(bright music)
I've always had a deep curiosity for,
I guess, logic puzzles.
Trying to understand how things fit together
and why they do so.
At some point in high school,
I learned about the fact that microbes
can eat all kinds of things
that other groups of organisms cannot.
So that kind of drew me to environmental engineering,
as an undergrad.
Two years into grad school,
I took a summer course in microbiology
and that just changed everything instantly.
I knew that I had to stop, kind of,
thinking about the world from an engineer's perspective
and transition over to become a biologist.
I ended up joining the Department of Organismic
and Evolutionary Biology at Harvard.
And when I was doing my PhD,
one thing I recognized was that
methane is mostly produced biologically
by this one group of organisms.
(bright music continues)
They produce 80% of the methane that's being released,
emitted to the atmosphere annually,
and which has a significant impact on climate change.
So decided I wanted to delve more into the biochemistry
and the metabolism of these bugs.
So Methanosarcina acetivorans was isolated
from these dense kelp forests
off of the coast of La Jolla in California.
So these kelp forests are so dense
that the insides are completely deprived of oxygen.
And for these methanogens to grow, that is a necessity.
They cannot grow in the presence of oxygen.
So in that dense kelp forest
is a place where there is no oxygen.
And in those places, there's a lot of carbon available.
It gives the methanogens the right kind of environment
for them to grow and then as a result
they were extremely abundant there,
and it was easy to find them.
So when I started my post-doc,
the CRISPR-Cas revolution had taken off.
There had been many groups that had developed CRISPR
technology for eukaryotic systems, you know,
ranging from Arabidopsis to mice.
Similarly, there had been some attempts
to develop CRISPR technology for bacteria
to edit their genomes.
There had been no such efforts on the archaeal front.
No one had tried to develop these tools
to work in an archaeal system.
So one the first things that I did was
that I tried to adapt CRISPR-Cas9 genome editing
to an archaea, to a methanogen
that I knew how to grow in the lab,
luckily it worked.
And that really enabled us to do a lot
that happened afterwards.
(bright music)
One of the things that I subsequently worked on,
after developing CRISPR for this particular methanogen
was to look at the genes involved in forming methane,
specifically an enzyme that's involved in methane formation
called methyl-coenzyme M reductase or MCR.
MCR is one of the most abundant enzymes in our planet.
And about 80% of all the net methane emissions,
on our planet, come from this particular enzyme.
But very little is known about how this enzyme
actually makes methane.
So we thought we could use CRISPR technology
to understand more about how this enzyme makes methane.
MCR is unusual in many different ways,
but the one facet that appealed to us
was the fact that this enzyme
goes beyond the 20 canonical amino acids
that we typically see in most proteins.
In MCR, many amino acids are modified
so that they look like something completely different now.
And what I wanted to understand
was why are there so many amino acids,
these building blocks for proteins modified in MCR
and are those modifications important for its function?
For instance, there is a very simple amino acid
called Glycine.
In one particular place in the protein
that Glycine had been modified to form
what we call Thioglycine.
And in principle, the difference between Glycine
and Thioglycine is the replacement of an oxygen atom
with a sulfur atom.
With Thioglycine it's extremely interesting in the fact that
we don't know of any other protein
or enzyme out there in nature
that has a Thioglycine in the protein.
So MCR up until now, it seems to be the only protein
that has Thioglycine instead of a regular Glycine
at that particular location.
So that was odd and unusual and extremely rare.
For the longest time researchers in the field,
who've noted this have considered
that Thioglycine kind of going beyond the Glycine
and modifying it must be really important for this enzyme,
possibly important for the formation of methane.
And so when we developed our CRISPR tools,
we thought, okay here's an interesting question.
Here's kind of this, you know, dogma in the field.
Why don't we go and look at what that Thioglycine does?
So to look at it, we identified the genes
that make those modifications,
that take a Glycine and make it into a Thioglycine.
And we modified those genes,
we deleted them.
And then we converted the enzyme from the form
where it has this unusual Thioglycine
back into a form that just has a regular Glycine
and looked at the differences between them
and try to ask the cell,
okay, how well are you growing?
How much methane are you producing?
And once we deleted those genes,
we essentially had cells that were making MCR.
But now that Glycine was no longer
getting modified to Thioglycine
and the cells were growing just fine
under most conditions that we tested and making methane.
And what that revealed to us was that
this Thioglycine is not that important
for the cells to make methane.
To some extent, we were kind of maybe disputing,
you know, a hypothesis that was well accepted in the field.
What we'd shown was the fact that
this enzyme just works fine with the regular Glycine,
which made us scratch our heads a little bit,
because what does it do?
Why go through the bother of making this,
you know, modification if it's not that important?
And to figure that out, we tried subjecting the cells
to a bunch of different conditions that we thought
were environmentally relevant.
And here I'm showing you one parameter
that often changes out in the natural environment,
which is temperature.
So we grew the cells up
at a bunch of different temperatures,
starting at 29 degrees Celsius,
which is on the lower end for these groups of organisms.
At these low temperatures,
we essentially detected no difference
between the wild type strain.
So the strain that has the modified Glycine,
the Thioglycine,
and the strain that has now just a regular Glycine
in its place, in green.
But as we increase the temperature from 29,
all the way to 45 degrees Celsius,
we noticed something interesting.
As the temperature increased,
we noticed that there was more and more of a growth defect
for the cells that did not have that Thioglycine
in their MCR anymore.
And what that indicates to us is that
as the temperature increases,
if you ever experience a high temperature stress
in the environment,
having that Thioglycine is really important
to keep your MCR happy and making methane.
And if you don't have it,
you probably don't do it as well.
So now our new hypothesis for this
Thioglycine modification in MCR
is that it actually helps the enzyme
stay stable and stay active under stressful conditions,
such as the one shown here,
that is high temperature stress.
So we started out with Thioglycine
because it was so unusual and so rare,
but when you look at MCR,
we find that in addition to Thioglycine,
it often has other amino acids
that also undergo these changes.
That also go beyond the 20 canonical ones
that we typically see.
And after this work, we followed up
on some of the other ones
and we're starting to see very similar patterns.
Most of the ones that we've studied so far,
and we've studied three so far,
are not crucial for this enzyme to make methane,
contrary to what we kind of believed for a while,
but they play important roles
under very specific environmental conditions.
Temperature is often one that we see
some interesting responses to that make us think
that maybe we were thinking of this incorrectly before,
you know, when an organism's growing out in the wild.
That organism experiences, extremely fluctuating conditions,
and especially if you're a unicellular archaeon,
you kind of have to have your defenses up
to make sure that you can make your way
through those conditions.
So if the temperature suddenly shifts,
you'd still need to grow and divide.
And we think now we're kind of changing our paradigm
and thinking now that these modifications to MCR
are more to adapt the enzyme to different environmental cues
like temperature, than for making methane itself.
So I spent my post-doc studying this one particular
methanogenic archaeon and developing tools for it,
but I'm extremely cognizant of the fact
that this is one particular strain, one organism,
and it's one amongst many that are found in the environment.
And what I'm trying to do in my lab
is trying to kind of, is being cognizant
of the diversity that's out there
and trying to see if we can leverage that diversity
in our understanding of methanogens and archaea.
It's an interesting time to be an archaeobiologist
because we're kind of in this, you know,
major transition in thinking about archaea
from an evolutionary standpoint.
And so that leads to interesting discussions
and conversations,
kind of at a big picture level about
what archaea mean and how we should study them
and what systems should we look at
to try to understand archaea?
I wish I could pinpoint one thing about archaeal research
that I find the most interesting.
I think what I find the most interesting about archaea
is the fact that there's so much.
They're such an understudied group of organisms
that I don't think there's one right question to ask.
I think there's so many things that we can look at.
(bright music)