cc >> Cassie Immel: Good evening,
everyone. Thank you for watching
Wednesday Nite at the Lab. I'm Cassie Immel.
Tonight, I will be presenting Dr. Chris Hittinger from
the genetics department. He received his bachelor of
science from Southeast Missouri State University and his doctor
of philosophy from the UW Madison Laboratory of Genetics
and the Howard Hughes Medical Institute.
His research interests are
yeast, carbon divmetabolism networks. of
His research fields include gene
genetics, molecular genetics, and population or the evolution
of yeast and fungi. Without further ado, I present
Dr. Chris Hittinger. [APPLAUSE]
Chris Hittinger: Thank you
to tell you this story. verAs you can tell, it's anty
exciting story, and I've been asked to stay still so that it
films well. So I
script in that matter. apologize if I go off
involves a search for the missing progenitor of lager
yeast on five continents and
techniques of microbial ecology and isolated wild species out in
the field with your hands with the latest cutting-edge
techniques from genetics, and I think that's something we can
get excited about. And I
because, as Cassie told us, there are supplements to the
educational experience this evening during the Q&A session,
and I certainly want to thank the generous folks at Capital
Brewery for providing that, and I certainly don't want to be
accused of delaying our ability to supplement our education
as much as possible. [LAUGHTER]
So this is a talk about yeast,
of what yeast are. Yeast and that raises the question
So what are fungi? Well, are one-celled fungi.
major groups of eukaryotic they're one of the three
every day. And this orgais sort of a view ofter
all life on Earth and how it's related.
We have single-celled bacteria down here.
2.5 billion years ago, sorry, about two billion years ago,
two bacteria came together and
then gave rise to most of the big macroscopic large organisms
that we encounter, and the
and also organisms like you and I, animals, and of course fungi.
Interestingly, fungi are
animals than they are to actually more closely related to
evolutionary biologists ptypically represent thoseat
relationships by the cursor,
you could actually see at the wbase of that triangle thereh
where the fungi and the animals actually come together and share
a common ancestor in the
So fungi are further divided dawn of the Precambrian.
They're the ascomycetes and basidiomycetes, and we probably
have a lot more experience
least their larger forms, encouand that's mushrooms,s, at
also corn smut, wheat rust are basidiomycetes.
Ascomycetes are cup fungi and molds, notably penicillin and,
of course, the bread mold neurospora, and yeasts.
Now, you'll notice that yeasts are also present in
basidiomycetes because when we
a little bit complicated. aThere are actually many, manyis
different forms of yeast. And this is something that's
actually evolved multiple times.
really, what are yeasts? So the correct question is
lineages of fungi that have all evolved independently this
single-celled lifestyle of
So what are yeasts? They're beisingle-celled fungi.us.
They're microscopic organisms,
this complex and yet they're descended from
There are over 1500 recognized species of yeasts.
So, if there's one take-home I
their regular jobs or at home on hope people will bring back to
many, many kinds of yeast, and television it's that there are
In fact, genetically they're they're incredibly diverse.
vertebrates. So there's actuallan awful lot ofthan all
diversity locked within this group of organisms.
Also, in contrast with bacteria,
asexual reproduction. So they undergo both sexual and
with a partner or they can they can produce sexually
we see this budding yeast here reproduce more of themselves, as
dividing and making a copy, a genetically identical copy of
itself. Now, it turns out while yeasts
have evolved multiple times from
people are talking about a fungi, it turns out the usually
all share a common ancestor 500 parmillion to a billion yearsat
ago, and those are the
Hemiascomycete yeasts. By looking at these pictures,
you can usually see why they're called Hemiascomycetes, and it's
because a normal ascomycetes
sexual structure, and this is actually a whole collection of
sexual structures; whereas, the Hemiascomycetes usually only
form four spores. And so
Usually, yeast means one of these guys here, this little
part of the tree that branches
maybe 500 million years ago. off from the rest of the fungi
I'm embarrassed to say that we Well, what do yeasts do?
them, and I mean that stiin the broadest sense. of
We don't know what they're doing
we don't know what they can do out in nature on their own, and
ecological question and also an fointeresting biotechnologicalng
question. We do know
pretty strong associations with various insect species or plant
species. And the ones that we know the
most about, of course, are the
our pets sick, or ones thour crops sick.us sick,
And many of you have probably encountered yeasts going by the
name of Candida. These
sometimes called thrush, cause yeast infections
or vaginal thrush, and these in particular oral thrush
overgrowing and are causing an infection.east
Yeast also produce a number of
dear to many of our hearts and the products that are near and
more entertaining. Soy make our lives more palatable or
They're probiotics. Perhaps sauless popular in somee.
circles are Vegemite
If you ever read the ingredients and Marmite.
processed foods you'll see on the back of many of your
brewer's yeast to lots that they add torula yeast and
And, of course, why we're here and lots of things.
alcoholic beverages toward today is because of these
And, of course, I should note that the same process that are
used to make alcoholic
bread rise, is also the process bthat we use in the biofuele
industry today which is of
state and to the increasing importance to this
Yeast can also spoil food and beverages, and that's another
important consideration. So there's sort of a balance
between these products that we
and the products that regawe regard as spoiled.rable
And, in fact, there's some
what people considered spoiled cultural differences in terms of
One thing that yeast often seem to share is that many of them
like simple sugars, but really
readily, and that's what we're only some of them ferment
because it's easy fermentation going to talk about today
down here. So, in thatparticular, when we'rects
talking about the champion
yeast, and many of you are fermenters are Saccharomycetes
here that Saccharomyces famillikes to grow on.oducts
Saccharomyces likes to grow in
rich in the simple these places because they are
Glucose is the most abundant sugar glucose.
It fuels life in many ways. It's the only source of carbon
that our brain uses. And
bread, grapes, and of course the it's found at high levels in
Probably many of you don't wort that is used to make beer.
Saccharomyces yeasts are realize but in nature
with oak trees in the actuanorthern hemisphere.tion
And, in fact, when we go sample wild yeasts, we're actually
usually too lazy to scrape
sometimes, especially if we have off the bark, although we do
it's much easier and quicker the landowner's permission, but
and grab soil back and just to go under the oak tree
And so we often get bringyeasts this way.lab.
If you're immunocompromised,
make you sick. So the Sacsame unfiltered yeast youally
may consume one night, if you're
cases, cause an infection. And, imof course, as I mentionedome
earlier, yeasts are very
industry, principally right now making corn into ethanol, but
increasingly making other products into ethanol as well.
So Saccharomyces love glucose.
What does it do It gewith the glucose?aces.
Well, most of us, most
glucose, as we learned in high organisms, when they encounter
work of 19th century school biology and thanks to the
they check to see microbif there's oxygen.teur,
And if there is oxygen, they
fully into carbon respdioxide and ethanol.down
And they do this because they get lots and lots of ATPs, which
are the energy
Now, most organisms do have a fermentation pathway, but, as
you can see, they get very
So they really only like to use it when oxygen is not available.
The products from that, of
which is what makes beer fizz, course, are carbon dioxide,
product that some people are and also ethanol, which is the
interested in the cofizzing in root beer.'t
Now, if you don't believe that
here than here, I think all tyou would have to do is with
parental guidance, of course, is
and try to light that on fire to tversus a small amount ofter
ethanol. And I think
there is still plenty of energy present in the ethanol and not
many energy present in the water.
Now, interestingly, Otto
century, actually observed that many tumor cells do something
completely different. They
oxygen is present, and instead don't check to see if
through glycolysis they run glucose directly
Interestingly, Saccharomyces and ferment it.
If glucose levels are high, they yeast do exactly the same thing.
and make ethanol anyway. ignore the presence of oxygen
because they've lost the Interestingly, this is not
This is actually a regulatory decision that they make.
So they check to see if glucose is high enough that they can
make a living off of these ATPs,
shut down respiration and crank up glycolysis and fermentation.
Now, why would you do that? Well, this is faster because you
don't need to get oxygen into the mitochondria and go through
all the many, many, many
And so if you can wastefully steps of respiration.
possible, that can give you brief competitive advantage.
Not only that, ethanol is toxic,
among the principal especcompetitors of yeast.ch are
So you can claim as much of the resource as possible.
As a bonus you poison your
you are able to tolerate, and then you can actually, after the
resource has been cleaned with
back and respire it and get a tfew more ATP back later in theo
process. So, when we
looking for the missing ancestor started this project
there were six species of lager yeast, we knew that
And I've shown their of Saccharomyces.
And the genus Saccharomyces is a evolutionary relationships here.
originated probably about co10 to 20 million years ago.es,
And that means all six of these species shared a common ancestor
at this point in the past. Now, if we fast forward in the
future, we can see that new species emerge, and that's what
is occurring at each of these branch points.
And so Saccharomyces cerevisiae and Saccharomyces paradoxus
share a common ancestor at
Now, of course, yeast evolve, their DNA evolves at a much
faster rate than our DNA, and that's why when we actually look
at the level of genetic divergence we see that these two
closely related yeast species actually are about as related
as human and mouse are
And so by the time we go out to the most distant member of this
genus, Saccharomyces uvarum,
human/chicken. And so that's how we end up with
the comparison that yeast are more diverse than all
vertebrates. There's
captured in yeasts. And I've a lot of diversity
relevant for this talk here, are drawn, perhaps the most
these organisms like to grow the preferred temperatures that
because of the temperature, aSaccharomyces cerevisiae hasy
been the historical yeast of
for thousands of years, and cpeople have used it to makes
ales, wine, for distilling, to
more recent times leavento make biofuels.urse in
Now, this is a dangerous
your boss, but I was at siWashington University in St.th
Louis and then University of
my immediate supervisor and mentor, Mark Johnston, and he is
an ale man. We've had a rather longstanding
disagreement about the relative merits of these two products.
He is a traditionalist and
and as you can see, I'm endorsing a lager product right
here. I don't think I consumed all of
those myself. I'm sharing with my friends back
there. But ales, as I mentioned, have
been brewed for thousands of years.
They've been with us
Whereas, lagers were invented in a very long time.
comes from the 15th century Bavaria, and that
And, in fact, by the 16th century, Bavaria had actually
passed very strict purity laws
actually, by the middle 16th called the Reinheitsgebot, and,
brewing during the hottest century it had actually banned
The lagering became a worldwide phenomenon by the 19th century
as Germans went to many, many countries including our own
country, and, in fact, many of
over in the German my ancestors would have come
So what is an ale and a lager for those of you who have not
had this particular argument with your boss?
Ales are things like pale ales, IPAs which are India pale ales,
a cosh, ESB, scotch ale, porter, stout.
Lagers are things like pilsners,
in this country, Vienna style helagers, Schwarzbiers, bock.est
And if all these names sound
sound vaguely English, vaguely German and these mostly
That's another sort that'sof short-hand.ence.
Most German beers that are
be of German origin, and many of popular in this country tend to
country that are popular the English style beers in this
So another classical way that people have distinguished ales
from lagers is that ales are
and that it ferments vigorously and stays up in the water column
until the end of fermentation.
yeasts fall, crash out of solution and ferment along the
bottom. Sometimes that's called
sedimentation or flocculation.
minimally filtered, in Also, ales are often served
Whereas, lagers tend to be some cases unfiltered.
crisp taste. The other prized for their clarity and
think you'll note why we were paying attention on the tree to
the temperature preferences is
warm temperature. And this, that of course, makes senseairly
because much of civilization
and refrigeration would not grhave been widely available.ean
So it makes perfect sense that
which was commensurate with ythe temperature around you.ct
Whereas, lagers are brewed at a much colder temperature.
And, in fact, they go through a long-term storage process which
is the basis of the word lager. It actually comes from the
German lagern, which
And there it's stored near means to store.
help age and freezing for several weeks to
So ales are brewed with Saccharomyces cerevisiae, and
this is true 95-plus
Whereas, lagers we have know at least since the mid-'80s are
brewed for a hybrid of
something else. And that Saccharomyces cerevisiae and
topic of tonight's talk. So, something else is the
scientifically called Saccharomyces pastorianus.
It's alternate name is
named for the carlsbbrewery in Denmark.indeed
And this is a sterile hybrid of Saccharomyces cerevisiae and
something else. Now, when I say it's a sterile
hybrid, I mean it's much like a mule, a cross between a horse
and a donkey. It can
but that progeny is not longer able to reproduce.
So a mule is not, excuse me,
reproducing either with a horse or with a mule.
Yeast can reproduce asexually.
are donkeys which can reproducing asexually repeatedly
in the laboratory, and we call these lager yeast.
So based on the cursory genetic information that we started to
roll in in the mid-'80s, we knew that it was a hybrid of
Saccharomyces cerevisiae, and it
was a hybrid of something down turns out we also knew that it
One of these early branching in this part of the tree.
But it turned out that there was chickenish level strains.
part of the tree, and depending on which strain you compared to
which, you could get DNA
about in more detail later, ranging as high as 15% which is
actually greater
So there was a lot going on here, and that really raises the
question of how many wild species there were in this
complex. So we get
information in 2009 when a little bit of
associated with the Suntory Brewing Company published the
genome sequence, and that's the complete list of genetic
information for the lager yeast Saccharomyces pastorianus, and
that made it really clear,
genome sequences of all these other species that there was
something missing because it was
genome, it was 7% or so off from clear that every gene in the
calling Saccharomyces uvarum. So, again, nothing was matching
this other part of the
So why couldn't we find it? Well, it turns out that if you
look at the global funding for
scale each country by the amount research and development and you
research and development, you oget a world view that isn
somewhat different from the traditional geography.
[LAUGHTER] Now, a lot of this is research
for defense purposes and
and other applications. So research on biomedical research
important kind of research, research on yeast biodiversity.
[LAUGHTER] But it turns out that it's a
pretty good proxy,
research that goes on yeast biodiversity because if you look
at the five big strain
all in the northern hemisphere collections of yeast, they're
the countries you'd guess them and they're all pretty much in
research and to be in based on general
So they're all in the dnorthern hemisphere.
The biggest one is actually in
CBS. And we have a very nice one
right down I39 in Peoria, Illinois, that one of our
collaborators, Clet Kurtzman, runs at the USDA there.
So this project began as a
Portugal, -- more than five colyears ago looking for new in
species of Saccharomyces, and in
in a particular kind of ptransporter that they new wasd
present in lager yeast and they were looking for where they
could find this species in the wild.
And they were searching in
Europe, they also had collaborators in Japan they were
sampling with, and they've been to Oceana a couple of times.
Nothing. No Saccharomyces eubayanus, no
lager yeast. >> Is
Mongolia figures so prominently? there any reason while
China, and that is Japan. >> That >> That's Korea?that is
Yep. And then there's North Korea.
[LAUGHTER] So I began collaborating with
Professor Sampaio and Goncalves on another project actually,
also related to Saccharomyces biodiversity, and I've isolated
about 200 strains from
But, again, at least in wild the United States.
plausibly be settings, nothing that could
Now, again, I stress that people Saccharomyces eubayanus.
yeast all the time from industrial settings, but these
are not wild progenitor, these
by people and have specific genetic marks that tell us that
they're no longer wild, and we'll talk about what those are
later. Where we really
with our collaborator down in Argentina at the very tip of
South America in northwestern Patagonia, and Diego Libkind is
the one who, by spreading the search to a fifth continent,
actually finally hit upon a wild
rof yeast. So why Patagonia?es
Well, Patagonia is a
It's really pretty consistently cool environment.
It's average temperature is not too dissimilar from what we
experience here in Wisconsin. It
extremes that we do because it's doesn't have quite the Arctic
the oceans, but it's pretty a consistently cool with anby
average temperature of between
And we can heat samples instead of on oak trees, they don't have
oak trees in this part of the southern hemisphere.
Instead what they have are
actually only very distant southern beech trees which are
have in the northern hemisphere. relatives of the beech trees we
And much like we see on oak Thtrees, though, when they'rees.
wounded they'll produce this
and other yeasts will actually sap, and Saccharomyces yeasts
see the sap being ferfermented in these cases.can
But perhaps what's more interesting is they get infected
by another kind of fungus,
sartoria, and this fungus actually ends up in a tug of war
with the tree's immune system and ends up blowing up into
these massive galls, they're
actually becomes very high in sugar as the fungus tries to
exploit to resources from the tree and the tree tries to keep
things under control and keeps
And it turns out that because they become rich in sugar and
these are fairly soft structures, people in Argentina
will actually chop these off and
salads as a little chop them up and put them on
Apparently, in pre-Columbian bit of a sweetener.
actually take these guys off, harvest them, take them back to
their village, dipped them in water, and extract the sugars,
and of course what we now know
make a fermented beverage. to be Saccharomyces yeasts and
frequently from what I It's no longer made very
about its relative quality understand, so that may tell you
So how do we to what we have today.
I've talked a couple times about actually isolate it?
getting yeast from the wild. So what we do is we take bark ore
soil, and usually soil is more
off bark, and in the case of convenient but we can also chop
harvest the fungal parasite. the sartoria you can actually
laboratory and basically you Takes that back to the
ethanol in this case is basically a cheap antibiotic,
and sometimes we add antibiotics
because bacteria as present hundreds or thousand of fold
higher concentration
So by giving them sugar and ethanol, we can wait for couple
of weeks depending on the temperature of isolation, and
then as the yeast start to
fermentation and the media produce carbon dioxide from
pretty good indication that we've isolated something froma
the wild. And then we can take it back and
put it on petri plates and check
tetrads we talked about, those to see if it's forming those
can expose it to DNA himyasci, and if it is, then we
And based on sort of a fingerprinting.
can triage the strains and tell whether they are something that
we might be interested in or something that we're not
interested in. And it turns out, from the
Patagonian studies in particular, we were getting
predominately two species back. And so Diego preliminarily
identified strains as either
Patagonia, and, again, this is wild Saccharomyces uvarum from
the European Saccharomyces strains that looked kind of like
been sequenced, and also uvarum whose genome had already
eubayanus which is the candidate possibly this new species
yeast. And it turns for tout this actually lager
seems to be a preference for
growing on which species which species of yeast is
And this is statistically significant, and it's a little
easier to see if I
So you can see that uvarum highlight them for you.
whereas eubayanus likes realpumilio and Antarctica.beyi,
And, again, this isn't an absolute distinction, but
there's clearly some partition
niche partitioning going on as of resources going on, some
resources and the other prefers othe other set of resources.of
And so because we're finding
in sympatry in the same basic tarea, that sort of raises thee
question of whether they're really separate species, and,
indeed, which one actually
And so I've shown you evidence contributed to lager yeast.
in that they are that they are ecological species
We also crossed them in the lab and attempted to create the
hybrid, and it turns out when you do that, they can only
reproduce sexual spores 5% to 10% of the time, which suggests
that there's a substantial
In other words, they're substantially sterile much like
a mule is. Now, phylogenetic species, this
is more a question of whether they are actually exchanging
genes historically in the wild. And so this is a question where
applying the latest sequencing
way to get a look at technology is really the only
And so what we did is applied illumina sequencing which allows
us to sample 36 letters of DNA at a time.
And then we mapped all of those reads to the genome assembly
that we already had for the European strain of uvarum,
allowing a bunch of mismatches. And how this technology works is
truly amazing and really could
the Lab talk on its own. I'm fill an entire Wednesday Nite at
much. Suffice it to not gsay that when thet too
original Saccharomyces uvarum genome was sequenced 10 years
ago, it cost more than $100,000,
we now can do it for $100. and, using this technology,
technology that is going to, as we'll talk about at the end,
change the way you interact with your doctor in the future.
But just to give you a quick
when I say the DNA differs. runHere's an example of theean
Saccharomyces uvarum
This just happens to be a random European strains right here.
we look at the strains we're snippet from the genome, and if
look at a piece of DNA that gmatches the existing sequence,n
and as we see if you look across
and adenines and cytosines, at there's only four possiblenes
letters but we see that at every
And geneticists sometimes get polazy and it also helps us to.
view the data if we just put
And so this makes it really clear that for this stretch of
DNA they have exactly the same sequence of DNA letters.
Okay, what about our new strain,
Well, if we look at this same stretch of DNA, it turns out
that it's the same for a lot
differences. And in of this case there are few
actually five differences.
from both the European and And so it's clearly distinct
But it's similar enough that Patagonian strains of uvarum.
So, of course, if we then look we can still recognize it.
the lager yeast, it matches aperfectly our Saccharomycesm
eubayanus strain. It's
them in some places, but where the same as all four of
yeast Saccharomyces pastorianus talso differs and it differs inr
the same way. Now, just to show you that I
didn't cherry pick things, I'm
chromosome. Now, of goincourse, a chromosome the
goes on, in this case, for
you all 500,000 letters because we don't have time, and I would
lose count probably at number 124,000.
That's usually where
[LAUGHTER] But what we can do is we can
average and sort of look
And if we do that, we can measure, going up here, how
different each strain is
Saccharomyces uvarum, and then from our European strain of
from our new Patagonian species, how different each strain is
And if we do that, we can see that our new strain of eubayanus
is, indeed, quite distinct
On average, it's 7% divergent, from the uvarum that we see.
DNA letters it differs. Whereas, so that means that 7% of those
we're getting from Patagonia, uvarum, looks almost exactly
like its European counterpart.
it's very, very similar. And, Theof course, if we comparebut
it, again, to the eubayanus,
Okay, so if we lay the lager it's about 7% divergent.
that lager yeast is, of course, very distinct from uvarum, but
when we compare it to eubayanus,
zero line. And, it isindeed, the genomeso the
are 99.56% identical, so at more than 99.5% of the letters
they're exactly the same. So, we needed to look at a few
other strains associated with brewing and that's partially
because other groups had
candidates for having given asserise to lager yeast.ial
And it turns out what we see is that many of these strains are
really a mish-mash of genetic variance that probably came
together at some point after
of these wild alleles lagerinthat we're seeing.xtures
And this is a perfect example.
isolated from turbid beer in 1895, I believe in the
Netherlands, and this was actually designated as the type
strain of Saccharomyces bayanus,
official taxonomic specimen that which means that it's the
particular name bayanus. But is forever associated with this
uvarum alleles for hundreds of we see that it actually has
matches, and that it looks thousand of kilobases, exact
And, indeed, for a lot of veits genome it actually isus.
heterozygous in that it has both
And here, basically, an entire eubayanus and uvarum alleles.
We have one copy of eubayanus haland one copy of uvarum.re.
So if we look at another strain,
contaminate that was isolated in this is another brewing
also a hybrid. However, the '40it's a hybrid innism is
slightly different places. So here it has entirely uvarum
alleles. It's very
And over here, it's entirely eubayanus alleles.
Now, an important feature of
can see whenever the line goes plotting things this way is you
Similarly, the orange line, whenever it goes down here, it
also goes down here. And
really two alternatives here, much as I've drawn it, that is
an uvarum alternative and a eubayanus alternative for this
particular species complex
Because if there were three and alternatives or fourins.
alternatives, those lines would not necessarily have to move in
parallel. So if we
genome, you can see that if we look at the whole
pairs, we see exactly the loosame set of patterns.ase
We see that the orange and the
mixtures. They go up green strains are very complex
place. Interestingly, and dwe see thatr the
pattern that I just mentioned where whenever the green goes
down on top, it also goes down
are probably only on botttwo alternatives.t there
And the other thing we see is that the wild strains that we're
getting out of Patagonia, the red and the blue strains, they
basically hug the positions that they're supposed to be in.
They're not picking up
other species, and similarly, the purple lines basically track
exactly what the blue line, and that tells us, again, that
pastorianus is a hybrid of
Saccharomyces eubayanus. cerevisiae and this new species
with the genetic information Okay, so what else can we do
ancestor? Well, we can ask if
Saccharomyces pastorianus was domesticated or if it underwent
genetic changes after that sterile hybrid form.
And so we're going to be looking
that really stick out. We've for radical changes, changes
for really about a year and a half now, and so we haven't had
a lot of a chance to work with it.
We're going to look for places
strains where we can look at where it differs from the wild
contaminants and the those three brewing strains, the
look for obvious cases as to Saccharomyces pastorianus, and
code. And, in what'particular, the onesetic
we're going to focus on are cases where there might be extra
or transferred genes from Saccharomyces cerevisiae because
we know cerevisiae has been in
and other processes for use fthousands of years.making
And we're also going to look for
that have been inactivated very recently, possibly as a result
of domestication. So we looked at the genomes of
all of those strains on the previous slide for contributions
from other species, in
from cerevisiae. We paractually looked at allons
possible candidates, but it turns out we found no foreign
contributions in any of the uvarum strains or in our wild
Patagonian strains of Saccharomyces eubayanus.
We, of course, found that Saccharomyces pastorianus has
lots of genes from
It is, of course, a hybrid. But, interestingly, we found
that pastorianus had not picked up genes from any other species,
any of those other
However, we find that the green strain that was a brewing
contaminant and the bayanus type strain from turbid beer
115 years ago, it turns out that
multiple genes from tSaccharomyces cerevisiae.up
Now, you remember they are
eubayanus, but they're also mospicking up strains fromand
cerevisiae that are involved, sorry, genes from cerevisiae
that are involved in maltose
glucose-glucose disaccharides, consumption, which are
And this is, of course, at very high levels in wort, which
you'll note that one of the main
course, malt and that is four ingredients in beer is, of
There are also genes that these strains picked up from
cerevisiae that are involved
This is common table sugar that we all put in coffee in the
morning. That breaks
bonds. Some brewers will actually add
table sugar to their beers, so
and it's also present at fairly high level naturally in wort.
And, of course, in the brewing
number of cerevisiae genes that care known to be involved in a
sedimentation and flocculation, which is another trait that's
very important to the brewing process.
This is the process of settling out and creating a bottom
fermenting yeast. So this means that these two
strains, in addition to being mixtures of uvarum and
eubayanus, are actually triple
and also picked up hybrids in that they've gone on
And so these are really gecomplex organisms.e.
So if we look at the genes
cerevisiae, we can look and see if it's ever happening in the
same way twice because that could indicate that a single
genetic event happened and then it was advantageous and spread
to other species. And here's probably the most
striking example we found. And
chromosome. This is a thiscase where the fancyf a
genetic statistics are actually harder to read than the actual
sequence data itself. So
sequence data. And you I'll direct you to the
I've split out read alleles canand blue versions.re,
And you can pretty readily see
are in red, all of the eubayanus that all of the uvarum versions
Saccharomyces cerevisiae versions are in blue, and then
It's a third option. And is different still.
right here, is all of the eubayanus alleles that are
involved in brewing all the sudden switch over to the
cerevisiae version. You can see they're all the
same. Whereas, the
version is actually different still from uvarum but is much
closer to uvarum than it is to
very closely related cerevisiae because these are
So this is an example of a sister species.
rare and fused a Saccharomyces single genetic event that was
eubayanus chromosome, and cerethis rapidly spread,to a
through the population. Here's
a case where a eubayanus gene actually gets broken by the
domestication process. And
you the red versions for uvarum sand the blue versions forg
eubayanus. We see there are lots of dots
meaning it's basically the same.
orange arrow there's What we see here is at that
It's no longer a dot. It's actually a missing base.
and because of the way the DNA actually a missing base,
throws the entire right half of the screen out of frame, and so
the code is completely
having exactly the same protein sequences that are encoded for
by the DNA, it's completely random and completely different
and non-functional. And,
cases, exactly the same lesion again, it's, in all three
Now, this is an interesting gene that is breaking the sequence.
sulfate transporter. And because this is a low affinity
actually a patent on it turns out there's
a lot of, the other sulfate transporter from Saccharomyces
eubayanus. And this will actually increase
the production of sulfites, which are made from sulfates
by lager yeasts, and this particularly interesting because
it turns out that lagers
sulfites than beers actuaproduced by ale yeast.more
And this is probably one of the reasons that lagers are able to
age longer and retain their flavor better is because
sulfites are natural
Okay, so I'm going to briefly summarize what we think happened
genetically based on the
Of course, in Europe for data I've shown you.
been ale type Saccharomyces thcerevisiae that have beenve
partially domesticated. Brewers have been passaging them
from batch of beer
Eubayanus, at some point, fused with this ale type yeast and
created a hybrid yeast, a sterile hybrid that is a mixture
of cerevisiae and eubayanus.
changes that happened, two of Twhich I've talked about, thec
inactivation of this sulfate
up maltose genes from Saccharomyces cerevisiae, genesg
for breaking down maltose,
And, of course, in the brewing a common sugar.
reach tremendous levels, vats, the populations of yeast
Towards the end of the process, cells start to die and they
release DNA. And it turns out we can actually
do this process in
You can get transformation as strains pick up DNA from other
organisms. And, of
contaminate strains seem to include Saccharomyces eubayanus
once it made its way into the brewery, and it can pretty
readily pick up these partially domesticated genes from
pastorianus because it has
eubayanus that can recognize substantial portions of
And because eubayanus is so closely related to uvarum, they
share a common ancestor at about
know that Saccharomyces uvarum 7% genetic divergence, and we
frequency in Europe, we think a lot of these complex triple
hybrids informed as brewing contaminants crossed to wild
European strains of uvarum, and that's how we end up with
several of the triple hybrids.
form, here's what Okay, swe think happened.cartoon
Pastorianus is a hybrid of
course, eubayanus. The cerevisiae and ale yeast and, of
invented in the 15th century, but based on the date of the
rise of trans-Atlantic trade, the modern lager yeast strains
probably came into existence
became popularized but probably before the lager strains were
spread around the world in the 19th century.
And I've shown you genes
sulfite production, and sedimentation all show the sort
of hallmarks that we would
the selection of humans, brewers expect for genes that were under
competitive selection going on between brewing strains.
And, of course, I've also tried to explain how some of the more
complex brewing contaminant strains and some of the more
complex brewing strains seem to have formed, in some places,
pretty complex triple hybrids.
that while the species complex Hopefully I've also convince you
pretty complicated, it really boils down to probably two
naturally occurring species that
traditional ale yeasts and have then been crossed to the
Or to really, really boil things remixed around a whole lot.
[LAUGHTER] Okay, so I down, ithink I have a few that.
minutes left here, so I want
I've convinced you that it's not just cerevisiae,
Saccharomyces are important in other processes.
And now that we are starting to understand about the natural
diversities out there, we're
industrial processes in a new way and learn what is doing what
process. We now know, for example, that
lagers are made with hybrids
cerevisiae. I've also of eushown you now that'smyces
it's pretty clear that a number of the brewing contaminant
strains have substantial
There are also reports that champagne has contributions from
uvarum, although I think with genome technologies this
probably needs to be reexamined.
reports using not quite genome There are also a number of
it's solid enough that I believe scale data but close, at least
often actually hybrids involving it, and that Belgian beers are
Saccharomyces kudriavzevii, and there are also some wine strains
and cider strains that appear to involve contributions from
kudriavzevii. So this is another species that
appears to be involved in some
processes, and we're very interested in following some of
that work up. And, in particular, Belgian
strains are famously diverse, for those of you who are beer
aficionados, and we suspect that there may be a lot of
interesting complex things
As I mentioned, it's only some going on in Belgian stains.
and so we're very interested in tlearning about other Belgian,
strains, and also Weiss strains are also another interesting
mystery. And we actually
strains that we've recently generated genome sequence data
for from New Glarus, and we're
at those to see which vcontributions are coming fromok
where. And, again,
going to be able to do that because DNA contains this very
powerful information, and the powerful information it contains
can be used for that sort of ancestor determination.
And this is from this month's National Geographic from
researchers at the National Human Genome Research Institute,
Heidi Parker and colleagues.
they've determined the genetic And this is an example of how
breeds, and, as you can see, contributions to various dog
into four major groups based on which fraction of which genest
come from each of these groups. And, indeed, you can actually
pay a company commercially to go
if you have a mutt and determine out and look at your dog's DNA
which various dog breeds you with some degree of probability
So that could settle many arguments between spouses.
or start new ones. [LAUGHTER]
In all seriousness though, this can be done in humans also,
and there are companies that
something about your ancestry will for as little as $200, you
or will tell you about your susceptibility to various
diseases. And this is something I think we
should all be thinking of, and, again, this is a topic that
would go well beyond a
We could be here until Thursday or Saturday morning discussing
personalized medicine, but this is something that we're all
going to have to deal with, is
of sequencing a yeast genome that as I mentioned the cost
$100,000 to less than $100, and inhuman genome sequencing iser
rapidly getting to that point where all of us are going to be
able to, if we want, to sequence
implication of those variations, our human genome and discuss the
with our doctor. And our susceptibility to disease
important because many tumors have particular genetic
signatures and that can affect
particular kinds of cancer. So prognosis and treatment for
be thinking of. But this this is an area we should all
that's creating this revolution, same kind of technology
revolution, and that is in I hope will also create another
brewing. And, again, what I might call personalized
flavors I think. A, there are a lot of diverse
strains that are in use by
interesting characteristics. We can do the sort of thing you
can do with dogs and tell you where they came from, what they
are, but, of course, stains can be manipulated and crossed in
the laboratory, and if you want to mix variation I think there's
a lot of opportunity to do so in
the kinds of genetic a way that is now informed by
And so to summarize a lot of tools that we have.
groups, I also want to stress that all these species of yeast
have variations within species, much like humans are different
from one another, except the
So within a species of yeast, we have variation as high as
3.5%. Now, the human/chimp level
variation is about 1%. so these are extremely diverse
strains of yeast. There are hundreds of strains in
freezers around the world.
ethanol from sugar, and they're all a little bit different.
And, of course, this goes to
understanding their ecology and evolution, but I think brewers
and biofuel producers are really
can exploit this variation. goAnd, in particular, for the we
biofuel industry, the problem is this.
It's that our main workhorse
and while this is great for is producing beer, which ise,
starting with maltose, a
real problem because it creates ga competition for food becausea
corn is the principal way that people feed this chain.
Whereas, we really need to
products which use alternative be moving toward cellulosic
The problem is the organism that sugars like xylose.
is really pretty hardwired is best at doing this aspect
So we need to understand how it is using glucose better so that
we can understand how to get it to use other sugars and how to
exploit the variation that
preferences for xylose, for between glucose and xylose,
and other potential sugars. So
Jose Paulo Sampaio and Paula very briefly, I want to thank
for wild yeast as part of this Goncalves who began searching
in Europe. And, of project more than five years ago
who actually discovered Saccharomyces eubayanus down
in the wilds of Patagonia deep in the Andes Mountains.
Jim Dover for taming the bucking
Wash U and then at Colorado and bhelping make this cutting-edget
technology something that we're all going to be able to take
advantage of to do cutting-edge genetics for the next 5 to 10
years. And, of course,
for letting me work on lager Mark Johnston
to be working on ale. [LAUGHTER]
And so I think we'll take maybe one or two questions in here
and then perhaps go enjoy the full sensory array
of the educational multimedia presentation involving taste
and smell and other delights. So thank you.
[APPLAUSE]