cc >> Tom Zinnen: Welcome,
everyone, to Wednesday Nite at the Lab.
I'm Tom Zinnen. I work here at the UW Madison
Biotechnology Center. I also work for UW Extension
Cooperative Extension, and on behalf of those organizations
and our other sponsors, Wisconsin Public Television, the
Wisconsin Alumni Association, and the Division of Continuing
Studies, thanks for coming to Wednesday Nite at the Lab.
We do this every Wednesday night, 50 times a year.
Tonight, it's my pleasure to introduce to you Nancy Keller.
She's a professor in the Department of Medical
Microbiology and Immunology and in the Department of
Bacteriology. She received her doctoral
training in the Department of Plant Pathology at Cornell
University. Her postdoctoral training with
the US Department of Agriculture in New Orleans first introduced
her to the genus Aspergillus where she worked on the
identification of enzymes required to produce the
mycotoxin aflatoxin. Since that period of time,
Dr. Keller has not only retained her interest in understanding
how Aspergillus fungi produce toxin metabolites and using this
knowledge to design controlled strategies to help combat
diseases caused by pathogenic Aspergilli, I would just say
Aspergilluses but that's all right.
[LAUGHTER] She has also expanded her
research to work in the field of drug discovery utilizing
knowledge gained from the natural product formation
by fungi. This is one of the funnest
titles we've ever had: "Aspergillus Can Kill Us
and Thrill Us." Please join me in welcoming
Professor Keller to Wednesday Nite at the Lab.
[APPLAUSE]
Nancy Keller: Well, hopefully we'll have a little fun tonight.
And can everyone hear me okay? >> Yes.
Okay. Wave if you can't or otherwise.
Okay. So, what I thought I would do
is introduce you, first, to Aspergillus.
I think many of you know Aspergillus but perhaps some
don't. For those of you who were
sitting here earlier and you saw the beautiful waving of these
objects on the YouTube, that was a video, an artist's rendering,
of Aspergillus. So, this Aspergillus is a genus
of fungus that was first described in 1729, I'll give you
a little history here, by a priest scientist.
I guess we call, what do you call a priest?
Reverend? Micheli or Priest Micheli.
And I was trying really hard to see if I could find some of the
pictures that he drew of Aspergillus from his very early
microscope. I couldn't find his but I did
find this drawing from Louis Pasteur.
Actually, maybe I can do what is asked and use this little
hoo-dicky here. So this is Louis Pasteur's
drawing of Aspergillus glaucus. And I part that really somewhat
resembles the Aspergillus is this particular figure right
here. And let me just show you what he
was trying to draw at the time. Aspergillus is a genus of fungus
that produces its sexual spores, and, yes, fungi have both sexual
and asexual spores. Pardon me, these are the asexual
spores, on top of a stalk that has a vesicle on the top and
cells that produce these spores that fly into the air.
And I'm quite sure we're probably breathing in some of
these spores right now. If you're a healthy person, it
doesn't make any difference because your immune system will
kill the spores. Now, where did this name come
from? So here's the really interesting
part with this fungus. So Reverend Micheli was a
scientist. He was also a Catholic priest.
Monsignor. >> Monsignor.
And what he did notice right away is that the Aspergillus
stalk actually looked quite a bit like an instrument that's
used by the priests in the Catholic church.
This instrument is known as an aspergillum, and hence he called
the fungus Aspergillus. And this gives you an idea of
how similar the likeness is between this fungus and the
Aspergillus. You can see here the priest
sprinkling the holy water, and it does look very much like the
spore is coming off of the Aspergillus.
And this, for those of you who are interested and mesmerized by
that YouTube I showed earlier, this is the address.
So, now that I've introduced you to Aspergillus, I'll take a
moment to tell you how I got interested in working with this.
Tom did give a bit of my background, but why, you might
ask, did I start working with fungi and Aspergillus.
And I will tell you that I actually did have an earlier
episode that brought me to fungi.
I was a Peace Corps volunteer before I went to graduate
school, and I was a volunteer in Africa in a country called
Lesotho. And where I worked, what I
noticed that oftentimes the food was really in very poor
condition. We usually had enough food, but
it was often tainted with mold. And a lot of my students got
sick over the course of the many years I taught there.
So I became very interested in agriculture and quality of food,
and when I was in the Peace Corps, one of the advisers in
Lesotho was actually an alumni of Cornell University where he'd
taken international agriculture courses, and he's the one that
got me interested in Cornell and plant pathology.
And ever since I got my degree, I have always worked with fungi.
I really enjoy fungi very much. They're fascinating organisms.
And I've spent most of my career with fungi with Aspergillus.
So this title we had that Aspergilli can kill you or
thrill you, well, it is true. And I want to introduce you to
some of the big players in Aspergillus.
There's probably about 180 species to 200 hundred species,
and some of them are very well-known.
There's a fungus called Aspergillus fumigatus which is
the most common cause of the medical diseases posed by
Aspergillus. And then the Aspergillus that I
first started working with was Aspergillus flavus.
Aspergillus flavus and its cousin, Aspergillus parasiticus,
are the very common food contaminating fungi which
produce the mycotoxin called aflatoxin in our food crops.
And we'll talk a little bit more about aflatoxin later, but it is
the most potent natural carcinogen known to humankind.
I'll also point out that Aspergillus flavus is the second
most common Aspergillus to actually invade the body and
cause various diseases. A disease of the mammal caused
by Aspergillus is known as an Aspergillosis.
Now, we do have Aspergilli that we like that are on the positive
side, and why are they useful? Well, Aspergilli, and other
fungi, are one of our most common organisms we need for
composting and destroying, recycling, carbon and nitrogen.
There's a particular Aspergillus called Aspergillus nidulans,
which is one of the best established model organisms for
genetics. We actually work with this one
quite a bit. The history of this particular
fungus is very interesting too and would take us into World War
II, but maybe after if you're interested.
Then we have the food production Aspergilli.
And Aspergillus niger is a fungus where we get a lot of
enzymes that we use both for food, citric acid.
Also this is the fungus where we get a lot of the enzymes that we
use to make what's, I think it's called stone-washed jeans.
Can I interrupt you? >> Yes.
What do you mean by fungus? >> By fungus?
A fungus is an organism that has its own kingdom, and, let's see,
I'll give you an example of one you probably know well:
mushrooms. Mushrooms are fungi.
A fungus is one; fungi are many. And they're just a kingdom of
life. >> Well, why is the mushroom a
fungus? >> Why is it a fungus?
Well, the fungi are put into this kingdom based on their DNA
sequences and the enzymes and proteins they make, and this one
is classified, mushrooms belong to this kingdom based on their
evolution and the sequences of their DNA and their proteins.
And they reproduce by spores. >> Most of them do.
Okay? I'm sorry, I was there making an
assumption that everyone knew what a fungus was.
Yes, they reproduce by spores, most of them.
I think you probably have seen Aspergillus, or a related fungus
called Penicillium, in your refrigerator on food that you
don't eat readily. Or on your shower curtains.
I'm quite sure you have all seen these fungi.
Now, the Aspergilli we like, for those of who like Asian food
especially, are Aspergillus oryzae and Aspergillus sojae.
I'll show you some pictures of these.
These are the fungi that are used in making, oh, soy sauce
and sake and various fermented foods.
And then we have a series of Aspergilli where we actually get
a lot of the pharmaceuticals we use to combat drugs, I mean,
combat other organisms. For example, these echinocandins
right here. These are actually compounds
made by Aspergillus nidulans and Aspergillus sydowii, and they
kill other fungi. So fungi are always competing
with each other. and they sometimes like to kill
the other fungus so they can get their own turf.
Lovastatin, maybe some of you are familiar with statins or
even use these statins. These are compounds that help
people with high cholesterol, and there are many others too.
So now I'm going to give you an example of some of these
particular good and bad Aspergilli.
Aspergillosis, as I told you earlier, is just the generic
name for diseases caused by Aspergillus.
And this is becoming a very serious problem in the world
because we're now able to keep people alive where normally we
wouldn't, and we keep them alive often by using, let's say,
steroids which lower the immune system.
So what we're finding now is a lot of people that have diseases
which lower their immune system are getting Aspergillus
diseases. For example, people that have
AIDS often get Aspergillus diseases, diabetics, people with
leukemia. You're particularly susceptible
if you've had some organ transplant.
The range of diseases are from fairly mild, a lot of people
have allergies, that can be mild or serious.
Often you can get a ball in your lung.
That sounds horrible but it's really not the worst thing in
the world. We're pretty good at removing
those aspergillomas, that's what the fungal balls are called.
It's when you start getting invasive growth that the fungus
is growing through your body that they're very hard to
control. Now, I told you there's about
180 species of Aspergillus, but there's not that many that cause
disease. One criterion is that you have
to grow at 37 degrees because that's the human body.
So these are some of the ones that are more frequently found
in humans. The genetic model Aspergillus
nidulans. Amphotericin B is an antifungal.
We're having a big problem now with a lot of these Aspergilli
being resistant to antifungal drugs.
Aspergillus niger is a dark Aspergillus.
Aspergillus terreus is particularly difficult to treat.
It is more resistant than the other Aspergilli to antifungals.
Aspergillus flavus, and the most common species, as I told you,
Aspergillus fumigatus. Here's an SEM, a scanning
electron micrograph, of the conidiophore, the asexual
spores, here's one right here, of this fungus.
You're going to see a couple gross pictures right now.
These are just some examples of what can happen if you get these
infections. I will tell you by the time you
see a lesion on your skin, that's not a very good sign.
I don't know what tissue this is, but you can see a
sporulating colony right there. This is fingernails that have
been infected with an Aspergillus.
The very common disease is this aspergilloma.
You can see here is lungs and here is a ball.
You really can get rid of these balls pretty well.
And this is a case where they didn't get rid of the ball very
well though. And this is a lung from someone
who died, and here is one of the, this is a huge
aspergilloma. And if you do sections and you
stain the lung or any other part of the body which has
Aspergillus in it, you'll see the fungus, and here you will
recognize that Aspergillus or the aspergillum.
So it's a dead give away that's the fungus you have.
Now, the other player which can kill you is the Aspergillus
flavus that produces toxins in your food, and when you eat the
food, you get sick. The toxins that fungi produce in
food and feed are called myco. Myco is from the Greek name for
fungus. So mycotoxins.
And really the history of mycotoxins is quite fascinating.
It is felt by historians, although we cannot prove this,
that the 10th plague of Egypt is most likely caused by
mycotoxins. If you read the documentation,
it really sounds exactly like a mycotoxin outbreak.
And I've given some of the history over the ages when we go
back where we can figure out that certain diseases were
associated with mycotoxins. I'm not going to go through all
of this now but bring you to what I call modern
mycotoxicology. And this is what actually made
us understand that fungi can produce toxins that are very
harmful. This happened in 1961 when there
was a shipment of peanuts from Brazil to England.
The peanuts were used as feed for turkey and chickens, and it
resulted in over hundreds of thousands of poultry dying from
England. This is a massive outbreak, of
course, and this really set the stage for the scientists to try
to figure out what happened. They called it turkey X disease.
And ultimately what they found, it was a compound produced by
the fungus Aspergillus flavus, that's where we got the name
aflatoxin, that caused this disease.
And what happens here are four turkey poults.
This one was fed aflatoxin. You can see it's stunted, and
the liver has a fatty discolored tissue look.
This turkey is not going to make it.
Since then, aflatoxin periodically comes into the
news. One of the more interesting news
items, I guess, about, geez, when was the Desert Storm war?
That was about... >> '91.
'91. Wow, okay.
So it was weaponized and put in scud missiles.
Probably it was all destroyed before it could do anything as
the bomb exploded, but nevertheless.
And all of us who were working with aflatoxin got phone calls
from the federal government to ask if we had sent any strains
of Aspergillus to anyone we shouldn't have.
That was a funny question to ask because Aspergillus flavus is
all over the world, but I did get a phone call.
This is what it's going to look like most commonly.
You'll recognize this corn ear, and this fuzzy green stuff is
Aspergillus flavus, and there's the molecule of aflatoxin.
I know only certain people enjoy to look at chemical structures.
I'm quite found of them so I won't give you too many.
You see that, you don't want to ingest it.
Aflatoxin, we're still working on it.
I do a lot of work with this. We still have not been able to
control it. The fungus is ubiquitous and
when the environmental conditions are right, it's going
to grow in our corn and our peanuts and our tree nuts.
What we do do in this country, though, is we analyze these
particular crops to look at the level of aflatoxin.
And the food in our country will not have high levels of
aflatoxin. However, as you can see here, we
do not look at our pet foods so carefully.
Since I've been working with Aspergillus there's been three
dog poisonings. I'll just go through this.
The first one was in Texas where I actually was working at that
time at Texas A&M. Much more severe is the constant
human poisoning in Kenya. Now, I have to back up a little
bit. I had told you aflatoxin was a
carcinogen and it is. And you can die from liver
cancer from aflatoxin, but when you have it in high amounts, it
is toxic and can kill people and animals in a week to a few
weeks. When you see a poisoning case,
you know there has been massive amounts.
And then in 2006, '05 and '06, there was another big dog
poisoning case in the United States.
There's been a whole lot of breakouts just recently.
There's frequently aflatoxin contamination in items from
China, in milk and cooking oil recently, and just again in
December there was aflatoxin found in dog food from three
different companies. And this is really becoming a
very serious problem with the dog food.
So there's a little interesting article if you guys might want
to read it. Let's switch, though.
I've given you a lot of bad news about the Aspergilli.
There's a lot of good news too. And so here is, and this is
quite an interesting juxtaposition, Aspergillus
oryzae. Why do I say it's interesting?
Well, we have now actually sequenced the genome of many
Aspergilli. I think all of you probably are
familiar that the human genome has been sequenced which has
given us a lot of very interesting information.
Well, so have many fungi. And what we found out is when
Aspergillus oryzae genome was sequenced, I am going backwards,
they said, huh, it's Aspergillus flavus.
What's going on here? Well, it turns out that
Aspergillus oryzae, which we make our food with, actually
does seem to be a tamed Aspergillus flavus.
There's a lot of speculation as to what happened over the years,
how we got this safe Aspergillus flavus.
That is something we could talk about after the lecture if
you're interested. But in any case, Aspergillus
oryzae is the fungus that is used to prepare a lot of very
yummy condiments, foods and liquids.
So it's really interesting. There's a YouTube, and I'm not
going to do this because this would take too much time, but a
very interesting YouTube which shows how you make soy sauce and
how they put the fungus into the soy beans.
So soy sauce comes from soy beans that's been fermented with
Aspergillus. This is a little recipe here.
You have soy bean cake, they do have wheat and down.
Here we add Aspergillus oryzae. You put it in these different
vats, and you get a mash called koji.
Is that in China? >> That was in Japan.
You can buy koji. And now what I have to show you,
I have a little surprise here, koji and soy sauce and
Aspergillus are so important in Japan that the national fungus
is Aspergillus oryzae, and they've made a toy Aspergillus
oryzae. My one graduate student is from
Japan, and when she went back home to visit...
[LAUGHTER] She brought me one of these.
[LAUGHTER] We actually hang this
in our lab. So this is Aspergillus oryzae,
the good Aspergillus. What else do we get from
Aspergillus? Thanks to penicillin, he will
come home. Maybe some of you have seen
this. This is from World War II.
So what's this all about? Well, I think some of you might
know about this finding, this accidental finding, by Sir
Alexander Fleming in 1929. He noticed he had a petri dish
of bacteria, and what he saw is that a fungus had landed into
this petri dish and it was killing the bacteria.
There was an inhibition zone that would not let the bacteria
grow. What he and others subsequently
found is that the fungus produced a compound called
penicillin. Now he actually identified this
from a fungus called Penicillium, but good old
Aspergillus nidulans also makes penicillin, and much of the
genetics in biochemistry was worked out with this model
system. It turns out that we get a lot
of useful pharmaceuticals from Aspergilli.
Now, we had mentioned lovastatin.
I'm just guessing some of you might actually have this
prescription. And here is the chemical
structure. Aspergillus terreus is the
fungus of choice to make lovastatin.
Now, I've given you a sort of brief introduction to the
different Aspergilli, and this is a nice little segue right now
where I'm just going to tell you a little bit about what we do in
our laboratory. As I said earlier, I know
chemical structures aren't everybody's cup of tea, but you
have to admire the diversity here, I'm sure.
And when you look at these, what I want you to think is that, oh,
wow, fungi make these. Not all these are from
Aspergillus. Actually, I couldn't even
possibly put in all the compounds from Aspergillus, but
this is just to show you that the compounds are very diverse.
And these compounds are very bioactive.
And that's the main reason why Aspergilli can sometimes be a
problem for us or a benefit. They make compounds that are
destructive to the animal body, though in small amounts it might
be good. Here's, I'm sure I have it,
yeah, here's penicillin here. Here's aflatoxin.
Here's the statin, lovastatin. Some of these are good.
This is another antibiotic. This is bad.
That's bad. Bad. Bad.
Really bad. Bad.
Good. Neutral.
We have one somewhat neutral one.
So what are we doing? I forgot to, I wanted to put in
this slide, too, to tell you that the production of these
compounds by the fungus of course has nothing to do with
humankind, but they're a benefit to the fungus.
And it depends in the lifestyle and the life stage of the fungus
when it's going to make all of these different compounds.
So right here is our conidiophore again from the
aspergillum. All Aspergilli will produce one
of these. And each one of these little
spores will fly in the air, and when they land on a substrate
that they can obtain nutrition, they're going to germinate.
Right there it's germinating. Now that substrate could be, oh,
a seed. Or it could be decayed
vegetative matter. Or it could be your lungs.
After it has germinated, it will form mycelium.
Under the right conditions that mycelium could form the asexual
stage again, and you have this circle.
Under some conditions, mycelium can produce the sexual stage,
and it makes these organelles, which are called sclerotia or
cleistothecia, where we get a sexual spore.
Now, I've colored some of these different parts of the fungus
different colors because they are different colors and a lot
of this has to do with some of the chemical structure.
And in these different tissues you get different compounds.
So, the fungus is very fluid and it will produce different
structures and different secondary metabolites depending
on the environment. One interesting thing is
Aspergillus produces its asexual spores in the light and will
only have sex when it's dark. [LAUGHTER]
There are other things that the fungus responds to,
and, in particular, I'm going to focus right here, is a lot
of these chemicals that the fungus is producing are in
response to stresses, particularly when the fungus
meets other microbes. And that would be, where's that
little arrow? Here it is.
And by other microbes it could be another fungus or it could be
a bacterium. So let me show you how we have
taken advantage of this information to understand how
the fungus makes these chemicals and hopefully ultimately find
some therapies to stop the fungi that we don't want from invading
us but also maybe find new drugs.
We made a fantastic discovery, really.
We were very fortunate to find a gene called laeA.
This is now in the fungal world very famous.
I don't think anybody here has heard of it, but anyone who
works with fungus now knows about it because it turns out to
be a global regulator of all those compounds I showed you
earlier. This square right here is what
we call a thin layer chromatography plate.
And if you put extracts of any organism on the plate and put it
in a solvent, the compounds will move up the plate.
And what you can see here, this is the wild type fungus, is that
as you put an extract of the fungus on the plate, you can see
many different compounds separating out.
And some of these compounds are the structures I showed you
earlier. What we found is this particular
protein, laeA, when you remove it, this delta means it's
removed, is you lose production of many of these chemical
structures. We'll just ignore this right
here. I think the main point I want to
show you is that this particular protein is required for the
expression of the genes that make these chemical, these great
variety of chemicals in Aspergillus and now, it turns
out, other fungi. Now, this little plate right
here gives you an example of just how important that laeA is.
I want you to look a little closely at this plate.
This is a petri dish, and what we have here is, can you see
these little white objects? >> Yeah.
Okay. These are larvae of a fungivore.
A fungivore is an insect that eats fungi, and what we did is
we gave it a choice. We put all of the larvae in the
middle of the plate, and in this section we had the wild type
Aspergillus, you can actually see wild type there, and in this
section we had the Aspergillus where we had removed that laeA
protein so it no longer produced many chemicals, many toxic.
And when we found is that all of the larvae, except for, let's
see, we got a little goofy one that's sort of out here by
itself, but all of the larvae made the choice to go to the
fungal strain that produced less toxins.
So we know a lot of these toxins are, indeed, a protection for
the fungus. What we then asked is if this
protein is important for making toxins, maybe it's important in
Aspergillus fumigatus, the fungus that causes disease in
humans. So what we did is we removed
that protein from the human pathogen Aspergillus fumigatus.
We found something very interesting.
It does appear to be a virulence factor.
This particular slide here is we used the Muri model, that is the
mouse model, to test for pathogenicity.
And what you can see is when you remove laeA, there is a greater
percent survival of the mice. We have a clue as to how this
happens. Here is the spore of wild type
Aspergillus fumigatus, and here is the spore of the mutant where
we removed laeA. You can see it looks a little
bit different. The more important slide are
these red ones. What we did is we labeled the
fungal spore with a dye that makes it appear green.
And right here is a macrophage. So in our lungs, we have
macrophage that one of their big jobs is to swallow up fungal
spores and kill the spores before they germinate and go
into the rest of our body. And as you can see, the
macrophage are so-so at gobbling up wild type Aspergillus, but
when we remove the laeA it just sucked them all in, and it kills
them much more readily. Part of this is due to the loss
of toxins in this fungus. So, we next asked, okay, laeA
regulates a lot of the toxins in Aspergillus fumigatus, but which
one of these are important for pathogenicity?
So what you're looking at now is a schematic of the eight
chromosomes of Aspergillus. So every fungus has a certain
number of chromosomes depending on the genus, and Aspergilli all
have eight chromosomes. What we could tell from
sequencing it is that there would be areas of the
chromosomes where we would recognize genes that we knew
could produce toxins. And these red arrows show you
all of the toxin gene clusters that are regulated by laeA.
So it really regulates a lot. One of the gene clusters we
wanted to find and investigate was a cluster of genes regulated
by laeA that make this molecule. This molecule is called
gliotoxin. Now, gliotoxin has been known
for about 80 years now. And it was isolated from
Aspergillus fumigatus and other fungi in the 1930s, 1920s and
1930s. And it was found to be a very
toxic compound. Whenever you see a sulfur bond
you might want to be a little leery about a compound.
And what they found out is that this compound actually killed
other fungi. Now we get into our interesting
story here about how fungi will make compounds to kill other
fungi. And so at first people were
thinking about maybe we could use this gliotoxin as an
antifungal to control fungal diseases in agricultural
situations. However, it also kills humans
very effectively, so that idea was trashed.
What we were able to do by understanding the proteins
likely to make something like this is find this cluster of
genes regulated by laeA and Aspergillus fumigatus.
And what we did is we deleted these genes, particularly the
first gene gliZ. The gene gliZ is a type of
protein called a transcription factor, and we knew it was
highly likely that if you removed gliZ none of the other
genes would work and you'd get no gliotoxin.
And indeed that was the case. This is another thin layer
chromatography plate, and we've grown the fungus and run the
extracts to look for gliotoxin. This is standard.
And here you see wild type, and here's gliotoxin plus a bunch of
other things you don't really want to deal with.
If you remove laeA, again you can see most of the compounds
have gone. This time in this strain we
removed gliZ only, and what you can see is gliotoxin is now
absent from this thin layer chromatography.
There's a few other compounds missing too, but most of them
are still there. So gliZ is not global like laeA.
It is specific for gliotoxin and gliotoxin derivatives.
And this is just a complement. We took the gliZ mutant and
added gliZ back to the fungus and gliotoxin is back.
And what we found, this was not surprising because other people
had shown this, it had been known that gliotoxin causes
self death. Apoptotic death is when
basically a cell commits suicide.
And when mammalian cells are exposed to gliotoxin, they
self-destruct. And what we were able to see
here, this is just a graph where we took extracts from our wild
type fungus and added them to neutrophils.
Neutrophils are the other cells in our immune system that
protect us from fungi and bacteria.
And this is the percent death of the neutrophils over, I'm not
even remembering how long we did this right now, but you can see
about 50% of the neutrophils died when we added an extract
from the wild type, but if you added extracts from the gliZ
mutant, hardly any died at all. So indeed, gliotoxin, which we
once thought of to use as an antifungal, is really a very
serious contributor to invasive Aspergillosis.
Now, I'm going to show you, though, how tricky the fungus
is. Now we're back again to this
particular gene cluster. All of these genes make proteins
that are required to make gliotoxin.
Now, I told you that people were thinking at one time, why not
use gliotoxin as an antifungal? Because it kills other fungi.
It does it very well. Why doesn't Aspergillus
fumigatus kill itself with gliotoxin?
Well, that's what is really very interesting, and you really have
to give fungi a lot of credit here, is when a fungus makes a
toxin that kills other fungi, it almost always has inside the
cluster of genes making its toxin one protein that protects
itself from its own toxin. And in the case of Aspergillus
fumigatus, gliT encodes a protein that destroys gliotoxin.
And what another lab did last year is they removed gliT from
Aspergillus fumigatus, and then they compared the effect of
gliotoxin when added in the middle of a petri dish in this
mutant to wild type. And as you can see, pure
gliotoxin does inhibit Aspergillus fumigatus a little
bit but if you don't have gliT, it's really killing it.
So this is a very common situation in fungi.
All those toxic molecules that I've shown you, the fungus that
produces it will have a protective device so the
antifungal doesn't hurt it, just the other fungi.
I'm going to end with one last story.
And this is a new story in our lab that hasn't been published
yet, but I think it will be shortly.
And I'm continuing with Aspergillus fumigatus, the
pathogenic Aspergillus, but we do this with others too.
So we found gliotoxin, now we're back to the same schematic.
Gliotoxin was actually this cluster.
Well, we wanted to start looking at other clusters that are
regulated by laeA to look at the compounds to see if they add to
pathogenicity. And this is our latest one.
We took one of those clusters and looked for the expression of
genes. This is gene expression in
Aspergillus fumigatus, and we find that in a wild type fungus
we have a series of four genes in a row that turn on at about
18 or 24 hours after the fungus grows, but if you remove laeA,
they're not expressed. What we have found, and this is
a graduate student in my lab, is that those four genes make a red
compound. You can see it's here in these
strains. But when you remove the genes
that make it, it disappears. And we found this compound, this
red compound, to be a compound called endocrocin.
It was not known that Aspergillus fumigatus made
endocrocin. Endocrocin is known, though.
Endocrocin belongs to a type of molecules called anthraquinones
coming from fungi that are used in the dye making system.
So these are swaths of cloth that have been dyed from
different anthraquinones from fungi.
Beautiful. And, indeed, Yun was able to
show that several of these genes make this red compound,
anthraquinone, and she is putting together a chemical
pathway. This probably is interesting to
maybe two or three of you. [LAUGHTER]
So we'll go by that. But what is most intriguing with
this, she wanted to ask, where is the anthraquinone made?
This endocrocin? And if you remember, I showed
you a few slides ago, that different compounds are made in
different tissues. So what Yun did is she grew the
fungus and then she would collect extracts from either the
auger the fungus grows on, the middle part which had mycelium,
the conidium stalks or just the conidia.
It's the conidia of Aspergillus fumigatus that come into your
lungs and cause the initial damage.
And you can see her results where she looked at extracts
from different parts of the fungus.
And what we see, this is the endocrocin here, two different
media, it is in the conidia that enter our lungs where most of
this endocrocin is formed. And what she's doing right now
is pathogenicity tests. She has made some strains where
she removed a gene required for endocrocin and looked at
survivability of these mutants she made compared to the wild
type. This time we're using a fly,
Drosophila model, and you can see from our initial data is
that endocrocin also appears to be a pathogenicity factor
because when you remove it, the flies survive longer.
Well, that wraps up my talk today, but I really need to end
by showing you the people who've done all the work over the
years. I am, basically, just an
observer now. I love going into the lab and
looking at the fungi, but I don't do any of the work.
These are just showing a lot of the people in my lab from this
summer. Graduate students, undergraduate
students, postdocs, visiting scientists.
So these are the people that deserve all of the credit for
what I just showed you. And that's it.
[APPLAUSE]