I am delighted to welcome you all to this lecture
of my friends, and I wish I could say
colleague, but not really. But my friend John
Cloudy. John and I go back many years.
I figured how many as we've known each other,
not as long as somebody in the audience I know
, but still a long time. He was at
Cornell and then for many years was actually a team
there and then moved to Harvard 10 years ago,
10 years ago. And now, as you can
see on the screen, Shine and a professor at
Harvard Medical School also has a joint appointment of the
chemistry at Harvard and actually teaches in that department.
So he's a true interdisciplinary, actually got him first
because his expertise and actually crystallography. And then he
really moved from that and into a biological chemistry in
a big way. Just a few brief introductions to
him. Uh huh. He is co director of
the infectious Disease program of the Road Institute and,
as I said, professor in biological chemistry and molecular
Pharmacology, part of medical school. He graduated from
Yale, PhD at Harvard, both in chemistry,
and then went to Iowa State and then moved to
Cornell when he was about 20 years before moving to
Harvard. His number Awards. The Sun Foundation Fellowship
, Camille Henry J. Fox Foundation, Guggenheim Memorial
Foundation Fellowship. He has the Bonus Culture Award of
the American Cancer Society. Cope awarded American Chemical Society
, and I kind of like about it. This
is the research achievement award for the American Society of
Competency, which is the society most closely involved.
Actually, was denominator for him for that award,
So I take pleasure in fact, received, He
said. What was your strategy worked or something?
So it's a real pleasure to invite John, uh
, to speak to us on learning chemistry back,
please. Welcome. So thank you, David.
Both for the invitation and the awards of some years
ago. And and the kind introduction, um,
it really is a is a treat to be here
and have a chance to tell you about some of
the things we're doing really more directed from drug discovery
. Point of view towards very, very early days
. So let me just dive in. Um,
so we tend to think of farming no farmers in
their fields and doing stuff, and this is a
very famous quote from Schultz about farming. It's as
agriculture is a specialized form of symbiosis that is known
to have evolved in only four animal groups. Humans
bark beetles, termites and ants. You didn't invite
me here to hearing anything about human farming, because
in spite of the years in Iowa, I don't
know anything about that. So I'm gonna talk about
bark beetles, termites and ants. And what's sobering
is they've been farming for much longer than humans have
been farming. They've They've been farming for much longer
than their humans. They've been farming for much longer
than there were primates. So these agricultural systems go
back a long time. And what he what the
authors meant by a specialized form of symbiosis was the
following. They were using symbiosis in the way a
real biologist would, meaning they were telling you just
about how to things were co located so that the
farmer and the crop obviously we're together, and that
makes sense. So just the fact that they're in
the same neighborhood makes it a symbiosis, and it's
specialized in that you never find the farmers without the
crop. You never find the crop without the farmers
. The farmers can't live without the crop. The
crop can't live without the farmers, and so they
both benefit. So it's a mutual is, um
so that's the specialization. So let me let me
introduce it. What? What they mean by a
farming system and symbiosis? Easier to understand with an
example. So So the example is the Southern pine
beetle because we're here, and and what this shows
is sort of the rear end of, uh of
a female southern pine beetle trying to get into a
pine tree. And the pine is trying to repel
. That's the the Beetle. And the pine tree
is fighting back in the way. Pine trees typically
would fight back there, pouring out a resin,
all kinds of noxious, sticky stuff. The female
keeps trying to get in. Once she starts making
progress, she sends out a small molecule, uh
, and that calls all the other females in the
area called an aggregation pheromone that what takes place next
is called a mass attack of females. The pine
tree succumbs, Uh, at that point, the
the males show up. Uh, there's breeding.
The females go into the tree. They build they
build a gallery underneath the bark of the tree.
Uh, they lay their fertilized eggs in the gallery
, Um, and then that's that. Now the
larvae actually don't eat the tree, which is what
was believed for many years. The larvae eat a
fungus that eats the tree. Trees are are hard
to digest, and many fungi have specialized in integrating
trees. And so the female beetle has a specialised
anatomical compartment. And as she goes through the tree
, uh, she's releasing fungal spores from this this
compartment there seating, uh, seating the track she's
been in, and, uh, and they start
chewing up the tree, and that's where where the
larvae get all of their nutrition. So that's,
um, that's the farming system. So that's the
Beatles are the farmers. The fungus is the crop
, and when the when the young larvae they go
through all their developmental stages in the tree, when
they leave, they scoop up the fungus and carry
it to the next tree. Okay, so those
are the main players. They're not the only players
as it turns out there. Um, so that's
that's sorry. That's what I just just told you
. They're not they're not the only players sort of
hitchhiking. A ride on the beetle is another fungus
, sometimes called the blue stain fungus, and it's
obvious Toma minus and, uh, and what the
blue stain fungus does is probably kills the tree.
But it also is a very fast growing, aggressive
fungus, and it can overcome the food fungus,
the crop fungus that the the Beetle that the Beetle
relies on. So in that sense, this is
a threat to the entire system, and, uh
, and then so that was known for a long
time. What was known relatively recently say, about
five five or so years ago was that if you
look at the fungus in the gallery or you grew
the fungus in the lab, it looks like that
. But if you look in this anatomical compartment and
the female fungus, sort of where the head joins
the thorax if you look there, what you saw
was a very different picture and as discovered by care
collapsing and what that different picture showed was that there
was certainly fungi in this compartment, but probably more
fun. There were probably more bacteria act into my
seat bacteria than there were fungi. And so the
question then came up. What are what are all
these acts you know, bacteria doing there? And
why do you find them in every in every female
? So that was That was the question, and
that was worked out by Care and Cameron Curry and
some others. And so the crucial bit of experimentation
is shown here, So the bacteria is a lovely
bacteria. It's a red bacteria and makes these white
spores, and so and so you can see you
can see the bacteria, which is called Southern Pine
Beetle 74 There. Now, if you grow it
with the with the food fungus, it grows sort
of like that. And if you grow it with
this of the fungal predator, uh, grows sort
of like that and what that means. What that
was taken to mean is that this this bacteria is
making a small molecule that's diffusing through the agar,
and it's suppressing the growth of the predator much more
than it's suppressing the growth of the fungal crop.
So it's making a selectively toxic fungicide. Okay,
so it's making a selective, selective fungicide. And
so once you have something like that, it's ideally
set up for chemistry. You have a producer,
you have an essay and so you can find out
what the molecule is. And, uh, the
molecule turned out to be turned out to be this
molecule, which we gave the name Mike and Jamison
. Because my kanji, um is the name for
this special anatomical compartment and the female. Now,
I won't say much about this molecule. Um uh
, as many of you will imagine, it's touchy
because conjugated uh, Pauline's are touchy peroxide czar.
Touchy. That molecule is touchy. And I suppose
I should apologize for showing it at something with drug
discovery in the title, because you'll probably never in
this room seal less likely less drug like molecules ever
shown. Nevertheless, it actually does what it's supposed
to do quite effectively. It suppresses the growth of
a predator and spares, um, spares the mutual
ist. And so what? What it introduced into
this idea of a farming symbiosis was a new idea
that it wasn't a binary symbiosis between the farmer and
the crop. But there was there was a bacteria
that came into play, and the role of the
bacteria was to suppress the parasites and pathogens. So
this will become a familiar picture over the next over
the next several slides. So that was the paper
announcing this. And so that's the fun. So
the names are all sort of here. So this
is the strategy, and this is deals more with
the tactics. Uh, but the point of view
of chemistry, it's a way, uh, it's
a way to find, um, small molecules.
And once you then recognize what? What? What's
going on? Then? You have, if you
will, almost an algorithmic way to find other Find
other examples. So you say. Okay, well
, that's true. The Southern pine beetle. What
about the European park beetle? So that's the European
bark beetle. That was the Beetle, actually,
where the mic and gene was discovered. And you
can find these molecules there which had exactly the same
role that they would. They would suppress a fungal
pathogen and spare the fungal crop. And so all
the names changed. But, um, and the
molecules change. And so, if we didn't know
about cyclo hexane meid, we could have learned about
it from the European bark beetle and the bacterial Symbian
that it carried. Okay, so That's the So
that's how things work. Work for beetles. And
there are like 200 genera of these beetles are over
6000 species, right? And I think they all
follow essentially the same strategy. So what about ants
? What does anti agriculture look like? And agriculture
looks something like this. So if you've ever been
in the tropics, the new World, tropics,
anywhere, and say central or South America likely to
have seen these sort of trails running through the jungles
or parks or agricultural lands and very often they're mostly
active at night. But even during the day,
you can see scurrying along these trails ants, and
they're carrying big chunks of leaves the way more than
they do. They're running back and forth, back
and forth, back and forth, and what they're
running is between a tree or a bush or something
and a hole in the ground and, uh and
, uh, and they go down into this hole
in the ground. Now, if you look in
the hole in the ground, it goes down for
maybe 10 or 15 ft. Then it opens up
and you can find the largest ant nests are probably
about the size of this stage can contain millions of
ants They can last for a decade. And what
you find is a football sized colony after football sized
colony connected by little little tunnels. And what's growing
in these little football sized shapes is a fungus.
Okay, so that's the fungus, the white stuff
. And, uh, and those are the ants
tending, tending the fungus in the garden. So
what they do is they get down the leaves.
Uh, they chew it up a bit and they
feed it to the fungus and then they the fungus
. Mm. And so they become then the major
herbivores throughout throughout the tropics. And a lot of
plant material goes down underground. Now, what was
so that was known since maybe the late 18 hundreds
? Um, also since the late 18 hundreds,
it was known that the ants typically looked funny,
right? And what do I mean by funny?
They might have these white patches on them. You
can you can see there. Or they might look
like they had been dipped in powdered sugar. They
have this white powder all over them, and it
was known what that was from the late 18 hundreds
. Up until about, I don't know, 2005
or six or so, and that was it was
wax because insects make wax. So what else would
white stuff on an incentive be? But wax turns
out it's bacteria and that the ants are very highly
specialized. They have what are called crypts, but
they're really little bacterial culturing chambers. And the bacteria
grow in these in these crypts. And that was
a discovery by Cameron. Um, and actually,
this was the beginning of our of our collaborative efforts
. And that shows an ant work around, uh
, with 22 of these scripts, which you can
see by the white stuff. And that's a microscopic
view where you can see the bacteria growing in,
uh, in the crypt. And then you would
say, Well, uh, what what are the
bacteria doing? But you already know what the bacteria
are doing. So these ant gardens, their fungal
gardens, are attacked by a predator fungus and called
Escobar pepsis. And, uh, and the predator
fungus can overwhelm the food fungus and the bacteria,
which are a different kind of acting on my seat
. All belong to the pseudo no cardio genus.
But the bacteria make small molecules that suppress that suppress
this fungus. So, um, so that's very
understanding. And the way it works is these funds
. These this kind of fungus tends to be a
slow growing fungus, sort of related to mushrooms.
Okay. And this kind of fungus tends to be
more like the black stuff that grows on your shower
. And ah, and and and so they're very
different kinds. And and it grows rapidly, and
that grows slowly. So those are the different kinds
of fungi. So But again, then what you
should see is that it's the same strategy farmer crop
uh, bacteria, uh, protecting, making small
molecules that protect from, uh, from these predators
. And so, if you say well, what
kind of small molecules to the pseudo no card you
make. Here's Here's an example. So this is
, um, the first of these that we identified
with Cameron and it's, um it's a molecule called
dented remission after after the kind of kind of ant
that makes it. And what you can see is
that it's too big to put and little space that
I allowed for molecules the general thing. But If
you look at it more carefully, you'd say,
Well, what I see is that it's a It's
a peptide. At least there's a peptide ring here
. There's some unusual amino acids. Uh, then
there's some clearly some post translational kinds of modifications or
some some modifications to what? Our standard amino acids
. And then there's a bit up here that actually
looks like it comes from a Pawlicki tied synthesis pathway
. So it's one of these hybrid, um,
in RPS PKs pathways that generates that generates. This
, uh, generates this molecule. Okay, so
, um, the short story, though, is
that the strategy for ants looks a lot like the
strategy, uh, for bark beetles. So what
about what about termites? So termites are have exactly
the same ecological niche in Africa and the old world
tropics, as the ants have the new world tropics
. So these are the kinds of termites that build
these huge Coloman er mounds and they go out which
can last for years and contain hundreds of thousands of
termites. And and what they do is they go
out and they forage for plant food. They bring
, they bring the plant food back, they sort
of chew it up and mix it with spores of
the fungus and build these beautiful sort of free form
structures look like modern art things called combs, which
are made up of this massive plant material and fungal
spores. And then the fungi start growing. So
that's what this is. And then the fungi start
growing out of that, and the termites eat the
fungi. Okay, so it's exactly if you will
the same the same, the same sort of system
up here. And once again, there are fungal
parasites that try to invade this system. Um,
and if you look at the termites they're carrying bacteria
usually stripped mice these kinds of bacteria and they make
small molecules. Here are a couple of them that
that preferentially kill. Those are fungi. And not
not those fungi. Okay, so that's sort of
the, uh, the end of the first part
of the of the talk. And what it establishes
is that this thing right, you can find everywhere
, right? So I already told you that there
are something like 6000 species of beetles that do this
, and that's been going on for 100 and 50
160 million years. Ants are much younger. It's
only about 50 or 60 million years. It started
in the Amazon with one, and now it's about
over 200 species. The termites are roughly the same
started about maybe 70 80 million years ago, and
there are about two or three, probably 300 species
of termites that do this. So it's an extraordinarily
successful strategy, and presumably these three different, um
, insects all converged on it. And it's a
strategy that involves a fungus and insect and the bacteria
so it involves. It's it's a trilateral symbiosis and
it's three of the five kingdoms of life are mixed
into this. And you would say this point.
OK, I got it. You don't have to
tell us that anymore. Why would you keep doing
it right? And you would keep doing it because
, well, this is the same. This is
always different. And if you're a chemist, right
and supposed to be looking for new small molecules and
you want to find things, let's say you wanted
to find anti fungal agents. That's a pressing need
, that this would be a great place to look
, and so and so that's what this article says
. But then I can't just tell you that I
should probably give you some examples of what you can
find and what questions you can try to answer by
looking by looking at this. Okay, so here
are, I think, some interesting structures. So
that's a cute one. It's a sort of a
volatile molecule. Um, suppresses one fungus attracts other
fungi. They're terrible to work with. Molecules like
this you can see there. Macros. I click
lac tams, uh, with a lot of conjugated
double bonds. There's sort of some interesting looking looking
things here, So those are all new molecules you
can find relatively quickly because this is sort of an
unexplored source. Here are a couple of connoisseur structures
. This one is interesting because, you know,
if you look at it, you say, Well
, you know, it's some fatty acid in here
that got mixed up with, um with some amino
acids. But in fact, how you form that
sulfur carbon bond is interesting. It's not obvious how
you go about doing it. And here's another one
you can say of those nasty macros. I click
clack tams, and that's that's interesting. What's interesting
about that pair is there actually is members of each
other. Okay, so this can undergo what's formally
. It deals all the reaction to pinch off all
of those rings. And this can add to hear
a conjugate addition to build to build that ring and
which one of these you get depends on what you
feed the bacteria. So you feed it one thing
and you get one. You feed it another thing
, you get something else. And so how is
that regulated? And how does, uh and how
does that and how does that go on? So
I won't give you any more examples along this vein
, But trust me, you can find lots of
lots of small molecules this way. Now it's one
thing, uh, to find interesting small molecules that
chemists like that. But it's that's it's not the
most marketable of skills, always and so and so
so that that raises the question. Why would anyone
pay you to do this kind kind of work?
And so the question is, could it be a
path to new drugs? And so, um,
this is from nature. This was a report.
Our report from a meeting in 2000 and nine,
when there was absolutely no evidence that it would be
true. There's no doubt that there's any evidence now
, but this was just way premature. And just
say fungus farmers show way to new drugs, which
could be could be true. Here's Here's a more
recent one about why you'd like to do it.
So this is a very interesting article from a year
ago. Emerging fungal threats to animal plant and ecosystem
health. And what it does is points out that
that there's considerable evidence that, uh, pathogenic fungi
could be making, which we don't worry about much
are are making a comeback. And they're threatening animals
there, certainly threatening plants. We've seen new,
um, fungal pathogens appear and they're threatening people.
So the fact that I learned from this that I
didn't know is that a few, uh, that
fungal diseases will kill more people. Now, this
year the malaria and TB combined. Okay, so
and so it's a serious unmet medical threat. So
and what better place to find anti fungal agents than
these bacteria that have been selected to make anti fungal
agents? So that's that's one argument. Now what
you could also say is, Let's get back to
, let's say then to Jamison. Um, that
one other way to conceptualize what these molecules were evolutionarily
selected to do was to kill fast growing you carry
. It's the path agenda, the predator fungus in
the presence of slow growing you carry. It's the
food fungus or the insect. And if you then
think about okay, what else could you do with
molecules that kill fast growing eukaryotic cells in the presence
of slow growing eukaryotic cells? You'd say, Well
, maybe they would make anti cancer agents. So
this is just a simple example from, uh,
from the NIH 60 selling panel And what it what
it shows is a number of cancer cell lines.
That's what what are listed in this column, and
then it shows for this molecule relative. I mean
, this kills all of them, but it shows
what kinds of selectivity, uh, that it has
against the different cell lines. So what you're looking
for is something that has bars going in one direction
in bars, going the other direction, that is
, you don't want something that's just generally cite a
toxic. You want something that's selectively site a toxic
. And this actually has a very interesting, unusual
fingerprint in this essay. The other thing that I
think is significant about this way of looking for things
, um is the following that One of the reasons
why I think natural products haven't flourished for drug discovery
is that very often typically, they're one offs.
You get a molecule. It's very active, does
something and, you know you work on it,
and then it turns out it has some liability.
And you don't have a plan B, whereas with
normal what I'll say synthetic chemistry based drug discovery.
They always have backup compounds or things other things they
can try. And, uh, and and then
one of the difficulties with natural products is you often
don't have these backup compounds. You don't have a
way to go in and make sort of deep seated
changes to the structure. So using this as an
example, we tried the following. So, uh
, so there's a dent in German license. And
so now, once you know what kind of molecule
you're looking for, you can begin to, um
, you can begin to be much more focused in
your search as you go through these bacteria. And
so and so that was the original one. And
then, uh, and then we found this one
. And your response would be, Yeah, that's
great. You managed, uh, to use a
slightly different starter unit, right? One carbon different
starter unit. And you know, so I'm not
that impressed. But if you keep poking around,
you then begin to find some other things. And
so you say Okay, this is a change.
This what was clearly derived from A from an Allen
in here is a glycerine here that got oxidized that
got that. Alunan is a threatening. That oxidation
doesn't occur here, so you can see some more
deep seated changes. But if you really keep poking
around, you can then find things where that amino
acids, So that that and that those three atoms
of of this cycle are dropped out. And so
you end up with a cycle three atoms smaller.
And this, then is the, uh, the
Ester link. And again, you can see changes
here and changes here. And then you can find
a spectacularly interesting functional group there where this hydroxy Mamic
acid has been transformed. I mean, it looks
simple to make an O H into an N H
. Two. But there are really no, uh
, no examples of that. So, um so
that's interesting. So and there are more right one
once, you know. And then once you have
the bio synthetic pathway and you have to have the
gene sequence, you can start doing, probing or
sequencing to find more and more of these things.
Here's another one. So this is golden a mason
, which is one of the compounds you found early
on in a term mine. Gildan a mason is
has become an important, uh, I would say
drug model because it's an HSP 90 inhibitor, which
is way up regulated in cancer cells because they're making
all kinds of screwed up proteins. So you need
you need something to help you help them out.
And so this inhibits the HSP 90. That helps
helps refold troubled proteins. And this was not a
particularly head. Some liabilities these things invariably do.
And so they were made, uh, derivatives.
So the things that are in clinical trials all have
replaced this with some amine. And actually, what
we found from the termites, then is something where
That's no mean. So you can say Okay,
that's kind of interesting. Uh, do you have
anything else? And then we found just a whole
raft of things. Um, some of these are
are quite speculative, but what they all represent is
additions in the same region of Golden A missin.
It's actually a region that sticks out away from HSP
90 in that in that in that structure and and
it removes, uh, sort of a metabolic liability
that other gildan emissions have had. So we can
find all kinds of of new, of new golden
image since. And so there is a way in
which you can you can expand, um, expand
on sort of well known structural templates. And I
think you can also answer very profound, very interesting
questions which no one has actually tried to tackle.
And so what's interesting about natural product bio synthesis?
Do you have a bunch of jeans? And for
bacteria? They're all in an order. They're usually
all on a continuous stretch of DNA, often in
the order in which they carry out their reactions and
, uh, and they make and they make a
molecule. But if you want to connect with other
kinds of biologists. What you could say is Look
, I'm studying a multi gene genotype, uh,
genotype from a multi multi gene genotype. Okay?
And the beauty of that is the phenotype is absolutely
clear. You have a molecule, you don't have
a molecule. It's not like you have blue eyes
or greeny blue eyes or anything. You know the
other problem with multi genic traits, and you then
very often know the role of each of these genes
in the phenotype. You know, like, Oh
, that's the one that puts in puts in that
bomb. Right? So you have. So you
have a way to begin to begin tackling this and
and really, what we should all be doing is
studying the evolution of natural products because we talk a
lot about natural products. I mean, I talk
a lot about natural products. David talks. You've
probably heard, you know, they're so special,
they're you know, they've got complex stereo chemistry,
their confirmation early, biased. They're this. They're
that all those things are true. But actually what's
special is that they all have an evolutionary history.
They were all selected to do something. Everything else
is sort of an accident of the starting materials that
nature has in the pathways that nature has. Now
, why aren't people doing that? One of the
reasons is shown here, and that is that in
bacteria, you get horizontal gene transfer. So this
is a molecule called called Andrew Mine. Uh,
this is where it comes from from a bacteria in
upstate New York. Uh, that works over apple
trees. Disease called fire blight. This is what
you get if you look at a very different kind
of bacteria from a salt pond in, uh,
Southern California. But they are gene for gene,
Almost residue for residue. Identical. How could these
two things completely geographically separated? Uh, tip typically
separated. I mean, they come from very different
lineages. How come they have the same genes?
And what that does is makes it very hard,
because if you think you have a parent strain,
so this might be a organic chemistry Biology. You
have a parents train and then two daughters strains,
and you want to look at molecules here or molecules
there and try to figure out where they came from
. What, you end up looking at these things
? The things that look like that. They look
kind of familiar but kind of different, right?
I mean, you know, like, where the
hell did that come from? Those two things have
never have never been together. And so and so
you get these sort of time, Eric kinds of
things appearing because you have jeans winging in from all
the other bacteria in the environment because they're always exchanging
sort of sort of DNA. And what's different than
now is if you do these insects myelogenous because,
you know, let's say, for example, if
you look at the leaf cutter ants, their family
tree, their file a genie is well known,
and the file a genie of their food fungus is
well known because it follows the file a genie of
the ant and the file. A genie of the
antagonistic fungus is largely the same, although occasionally things
do creep in from the outside, and then it
follows that the, um, the mutual mystic bacteria
largely have that same that same file a genie and
more important, even than trying to see the vertical
gene transfer as opposed to horizontal gene transfer, what's
much more important is here you actually know what the
selection principle is. That's the other reason why it's
hard to talk about the evolution of natural products.
You don't know what they were selected for. We
know what we use them for. We have no
idea how they were, how they were derived in
nature. And so and so here, you know
. Okay, It was supposed to kill this,
make something that would kill this and not this.
Okay, so So that's that is agriculture. Uh
, and why? I think it's an interesting place
to look for interesting chemistry, including, I think
, uh, interesting new drug possibilities. Then the
last part of the really a three part seminar is
another farming story, something called primitive farming. So
what you just learned about is now called for the
last two years advanced farming. So there's something called
primitive farming. And it was an exceptionally interesting paper
in 2011 from, uh, the Queller and Strassman
lab they're now at When they published this, they
were at rice. They're not Washington University, and
it's the PhD thesis of Debbie broke. And so
if you just look at theirs, then this is
now completely familiar. Has been a large part of
the ecological success of humans. A handful of animals
blah blah, blah, advanced agriculture. That's and
then they go on, they say. We want
to say that the social media, addicted to stealing
described him as a primitive farming symbiosis. Which raises
a couple of questions. One. What's the social
amoeba? And to what's What's primitive farming? Okay
, so I'll spend the last few minutes trying to
tell you about that. So, uh, what's
the social media? Uh, that's a social amoeba
. So if you look at the life so this
social media has been unbelievably well studied in the laboratory
, and it's been a model organism for all kinds
of things. So it's it's life cycle is the
following? Uh, there's a spore, uh,
which, uh, deployed spore and it, uh
, it gets deposited. And it it grows into
a single celled amoeba that crawls around and that amoeba
eats bacteria that it finds. Okay, so it's
a single cell you carry out that eats, uh
, it's bacteria. And then, as it eats
more bacteria and it keeps dividing to make more more
amoebas, the food goes down, the amoeba population
comes up and they realize. Okay, this can't
go on forever, So they, uh they begin
to aggregate. And so these, uh, these
multiple amoebas come together. They form, they come
streaming together. Uh, they form this slimy mold
looking like thing. They come along, they form
a slug, and then the slug differentiates. So
about 20% of the cells end up in a sterile
stock, which lifts it off the surface, and
the other 80% end up in a fruiting body up
here at the top, where all the spores are
. Okay, So why do people study Dick t
They study it, uh, for developmental reasons.
Because you've got all of these different forms. Uh
, they can study it for actually a lot of
early cell biology. Things were discovered here. You
can think about your discovering the origins of multi cellular
charity, right? How single celled animals become multi
celled animals. You can think you're studying the beginnings
of sociology. How do things learn to cooperate?
You can think you're learning to studies chemical signaling.
So the signal to come together is cyclic GMP,
right? They follow a cycling KMP gradient. Um
, if you want to know, really a cool
story. It's this molecule called def differentiation inducing factor
because one of the things that's really important is if
you're an amoeba, single celled amoeba now part of
a slug crawling around you'd much rather evolutionarily speaking,
be at the top So your DNA goes on if
you're in the fruiting body, that in the sterile
stalk and so that's governed by this molecules. So
the differentiation is between things that end up there and
things that end up there. So so So people
have studied dick tion many labs for many years and
learned learned a lot of things. What Deb Bach
did that was different was actually looked at Dick T
in the wild, and she looked at victory in
the wild. And she looked. She looked inside
these fruiting bodies, and this slide is really miserable
. But anyway, there are. She could see
ones that were called farmers and some that were she
called non farmers and the farmers. I don't know
if you can see these. Those are those are
the spores and those things that look like sort of
dirty blotches. Those are bacteria, so they had
bacteria in with the spores in the fruiting body in
the non farmers. They just had the fruiting body
that just just came there. And so then you
would ask, What's better? Farmer non farmer Cain
or Abel? What? What, what do you
like? And it turns out it's completely context dependent
. So if you take one of these spores from
a non farmer and put it on a Petri dish
with no bacteria, okay to simulate landing in a
in an area where there are no bacteria, natural
bacteria and you ask, How does it do?
How many, how many survive and keep going?
The answer is essentially none. Okay, if,
on the other hand, you're a farmer and you
end up landing on a dish with no bacteria,
who cares? You brought your own bacteria, they
grow, you eat them and life is good,
so you kill him, right? Literally Now,
On the other hand, if you land on something
that's already been seated with bacteria, the non farmers
do better than the farmers. Why is that?
Because the non farmers sorry because the farmers can't eat
everything right. They have to practice what the article
called prudent grazing because they have to leave some bacteria
to get scooped up and incorporated in the sports.
And so they, if you will have fewer bacteria
to turn into amoeba cells. And and so,
um and so they do worse, slightly worse in
the competition. And so in the wild, it's
about 20% non farmers and about 30% farmers from what
she saw. So that actually then adds a very
interesting a bunch of considerations to how to think of
farming. So this part isn't isn't so confusing we've
got We've got the farmers, which are the farming
dick tease, and we've got their crop, which
are the bacteria that they eat. But now,
complicating the picture, we have these non farmers,
which are essentially identical to the farmer. Okay,
because no one ever knew that there were. There
were two different things until Debbie started looking, and
they have sort of an odd, odd relationship now
the other. The other thing that's interesting is that
the bacteria in the fruiting body sort of like a
market basket of bacteria. It wasn't just one and
about. As you can see from that thing,
about half of them could be eaten, and about
half of them weren't eaten. In fact, about
half of them couldn't beat. They would rather die
than eat those and so and so that raised the
question that the article, uh, posed generally.
So this conservative food for the non farmers generally for
farmers and non farmers like the function of the other
half, that's the other half of the bacteria,
if any, is unknown. But by now,
you can probably make a pretty good guess what the
function of the other half of the bacteria is.
It's got to be that they make things that benefit
the symbiosis, maybe by killing pathogens, maybe by
doing something else. So we made have taken,
I think, an initial step in trying to de
con veloute sort of what's going what's going on here
. So he picked one farming strain actually collected in
Virginia and one non farming strain collected in the same
teaspoon full of soil from the same place in Virginia
. Okay, so this is Queller. Strain 1
61 and strain 1 60 and these two must have
been exposed to each other because they're from the same
tiny niche. And we decided we being Pierre Stele
, fourth, who did all of this decided to
focus on some pseudomonas fluorescence because Pseudomonas was reasonably easy
to work with, had some known chemistry, had
a bunch of things that made it attractive. And
so we worked with with Pseudomonas for essence. And
there was the strain that they ate, which was
called be imaginatively. And there was the strain that
they didn't eat, which was called a. So
It's really a story of four things, but it's
really a story of two things and then two very
closely related relatives. So it's the dick T farmer
and non farmer and the crop and the and the
something pseudomonas fluorescence. So the the the argument I
don't have to spell out was that this would probably
make something that would protect, uh, that would
protect the dick t from something. And so one
of the essays we did is it turns out that
nematodes like to eat Dick t. So like C
. Elegans, the standard lab animal likes to eat
a dick t. And so we did a that's
we did a C. Elegans avoidance essay. So
what you can't see is there's a Petri dish here
and what what, you make the problem is the
lights are wash out a lot of things. And
so, uh, these are you make a streak
of E. Coli, which these are the the
E. Coli that C. Elegans normally eats in
the lab. Their favorite food. This is the
, uh, streak of of the pseudomonas fluorescence that
we're testing. And then, um, and then
you put down a bunch that's fine. Put down
a bunch of young adult nematodes and then you say
, What do they do? Do they crawl over
here to get to what they really like to eat
? Or do they stay back here? And the
answer you can see is if it's a the strain
that we think makes something they won't go near it
. That is, the nematodes all stay there.
On the other hand, if it's, um,
if it's be the strain that can be eaten,
the nematodes quickly zip over here and spend spend their
time over here eating their preferred E. Coli food
. Okay, so that says that's one example of
the way in which the non eaten bacteria can protect
can protect the dictator. So we did, Um
oh, and then it turns out these are really
bad actors, so if you drop, they won't
cross it. I've told you that, but if
you just drop them on it, they die.
Okay, so So, yeah, so they really
don't like like that. So if you then do
, um Well, yeah, you are the following
experiments if you take the crop and these are the
same scale. So those are what those pseudomonas for
essence look like. And these are what these look
like. And you do what Pierre shamelessly described as
untargeted differential Metabolism X. Okay, So what does
What does that mean? Metabolism IX means he ran
an H p L c Differential means he ran to
h p l C and untargeted means he had no
idea what he was looking for. So if you
do that, what you find is that the crop
makes, uh, makes it very well known.
Uh, pseudomonas molecule, uh, called parochial,
in which everywhere you look up in the literature is
described as a scenario, for although I think it's
very unlikely that that's actually it's it's purpose. If
you look at the one we call the ally,
it makes some very well known compounds that makes this
diacetyl flora Delusional, which, while it looks rather
innocent, is a well known sort of antibacterial antifungal
compound. It makes parole nitrogen, which looks almost
like something out of Monsanto's lab. Um, but
that also is very well known anti fungal antibacterial.
It makes HTN, probably don't need to tell this
audience. That's, um, that's a bad actor
. And then it made this molecule, which,
which we called the Crow mean because, well,
for what it's a crow mean until we never could
agree on a name for it. And so it's
now just come to the world is crummy. And
so I think I don't need to say much about
this, this or this. Those are all sort
of well known molecules that you can imagine. Uh
, it's well known not only molecules. It's well
known that Pseudomonas makes them, and it's well known
what their biological properties are. So those are classic
, if you will defensive molecules. This was more
of a puzzle because we didn't know exactly what that
did. It was actually, it's actually kind of
a cute molecule, for if you're a connoisseur,
it doesn't be bio synthesis of that is is an
obvious so so uh, Pierre and and Debbie began
working on on what that was, and it turns
out it's very, very interesting. So this is
, um, along here is the percent change in
spore production. So if you look at the fruiting
bodies and you you open them up. So this
is what Debbie does Incredibly tedious work and count the
spores inside the size doesn't tell you anything. Size
doesn't matter. Here is in many things, and
And what And, uh, but what matters is
how many spores are in there, Okay. And
so and so this is, if you will,
the average number of spores that you could find in
a fruiting body. And then if you start treating
them, going along here with, uh with with
the chrome mean and this is like nanograms per mil
. So this is, uh, this is about
one or two Nana Moller starting it here. What
you can see is two things. One is that
it dramatically increases the reproductive success of the farmers and
at a slightly higher. But the actually environmental concentrations
are up here at a slightly higher concentration. It
suppresses reproductive success of the non farmers, right?
So this is actually a completely different strategy, right
? This is a This is a new feature.
And so what it looks like then, is this
that we this part we understand this part. We
understand these compounds which are made by this all can
suppress parasites or pathogens. But this this has now
changed in the old day. And the other symbiotic
relationships these two were mutual ist on the theory that
the enemy of my enemy is my friend, right
? But here, their mutual list for that reason
but they're also mutual is because this increases the reproductive
success of that and it also suppresses the reproductive success
of that. Okay, that's a very interesting thing
to add. And in general, with this,
what's what's the? The interesting question is, how
are these two distinguished and how are these two distinguished
? So this is what Queller and Strassman are working
on. So I'll tell you about how those two
are distinguished. So you say you're both Pseudomonas for
essence. How similar are they? So you do
16 s and they have identical 16 s sequences.
So you sequence the whole genome and they're essentially identical
. What does that mean? It means they're about
two or 300 differences, and about 50 of those
are non synonymous. Differences that is would yield a
different apparently yielded different amino acid. Um, a
coding in a coding region. So Pierre looked at
those and then was very excited and he said,
I understand what's going on. It's in Jackass.
I had no idea what he was talking about,
but I said, That's great. And so oh
, it's in Jack. I really didn't know what
he was. So yeah, so there there's a
a gene called Jack makes a protein called Jacquet,
and they were completely identical in the two. Uh
, the two strains, with the exception of one
base pair, changes See to a t, uh
, down here and what it did is it changed
, um, glutamine to a stop code on.
And what it meant then, was that the one
made by the food was missing all of this.
Okay, And this turns out to be a helix
turn helix motif to turn on to bind to DNA
to turn on genes. So what? Gak Gak
s is a typical two components signaling pathway. So
jackass gets the signal from the environment his sardine gets
phosphor related gets passed along, and eventually that high
energy phosphate is passed on to an ESP Arctic acid
here in, uh, in the gak a gene
. So this is this is the sensor, Uh
, this is the one that carries the message.
And when that residue gets fuss for related, this
helix turn helix motif takes on a different different shape
so it can bind regulatory genes and turn on all
kinds of things and bring up a response. And
the response actually turns out to be the production of
secondary metabolites. So it turns on parole, nitrogen
production, chromium production, HCM production, and it
turns off parochial in production. Okay. And so
now you can see what's going on that if you
have a stop signal, you don't have that.
If you don't have that, you don't have that
. You don't get any response. So that one
base change it turned, turned the two into each
other. So to check that that was true.
We made a knockout. So this shows you what
the food one is. It makes the parochial.
And as I told you, this is what we
decided to call an ally and it makes the crow
mean and it makes parole nitrogen and makes h c
n. And so what we did is take that
and knocked out the gak a gene. And if
you do that, it becomes this right that becomes
this which is identical to this. And so it
turns, turns the whole thing around. So we
can feel in one thing that the difference between those
two, which is so different, metabolically so different
functionally, is actually one base pair. That's all
it takes. Turn one into the other. Okay
, um, so ah, I'll just close that
these bacterial Symbian are a great way to learn a
number of things chemistry. But also, I think
you can learn some very interesting biology by doing it
at this level at the molecular level. And the
other thing is that advanced primitive agriculture is actually pretty
advanced. It's just different and and in an interesting
way and leads to some more things to look for
. So let me thank the people involved. So
, Cameron, for the first part on what I'll
call advanced agriculture and some people in his lab,
Dave and Joan and Deb, you know, Debra
and at Washoe. It was all started really by
Dong Chan, who's now gone back to Seoul National
University. And then Gavin and Josh and Boris carried
it on. Now it's being done by Kion,
Tim and Claressa. Uh, and the other,
uh, was, uh this was done almost entirely
by Pierre and Alexandra in, uh, in my
lab. And Tina worked actually on both on both
projects. So anyway, I think these people for
for support financial support these people for all their hard
work and you once again for the invitation. So
thanks. Yeah, John, thank you very much
for a wonderfully interesting and diverse talk. We are
running a little bit late, so let's take just
two questions. One from a student, one from
Okay, I see a factory with a hand up
there. Okay. So take into the story of
compromising is a very complicated structure. Registrants come from
one hydrogen bomb. Loss of hydrogen bomb becomes,
for instance, so in this scenario, how this
revolution of metal, I'd pray and has Anybody can
learn from these models about resistance, so you can
certainly learn about these models from resistance. We haven't
done it much because the fungi grow slowly and are
genetically complicated. But and so we've put that off
until we find something we think might be more significant
. And then we'll look into what the what the
resistance is is a way of finding out what its
mechanism of action is. Because that would be a
driver. Or this morning you should start seeing Oh
, no question. No question. Mm. All
right. We have a student question. Yes.
Uh, you said sometimes he's kind of delicate,
You know, evolutionary interactions between one predator species and
then bacteria is can produce and the foundation to stop
. You mentioned that sometimes things get in the outside
. Do you see any kind of red queen effect
where these bacteria pathogens are fighting each other evolutionarily or
another? Oh, so there's no question. I
mean, so one of the things you could ask
is why, in a relatively short time, how
did you get so many different species? Right?
And that clearly is sort of a red queen effect
, where because new, because predators evolve. And
then you have to evolve new strategies right, which
you can do on an individual basis, and then
those can continue to escape those predators. But then
the predators will evolve new strategies. And so it
just keeps going. Yeah, so? So I
think that the evidence that that that's going on is
the spectacular, if you will. Evolutionary success,
uh, leading to the large number of examples.
Okay, So, regrettably, we do need to
enter. There's just a couple of announcements before I
close. First of all, just to thank the
TLS s for sponsoring this lecture is also co sponsored 1039 --> A:middle L:90% .