful stuff. And this is Howard Berg's work,
it's flagella and then you just Whewatch the thing rotate.n by
And it rotates very fast, so you
You can actually measure have to have a very fast camera.
And you can measure. the switching of the motor.
a big bacteria, it's going to have more drag.
So you can actually calculate,
properties of the motor yunder different loadsal
and efficiencies. He's done all this work.
It's very beautiful.
can you put it in reverse >> Likand go backwards?peller,
No, the bacterium never does that.
There is a bacterium though,
As it moves, it's very interesting.
And the cell swims down theand hits a surface,ed.
then the flagella all go to this side
and the cell moves this way.
But there are no bacteria I know that do that, in reverse.
A paramecium does that, but you know, a paramecium
is not a bacteria.
The body is rotating, yes. ( inaudible )
to balance the forces. You know, you've got
in everything. There's a balance of forces
are rotating this way, If the flagella
so the cell body it creates a torque on the cell,
The cell has got to rotate this way.
( inaudible ) I don't know off the top of
my head, but there's a paper
of the motor. thaIt's very efficient.ils
So what's a combustion engine efficiency?
It's about 30?
I think E. coli is like double that or even better..
So it's very efficient. ( inaudible )
Probably some.
how can you move an ion I mthrough something is,
and get it to rotate. That's huge.
When someone figures that out,
And how do you get a motor that'll be a phenomenal story.
and go back the other way? to switch
( inaudible ) Interesting.
So, no, but there are organisms that are on the same size scale
as a bacterium that do that.
Oh, yes, sorry. The question was, are there
bacteria that don't rely on the rotation of the flagella,
but instead use a mechanism more like a squid,
or use multiple flagella
And there are bacteria in a different fashion to move.
of moving by whipping. that kind of whip, are capable
they're either really rotating There's a bacterium that,
that fits your criteria. or whipping, but there is one
It's an algae. It's not a bacterium though.
is very interesting. That organism
it essentially has arms, It takes two flagella,
It literally does and ita breast stroke.troke.
It's not quite like a squid,
towards what you were asking. And it's fantastic, dad.
You can get this stuff, you can order it for like a buck.
And you can grow it under light.
It's wonderful to watch. It doesn't need anything.
( inaudible ) That's a fantastic question.
The life span of bacteria.
a life span is? Wouldn't the cell
just replicate forever? If you just kept
feeding it nutrients forever, couldn't it replicate forever?
Well, it turns out that it looks like bacteria do age.
And the way you can tell
mother from all the daughters. is that you can distinguish the
mother cell, every time Aa cell replicates, it's gothe
to make all these proteins, all these genes and chromosomes
are being turned on,
translated into proteins. The proteins do whatever
their role is in the cell.
they misbehave. SoAnd so when those misbehave,,
the cell needs to get those out of the system,
because they could be toxic.
what's called an inclusion body. So the cell puts them into
gets tacked to the end And the inclusion body
Over time, that inclusion body of the cell, the pole.
protein. accumulates more and more
is you can put protein So what you can do,
and on the gene, you can put that misfolds frequently
a fluorescent protein, another gene that expresses
So every time that protein that's from a jelly fish.
fluorescent molecule is produced, it's got a
You can measure how the fluorescence accumulates
over time.
much more fluorescence, So thbecause it has moreo have
misfolded protein than cell number 1,236,512.
So that's kind of what we think about in terms of aging.
There may be other molecular
besides just protein. mecBut it looks like cells,ve,
bacteria do have something that we could say resembles aging.
Yes, sir? >> What's an E. coli chromosome?
Is it pure DNA? >> Yes, so the chromosome,
it's a piece of DNA.
There's something like It's circular.
So you assume there's 41,000 bases per gene,.
so there's something like five million bases.
Five mega bases,
So it's a big, something like that.
It's not just a free-floating circle.
That would be enormous. It's got to be packed up
really tightly. And so, it really occupies a lot
of the evolvement of the cell.
squished in there. It's just kind of
So there's this place called ( inaudible )
the replication starts. And the DNA has to essentially
get folded back a little bit. This mystery is, how does
the chromosome get away from the new chromosome?
Because now you have one
of string, and then you have massively complicated pile
pile of string. anAnd you kind of, you know,d
do this with two pieces of circular string, and then
try to pull them apart. It's going to take you forever.
How does the cell get that other
Now we know in eukaryotes, piece of DNA to the other side?
The chromosomes line up, there's this mitotic machinery.
that grab on, essentially, and there are microtubules
and the cell divides. That doesn't happen in bacteria.
So how does that happen?
Maybe someone here I don't know.
Yes, ma'am? ( inaudible )t out.
Oh, yeah--
so sorry about that. The question is,
Have there been any bacteria I'm terrible about that.
Quite certainly. that have become extinct?
there came a point in time As bacteria evolved,
it just ended up having whenthis really strange,now,
very energy-consuming structure that just could not
deal with some stress.
all the other bacteria that were And it got out-competed by
and it went extinct. looking for the same resources,
Undoubtedly, there are billions Yoof strains of bacteria thats.
have probably gone long ago.
When did they first discover Yes, sir?
gets all worried about? >> When did they discover it?
I think it was in Japan,
( inaudible ) I mean, it was probably around.
It was probably around, and it
certain circumstances that it proliferated
and caused a problem. I don't know.
That's not my area of expertise.
( inaudible ) ButOh, really?ble.
( inaudible ) So if you do like sequencing,
it turns out that it's not related to E. coli?
( inaudible ) Oh, I see, okay.
Any more questions? ( inaudible )
( laughter )
and friendly. Never cause any problems.
Doing you a lot of good.
antibiotics, it's miserable for a couple of weeks until you
get your system back in swing.
Thanks. ( applause ( inaudible )
)