'Exciting biology' uncovers plants' high-fat diet for

fungal benefactors
One of biology's most enduring relationships, credited with helping plants
to colonise land more than 400 million years ago, has yielded a
fundamental survival secret with implications for agriculture and
biotechnology.

Plant scientists have discovered that a particular form of fungi, which

invades plant roots and then helps the colonised plants to absorb nutrients
from soil, receive life-sustaining carbon from their symbiotic hosts in the
form of long-chain fatty acids, a building block for essential lipids.
Previously, scientists had thought that the fungi received carbon from their
hosts only in the form of carbohydrates, which the fungi used to make their
own fatty acids and then the more complex lipids necessary for survival.
It's now clear that the main source of carbon from the host plants are fatty
acids, and that these fatty acids are necessary before the fungi can create
the more complex lipids that are needed for storing energy, for signalling
and for cellular membranes.

The latest work, published by the journal Science, comes from a joint team
of scientists from the John Innes Centre, based at Norwich Research Park,
and Rothamsted Research. This field of science is very competitive
currently, with research groups in China, Germany and the US all chasing
similar breakthroughs.

Professor Giles Oldroyd, project leader in cell and developmental biology at

the John Innes Centre, and co-leader of the research team said: "It has long
been thought that the plant delivered sugars to the symbiotic fungi. Our
research demonstrates that in addition the plant delivers lipids to the
fungus. We hope that through a better understanding of these plant/fungal
symbioses we may be able to improve their use in agriculture and thus
increase the sustainability of agricultural systems."

The symbiotic relationship at the heart of this research, and one of the most
widespread associations in nature, is that between the great majority of
plants, at least 80% of them, and arbuscular mycorrhizal fungi, which
create special feeding structures within the plants' roots called arbuscules.
The fungi, which develop hyphae to increase the roots' surface area, can
provide the plant with up to 80% of its nutrients from soil while the plants
can yield up to 30% of the carbon they derive through photosynthesis to the
fungi.

Professor Peter Eastmond, senior biochemist in the Department of Plant

Sciences at Rothamsted and co-leader of the research team said: "This is
exciting biology. We've discovered that the fungus is effectively re-
programming the plant to pump out lipids,"

"There are important implications for sustainable agriculture, particularly

in nutrient-poor soils where you need to make the most of resources," said
Prof Eastmond. "And also for biotechnology, in creating green pathways to
produce lipids in plants, for biofuels and for precursor chemicals for
industrial applications, as an alternative to fossil fuels."

Rothamsted began to investigate the relationship's metabolism in 2011

after ground-breaking genetic studies at the JIC had been the first to
identify and isolate two genes essential for sustaining the symbiosis, RAM
(Required for Arbuscular Mycorrhization)1 and RAM2.

"We grappled with understanding why these genes were so important until
we came up with the hypothesis that the symbiosis created a lipid factory in
the plant that fed the fungus," said Prof Eastmond. "This went against what
the literature said...we were proposing to overturn what's in the text books."
The team came up with a trio of robust and ingenious experiments that got
around the inseparable union to distinguish whether one or both of the
symbiotic partners were producing lipids. Each experiment independently
endorsed the hypothesis.

"There's a lot more work to do in following up this discovery," said Prof

Eastmond. "It will have a lasting impact on the understanding of the
metabolism of this symbiotic relationship and lead researchers in many
new directions."
Feeding fat to fungi: Evidence for lipid transfer in
arbuscular mycorrhiza

Confocal microscopy image of fungal arbuscules (green) inside plant root cells
(red). Credit: Dr. Maria J. Harrison, BTI

Nearly all organisms live in symbiosis with a vast, diverse array of

microbes. Arbuscular mycorrhizal (AM) symbiosis is the interaction
between plants and a group of fungi called Glomeromycota. Most land
plants, including several crop species, are able to interact with these fungi,
which have been long known to positively affect plant growth and nutrition.
The fungi live in plant roots where they elongate their tendrils (called
hypha) into the surrounding soil, like an extension of the root system, to
better access and transfer nutrients to the plant. In return the plant serves
the fungus food made during photosynthesis.

Previous research has shown that an AM fungal dinner at least consists of

high-energy sugars, but published today in New Phytologist,
researchers from the labs of Dr. Maria Harrison at the Boyce Thompson
Institute and Dr. Peter Drmann at the University of Bonn have produced
the first experimental evidence to suggest that AM fungi also get their
fat (lipids) from the host plant.

"Until recently, it has been assumed that the fungus obtains sugar from the
plant and can manage to make other essential nutrients itself," Harrison
explains. "However, this is not the full story."

The transfer of lipid may be just as important. Living inside a plant root,
the fungus will enter the root cells and create a tree-like structure called an
arbuscule, giving the symbiosis its namesake. As an arbuscule is formed,
new plant and fungal membrane must be generated, like the bark on the
extending branches of a growing tree. To do so both organisms ramp up
production of lipids, which are important building blocks for cell
membranes. Sure enough, plant root cells that harbor fungal arbuscules
increase their expression of genes involved in producing such
lipids by 3000-fold.

Taking it a step further, Bravo et al. showed that individually mutating two
of those genes, FatM and RAM2, in the model legume Medicago truncatula
not only disrupts the fungus' ability to form healthy arbuscules, but also
reduces plant production of a lipid necessary for fungal growth, called 16:0
-monoacylglycerol (MAG). This result indicates that FatM and RAM2 may
play specific roles in the biosynthesis of 16:0 MAG, and further highlights
the importance of this particular lipid for AM symbiosis.

What makes this food exchange even more important is the fact that AM
fungi can't synthesize 16:0 MAG themselves, indicating the fungi depend
on their host plant as the source of this molecule. The exciting discovery
that the plant provides the fungus with this essential lipid helps explain the
fungus' mysterious obligate nature, meaning it is completely dependent on
its host for survival and reproduction.

"Without the basic lipids that are obtained from the plant, the AM fungus
cannot produce the complex lipids that it requires to live," says first author
Armando Bravo.

One of the next steps in this research is to investigate how the plant
regulates 16:0 MAG production, and whether more fat for the fungus
means more nutrient transfer to the plant in return. In a world where
agricultural lands are being depleted of essential nutrients, AM symbiosis
may be the key to healthier plants and greater crop yield.
Symbiosis with mycorrhizal fungi provides plants
with enhanced access to scarce resources

Shrub-like arbuscules in root cells, stained with a fluorescent dye. Scale bar = 25
m. Credit: Priya Pimprikar

Symbiosis with mycorrhizal fungi provides plants with enhanced access to

scarce resources. LMU researchers recently discovered a mechanism that
may allow host plants to control the extent of symbiosis formation.
From cereals to trees: More than 80% of land plants form symbioses with
arbuscular mycorrhizal fungi. The fungi colonize the roots, and provide
their host with inorganic nutrients primarily phosphate which are
otherwise poorly accessible to plants. In return, the plants provide their
fungal guests with energy-rich carbohydrates. This type of symbiosis
evolved more than 400 million years ago and is indispensable for the
viability of many plant species. To enable the fungi to supply the host with
nutrients, their hyphal threads must first enter into the root, in which they
form branched tree-like structures known as arbuscules (from the Latin
arbuscula = a shrub).
The arbuscules then release the minerals, which the fungus has taken up
from the surrounding soil, to the plant. "The successful formation of such
alien structures inside plant cells demands a fundamental reconfiguration
of the metabolism of the root cells and must be tightly regulated by the
plant", explains LMU biologist Caroline Gutjahr. Together with the
members of her research group, Gutjahr recently identified one of the
crucial elements in this process. The findings have now appeared in the
journal Current Biology.
Perhaps not surprisingly, the degree of colonization of roots by the fungi
depends on the physiological status of the plant, including the level of its
actual requirement for the nutrients supplied by the symbionts. Thus, if the
plant already has access to an adequate supply of phosphate, formation of
arbuscules is actively inhibited. "However, up until now, no molecular
mechanism was known that might be capable of controlling the extent of
arbuscule formation in accordance with the plant's physiological needs,"
Gutjahr says.

Molecular nexus
In order to pinpoint mechanisms used by the host to control arbuscule
formation, the researchers focused on a mutant strain of the plant Lotus
japonicus (a species of legume, related to beans, peas and lentils), in which
the process is perturbed. "In this strain, we identified the gene affected by
the mutation as RAM1, the product of which is required for the activation of
other genes, and hence for the production of the proteins they encode", says
Gutjahr. "These proteins, in turn, are very probably required to permit
arbuscule formation to proceed. Their precise functions remain to be
characterized in upcoming projects."
Phys.org - News middle block
The RAM1 gene itself is strongly activated during arbuscule formation, so
the mechanisms responsible for its own activation are of great interest.
Gutjahr and her colleagues have now shown that two different regulatory
proteins are required to induce the gene. The first is the transcription factor
CYCLOPS, which was already known to play a key role in the regulation of
root symbioses. The second protein, called DELLA, was also familiar, albeit
in another context. DELLA forms part of a signal transduction pathway that
is activated by the plant hormones called gibberellins, which are essential
for the control of the plant physiology and growth. "To our surprise, we
found that, in the context of RAM1 activation, CYCLOPS and DELLA
interact with one another directly," says Priya Pimprikar, a doctoral student
in Gutjahr's group and first author on the new study. "With this interaction,
we believe we have identified, for the first time, one of the central nodes
upon which information relating to symbiosis on the one hand, and plant
physiology on the other, converges, thus enabling the plant to determine
the extent of root colonization in accordance with its current need for
phosphate, for instance", says Gutjahr.

Reducing the need for scarce resources

The symbiosis between plants and mycorrhizal fungi is also of considerable
economic and ecological significance, as intensive farming requires huge
inputs of artificial fertilizer each year to maintain the fertility of the soil.
These fertilizers contain mineral phosphates which are only available in
finite quantities, and some estimates suggest that reserves of these
compounds may run out within the next 100 years. "Arbuscular mycorrhiza
can help reduce the need for the application of phosphates in agriculture,"
Gutjahr points out. "And a better understanding of the mechanisms
underlying arbuscule formation is an important prerequisite for the success
of plant breeding efforts to enhance the efficiency of this type of symbiosis."

How plants interact with beneficial microbes in

the soil
Scientists have wondered for years how legumes such as soybeans, whose
roots host nitrogen-fixing bacteria that produce essential plant nutrients
out of thin air, are able to recognize these bacteria as both friendly and
distinct from their own cells, and how the host plant's specialized proteins
find the bacteria and use the nutritional windfall.
Now a team of molecular biologists led by Dong Wang at the University of
Massachusetts Amherst, working with the alfalfa-clover Medicago
truncatula, has found how a gene in the host plant encodes a protein that
recognizes the cell membrane surrounding the symbiotic bacteria, then
directs other proteins to harvest the nutrients. Details appear online in the
January edition of Nature Plants.
As Wang explains, plants often recruit microbes to help them satisfy their
nutritional needs, offering the products of photosynthesis as a reward. A
process used by most land plants depends on a symbiotic relationship with
mycorrhizal fungi. These form structures known as arbuscules that help
plants capture phosphorus, sulfur, nitrogen and other micronutrients from
the soil. This method is akin to scavenging, Wang says, because the amount
of nitrogen available in soil is quite limited.
By contrast, the less common process, found mostly in legumes, goes one
giant step further: it uses bacteria called rhizobia, which live in root nodules
and fix nitrogen from the air and make it into ammonia, a plant fertilizer.
Symbiosis with rhizobia means legumes can make ammonia by fixing
nitrogen in the air, which at 78 percent of the atmosphere, is "essentially
limitless," the biochemist adds.
Thanks to this feat, legume plants can get as much nitrogen fertilizer as
they need, rather than relying on often scarce nitrogen in the soil. This is
why beans are so nutritious, Wang notes. "The next time you eat your tasty
tofu or edamame, you have those little bacteria, and their 'marriage' with
legumes to thank."
"Talk to anyone in our field, and the dream is to make it possible for our
crops that can't fix nitrogen to get that ability," Wang suggests. "This
discovery moves us one step closer. Beans are special, but what our result
says is they are not that special because some of the basic infrastructure is
already there in plants that use arbuscular mycorrhizal fungi instead of
nitrogen-fixing bacteria, which no one understood before."
Phys.org - News middle block
The researchers led by Wang with postdoctoral researcher Huairong Pan
and doctoral students Onur Oztas and Christina Stonoha at UMass Amherst
plus others in China, discovered that in both processes bacteria and fungi
exchange nutrients with the plant across a cell membrane recognized by a
specially encoded protein made by the host plant that defines its borders.
"It's as if the plant has discovered how to create a sort of free trade zone,"
Wang says.
To explore how this trade works, the team, supported by a grant from
UMass Amherst, investigated activities of the gene SYNTAXIN 132, which
encodes receptors (SYP132) that identify cell membranes and interact with
secretory vesicles. They found that the gene usually makes one sort of
transcript that always seeks out the plant cell's surface membrane. But if
rhizobia are present in the host, that same gene will make a second type of
protein that is able to find the membrane surrounding the bacteria.
Surprisingly, symbiosis with arbuscular mycorrhizal fungi shares the same
SYP132 receptor. Scientists now understand that the host membrane - both
in legumes and beyond - around the fungal arbuscule has a lot in common
with the membrane around the nitrogen-fixing bacteria.
He adds, "The gene somehow knows the bacteria are present and makes the
alternative type of protein, which finds the membrane around the bacteria.
This means that the host can tell the two membranes apart, can sort them
out."
"Our discovery is that the two SYP132 proteins are not the same, even
though they come from the same gene. The gene makes two transcripts,
which involves an unusual process near the end of the gene, like a movie
with two different endings. Nobody knew there exist two proteins," Wang
explains. "So the answer is that one gene can use alternative terminal exons
to make two proteins with different sequences at the end. And it's the ends
that determine where in a cell the protein ends up."
He says the new knowledge he and colleagues uncovered is important to
future agricultural advances because the same process works similarly in
most plants, nitrogen fixing or not.

Evolution of root nodule symbiosis with nitrogen-

fixing bacteria
Nitrogen is essential for all plants and animals, but despite being
surrounded by itthe element constitutes 79% of air on earthonly a few
bacteria can absorb it directly from the environment. All other species are
ultimately dependent on these microbes as a source.
A new paper published this week in the open-access journal PLoS Biology
investigates the genetics behind the symbiotic relationship between these
nitrogen-fixing bacteria and plants, and presents evidence of specific
genetic changes that might have led to the evolution of symbioses with
nitrogen-fixing bacteria from a more ancient form of symbiosis.
About 80% of all land plants have a symbiotic relationship with fungi of the
phylum Glomeromycota. The fungus penetrates cells in the plants roots,
and provides the plant with phosphates and other nutrients from the soil.
This kind of symbiosis is called an arbuscular mycorrhiza, and evolved
more than 400 million years ago. Professor Martin Parniske and colleagues
started their study by looking at genes known to be involved in arbuscular
mycorrhiza, to see whether they could find evidence of any specific genetic
differences in plants that form symbioses also with nitrogen-fixing bacteria.
In this so-called root nodule symbiosis bacteria live in the root cells of the
host plants, where they bind elementary nitrogen from the air in special
organs, the nodules, says Parniske. In return, the microbes get high-energy
carbohydrates produced by photosynthesis in the host plant.
It had already been speculated that genes involved in the arbuscular
mycorrhiza symbiosis might have been recruited for nodulation, as these
symbioses both involve intracellular relationships. One clue was that
several genes, including the so-called symbiosis-receptor-kinase-gene
(SYMRK), are involved in a genetic program that links arbuscular
mycorrhiza and one form of bacterial nodule symbiosis. And the analysis of
SYMRK in several species of plant provided the striking evidence that
Parniske and his colleagues had been hoping for.
Our results reveal that an expansion of the functions of SYMRK
constituted an important step in the evolution of intracellular nodule
symbiosis, reports Parniske. Most plants have a short version of SMYRK,
which is required for AM symbiosis. A longer variant of SMYRK was found
only in plants involved in the symbiotic relationships with nitrogen-fixing
bacteria. Importantly, the longer version was found in both legumes (which
form symbioses with rhizobia the textbook nitrogen-fixing symbiosis) and
in actinorhiza (such as alder) which form symbiotic relationships with
frankia bacteria, about which there is little genetic information. The results
therefore suggest a common evolutionary origin of intracellular root
symbioses with nitrogen-fixing bacteria.
This work is an important step towards understanding the evolution of
nitrogen-fixation in plants, and even whether plants that dont form
symbioses with nitrogen-fixing bacteria could be engineered to do so, thus
increasing their nutritional value.

Researchers discover how bacteria sweet-talk

their way into plants

Root nodules, each containing billions of Rhizobiaceae bacteria. Credit: Public

Domain

A University of Otago researcher is part of an international team of

scientists that has discovered how legumes, which include important
agricultural species such as white clover, are able to tell helpful and harmful
invading bacteria apart.
Their study, which changes the understanding of carbohydrates as signal
molecules, is newly published in the leading international journal Nature.
The research has implications for improving the understanding of how
other plant and animal species interact with bacteria in their environment
and defend themselves against hostile infections.
Clover and other legumes form a unique symbiotic relationship with
bacteria known as rhizobia, which they allow to infect their roots. This leads
to root nodules being formed in which the bacteria convert nitrogen from
the air into ammonia that the plant can use for growth.
Exactly how these plants are able to distinguish and welcome in compatible
rhizobia for this self-fertilising activitywhile halting infection by
incompatible bacteriahas been a mystery.
Now Otago's Professor Clive Ronson and colleagues in a research team led
by Professor Jens Stougaard in Denmark at the Centre for Carbohydrate
Recognition and Signalling (CARB) have determined how legumes perceive
and distinguish compatible bacteria based on the exopolysaccharides
featuring on the invading cells' surfaces.
Professor Ronson says that exopolysaccharides, along with other
carbohydrates, are found on the cell surfaces of all organisms.
"Bacteria display a wide selection of polysaccharides, and many of these are
important for cell-to-cell interactions, immune evasion, pathogenesis,
biofilm formation and colonisation of ecological niches," he says.
Using an interdisciplinary approach involving plant and microbial genetics,
biochemistry and carbohydrate chemistry, the CARB researchers have
identified the first known exopolysaccharide receptor gene, called Epr3.
They found that a membrane-bound receptor kinase (a type of enzyme)
encoded by the Epr3 gene binds directly with exopolysaccharides and
regulates beneficial bacteria's passage through the plant's epidermal cell
layer.
"Now that we have identified this key mechanism that allows a host
organism to distinguish friendly bacteria from those that cause disease, this
opens up exciting new research avenues across a number of fields," says
Professor Ronson.
Microbiome studies in plants, animals and humans are some of the areas
that will benefit from the new discovery. The mechanism governing
microbiota colonisation of hosts is poorly understood and the identification
of an exopolysaccharide receptor is likely to inspire new approaches to
understand the interaction between multicellular organisms and microbes,
he says.