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Archaea and the Tree of Life

Transcript of Part 2: Mysteries of the Methanogens

00:00:03:01	I've always had a deep curiosity for,
00:00:06:17	I guess, logic puzzles.
00:00:09:08	Trying to understand how things fit together
00:00:12:02	and why they do so.
00:00:15:23	At some point in high school,
00:00:17:10	I learned about the fact that microbes
00:00:19:20	can eat all kinds of things
00:00:21:15	that other groups of organisms cannot.
00:00:23:21	So that kind of drew me to environmental engineering,
00:00:26:08	as an undergrad.
00:00:27:11	Two years into grad school,
00:00:28:14	I took a summer course in microbiology
00:00:31:15	and that just changed everything instantly.
00:00:35:05	I knew that I had to stop, kind of,
00:00:37:00	thinking about the world from an engineer's perspective
00:00:40:07	and transition over to become a biologist.
00:00:43:10	I ended up joining the Department of Organismic
00:00:46:14	and Evolutionary Biology at Harvard.
00:00:49:05	And when I was doing my PhD,
00:00:51:10	one thing I recognized was that
00:00:53:24	methane is mostly produced biologically
00:00:56:00	by this one group of organisms.
00:00:59:00	(bright music continues)
00:01:02:08	They produce 80% of the methane that's being released,
00:01:07:03	emitted to the atmosphere annually,
00:01:09:14	and which has a significant impact on climate change.
00:01:12:06	So decided I wanted to delve more into the biochemistry
00:01:15:01	and the metabolism of these bugs.
00:01:21:07	So Methanosarcina acetivorans was isolated
00:01:24:20	from these dense kelp forests
00:01:27:09	off of the coast of La Jolla in California.
00:01:30:06	So these kelp forests are so dense
00:01:32:18	that the insides are completely deprived of oxygen.
00:01:36:14	And for these methanogens to grow, that is a necessity.
00:01:39:04	They cannot grow in the presence of oxygen.
00:01:42:10	So in that dense kelp forest
00:01:44:24	is a place where there is no oxygen.
00:01:47:06	And in those places, there's a lot of carbon available.
00:01:51:07	It gives the methanogens the right kind of environment
00:01:54:04	for them to grow and then as a result
00:01:56:13	they were extremely abundant there,
00:01:58:20	and it was easy to find them.
00:02:00:12	So when I started my post-doc,
00:02:02:13	the CRISPR-Cas revolution had taken off.
00:02:05:04	There had been many groups that had developed CRISPR
00:02:09:00	technology for eukaryotic systems, you know,
00:02:11:14	ranging from Arabidopsis to mice.
00:02:13:21	Similarly, there had been some attempts
00:02:15:19	to develop CRISPR technology for bacteria
00:02:18:02	to edit their genomes.
00:02:19:18	There had been no such efforts on the archaeal front.
00:02:22:20	No one had tried to develop these tools
00:02:25:05	to work in an archaeal system.
00:02:32:07	So one the first things that I did was
00:02:34:07	that I tried to adapt CRISPR-Cas9 genome editing
00:02:38:05	to an archaea, to a methanogen
00:02:40:18	that I knew how to grow in the lab,
00:02:44:15	luckily it worked.
00:02:47:02	And that really enabled us to do a lot
00:02:49:05	that happened afterwards.
00:02:52:00	(bright music)
00:02:54:20	One of the things that I subsequently worked on,
00:02:58:01	after developing CRISPR for this particular methanogen
00:03:01:23	was to look at the genes involved in forming methane,
00:03:05:11	specifically an enzyme that's involved in methane formation
00:03:09:13	called methyl-coenzyme M reductase or MCR.
00:03:13:01	MCR is one of the most abundant enzymes in our planet.
00:03:17:03	And about 80% of all the net methane emissions,
00:03:20:12	on our planet, come from this particular enzyme.
00:03:23:04	But very little is known about how this enzyme
00:03:25:22	actually makes methane.
00:03:27:15	So we thought we could use CRISPR technology
00:03:29:23	to understand more about how this enzyme makes methane.
00:03:34:05	MCR is unusual in many different ways,
00:03:37:07	but the one facet that appealed to us
00:03:39:21	was the fact that this enzyme
00:03:42:02	goes beyond the 20 canonical amino acids
00:03:45:04	that we typically see in most proteins.
00:03:48:02	In MCR, many amino acids are modified
00:03:50:18	so that they look like something completely different now.
00:03:53:17	And what I wanted to understand
00:03:55:09	was why are there so many amino acids,
00:03:57:17	these building blocks for proteins modified in MCR
00:04:01:04	and are those modifications important for its function?
00:04:04:18	For instance, there is a very simple amino acid
00:04:07:05	called Glycine.
00:04:08:23	In one particular place in the protein
00:04:11:06	that Glycine had been modified to form
00:04:13:13	what we call Thioglycine.
00:04:15:17	And in principle, the difference between Glycine
00:04:17:12	and Thioglycine is the replacement of an oxygen atom
00:04:21:02	with a sulfur atom.
00:04:22:18	With Thioglycine it's extremely interesting in the fact that
00:04:26:06	we don't know of any other protein
00:04:29:03	or enzyme out there in nature
00:04:31:04	that has a Thioglycine in the protein.
00:04:35:09	So MCR up until now, it seems to be the only protein
00:04:38:19	that has Thioglycine instead of a regular Glycine
00:04:41:21	at that particular location.
00:04:43:23	So that was odd and unusual and extremely rare.
00:04:48:08	For the longest time researchers in the field,
00:04:50:06	who've noted this have considered
00:04:51:23	that Thioglycine kind of going beyond the Glycine
00:04:55:08	and modifying it must be really important for this enzyme,
00:04:59:11	possibly important for the formation of methane.
00:05:03:14	And so when we developed our CRISPR tools,
00:05:05:16	we thought, okay here's an interesting question.
00:05:07:15	Here's kind of this, you know, dogma in the field.
00:05:10:10	Why don't we go and look at what that Thioglycine does?
00:05:14:00	So to look at it, we identified the genes
00:05:16:12	that make those modifications,
00:05:18:01	that take a Glycine and make it into a Thioglycine.
00:05:21:11	And we modified those genes,
00:05:22:23	we deleted them.
00:05:24:06	And then we converted the enzyme from the form
00:05:26:16	where it has this unusual Thioglycine
00:05:29:15	back into a form that just has a regular Glycine
00:05:32:04	and looked at the differences between them
00:05:34:08	and try to ask the cell,
00:05:35:14	okay, how well are you growing?
00:05:38:00	How much methane are you producing?
00:05:40:09	And once we deleted those genes,
00:05:42:02	we essentially had cells that were making MCR.
00:05:45:20	But now that Glycine was no longer
00:05:48:14	getting modified to Thioglycine
00:05:50:21	and the cells were growing just fine
00:05:52:14	under most conditions that we tested and making methane.
00:05:57:04	And what that revealed to us was that
00:05:58:19	this Thioglycine is not that important
00:06:01:02	for the cells to make methane.
00:06:03:04	To some extent, we were kind of maybe disputing,
00:06:08:01	you know, a hypothesis that was well accepted in the field.
00:06:11:09	What we'd shown was the fact that
00:06:12:20	this enzyme just works fine with the regular Glycine,
00:06:15:24	which made us scratch our heads a little bit,
00:06:17:24	because what does it do?
00:06:19:07	Why go through the bother of making this,
00:06:22:00	you know, modification if it's not that important?
00:06:24:18	And to figure that out, we tried subjecting the cells
00:06:28:14	to a bunch of different conditions that we thought
00:06:30:12	were environmentally relevant.
00:06:32:12	And here I'm showing you one parameter
00:06:34:16	that often changes out in the natural environment,
00:06:37:05	which is temperature.
00:06:38:18	So we grew the cells up
00:06:39:19	at a bunch of different temperatures,
00:06:41:13	starting at 29 degrees Celsius,
00:06:43:14	which is on the lower end for these groups of organisms.
00:06:47:05	At these low temperatures,
00:06:48:11	we essentially detected no difference
00:06:50:06	between the wild type strain.
00:06:52:14	So the strain that has the modified Glycine,
00:06:54:19	the Thioglycine,
00:06:56:06	and the strain that has now just a regular Glycine
00:06:58:22	in its place, in green.
00:07:00:14	But as we increase the temperature from 29,
00:07:03:19	all the way to 45 degrees Celsius,
00:07:06:03	we noticed something interesting.
00:07:07:24	As the temperature increased,
00:07:09:15	we noticed that there was more and more of a growth defect
00:07:12:24	for the cells that did not have that Thioglycine
00:07:15:17	in their MCR anymore.
00:07:17:15	And what that indicates to us is that
00:07:20:09	as the temperature increases,
00:07:22:01	if you ever experience a high temperature stress
00:07:24:12	in the environment,
00:07:25:16	having that Thioglycine is really important
00:07:27:19	to keep your MCR happy and making methane.
00:07:31:07	And if you don't have it,
00:07:32:07	you probably don't do it as well.
00:07:34:10	So now our new hypothesis for this
00:07:36:12	Thioglycine modification in MCR
00:07:39:07	is that it actually helps the enzyme
00:07:41:17	stay stable and stay active under stressful conditions,
00:07:45:02	such as the one shown here,
00:07:46:17	that is high temperature stress.
00:07:48:20	So we started out with Thioglycine
00:07:50:17	because it was so unusual and so rare,
00:07:53:01	but when you look at MCR,
00:07:56:08	we find that in addition to Thioglycine,
00:07:59:00	it often has other amino acids
00:08:00:24	that also undergo these changes.
00:08:02:17	That also go beyond the 20 canonical ones
00:08:05:13	that we typically see.
00:08:07:06	And after this work, we followed up
00:08:09:10	on some of the other ones
00:08:10:16	and we're starting to see very similar patterns.
00:08:13:23	Most of the ones that we've studied so far,
00:08:15:20	and we've studied three so far,
00:08:17:19	are not crucial for this enzyme to make methane,
00:08:21:05	contrary to what we kind of believed for a while,
00:08:24:10	but they play important roles
00:08:25:20	under very specific environmental conditions.
00:08:28:17	Temperature is often one that we see
00:08:30:12	some interesting responses to that make us think
00:08:34:10	that maybe we were thinking of this incorrectly before,
00:08:37:23	you know, when an organism's growing out in the wild.
00:08:40:12	That organism experiences, extremely fluctuating conditions,
00:08:44:17	and especially if you're a unicellular archaeon,
00:08:47:18	you kind of have to have your defenses up
00:08:49:12	to make sure that you can make your way
00:08:51:02	through those conditions.
00:08:52:02	So if the temperature suddenly shifts,
00:08:53:21	you'd still need to grow and divide.
00:08:55:22	And we think now we're kind of changing our paradigm
00:08:58:22	and thinking now that these modifications to MCR
00:09:02:04	are more to adapt the enzyme to different environmental cues
00:09:07:02	like temperature, than for making methane itself.
00:09:14:06	So I spent my post-doc studying this one particular
00:09:17:05	methanogenic archaeon and developing tools for it,
00:09:21:04	but I'm extremely cognizant of the fact
00:09:22:21	that this is one particular strain, one organism,
00:09:27:23	and it's one amongst many that are found in the environment.
00:09:30:23	And what I'm trying to do in my lab
00:09:33:05	is trying to kind of, is being cognizant
00:09:36:00	of the diversity that's out there
00:09:37:15	and trying to see if we can leverage that diversity
00:09:41:00	in our understanding of methanogens and archaea.
00:09:44:03	It's an interesting time to be an archaeobiologist
00:09:46:23	because we're kind of in this, you know,
00:09:49:21	major transition in thinking about archaea
00:09:52:17	from an evolutionary standpoint.
00:09:54:22	And so that leads to interesting discussions
00:09:57:16	and conversations,
00:09:58:20	kind of at a big picture level about
00:10:00:20	what archaea mean and how we should study them
00:10:03:06	and what systems should we look at
00:10:05:09	to try to understand archaea?
00:10:07:05	I wish I could pinpoint one thing about archaeal research
00:10:10:15	that I find the most interesting.
00:10:12:01	I think what I find the most interesting about archaea
00:10:15:04	is the fact that there's so much.
00:10:17:03	They're such an understudied group of organisms
00:10:19:11	that I don't think there's one right question to ask.
00:10:22:12	I think there's so many things that we can look at.

This material is based upon work supported by the National Science Foundation and the National Institute of General Medical Sciences under Grant No. 2122350 and 1 R25 GM139147. Any opinion, finding, conclusion, or recommendation expressed in these videos are solely those of the speakers and do not necessarily represent the views of the Science Communication Lab/iBiology, the National Science Foundation, the National Institutes of Health, or other Science Communication Lab funders.

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