Research

Bad Boys of Biology Turned Good

November 17, 2020 By Lauren Hines in Research Tags: reactive oxygen species

Yosef Fichman, post doctorate fellow in the Mittler lab, walks through how the lab uses arabidopsis plants for certain experiments. | photo by Lauren Hines, Bond LSC.

By Lauren Hines | Bond LSC

Shaking a bad rap can be hard.

However, the Ron Mittler lab at Bond Life Sciences Center has shifted the scientific community’s thinking from seeing reactive oxygen species (ROS) as destructive to vital.

“It’s always a challenge when you do something for the first time, and people don’t always believe you,” said Mittler, principal investigator. “They want you to prove it.”

ROS are reactive molecules that perform chemical reactions easily. While these molecules cause destruction in cells with these reactions, they can also send widespread messages and regulate many essential processes. However, this benefit of ROS wasn’t always known.

Back in the Day

In the 1980s, the scientific community recognized ROS as important. But according to Mittler, the community also thought ROS only offered destruction to human cells and caused aging.

“All this is not true,” Mittler said. “It’s not that simple to just say that, but there was a big hype about reactive oxygen species being painted as the bad boys of biology, and so forth.”

Then, in 2009 Mittler discovered the ROS wave.

This wave fills in the communication gap in plants left by the lack of something like a central nervous system. Instead, plants use the ROS wave and systemic signaling to respond to stresses. ROS triggers a cell reaction in one area of the plant that triggers the cell next to it and so on creating a wave of activation throughout the plant. This way, the plant can acclimate to its environment and is important since plants don’t have the luxury to get up and move when something threatens them.

Mittler was at the University of Nevada in Reno studying responses to ROS. He was originally looking to develop a reporter system to screen for mutants that are deficient in sensing ROS.

“We noticed that we don’t only record local responses that when we treated one single leaf, we got a systemic response, and, from there, using additional experiments, we developed the ROS wave concept,” Mittler said.

Mittler published his new discovery in Science Signaling, and more papers came, exploring systemic signaling and similar processes, highlighting the importance of the ROS wave to many biological responses.

“Slowly people started to realize that reactive oxygen species are actually important, and they’re doing a good thing,” Mittler said.

What’s Happening Now in the Lab

More than 10 years down the road, Mittler still focuses on ROS. Current Mittler lab post doctorate fellow Yosef Fichman, has taken the baton in the study of ROS waves.

“I was born to a family that was working in agriculture, and my father grew oranges in Israel,” Fichman said. “For a farmer or a person working in agriculture, life revolves around growing plants 365 days a year. In a way, I was always fascinated by plants.”

Fichman and others in the lab have been working towards finding more pieces to the process.

“There is a cliché that says proteins never work alone, so we’d like to know which other ingredients are involved,” Fichman said. “In my latest paper, using mutant plants, we were able to detect more ingredients to this mechanism and expand our knowledge.”

Fichman and others in the Mittler lab worked to image the ROS wave, tested different stresses for the ROS wave to respond to and found more pieces to the puzzle. Fully understanding this process can help researchers eventually create genetically modified crops that withstand more stresses.

What There’s Left to Discover

Currently, researchers don’t know exactly which compartment of the cell is most important in regulating this process or if compartments interact with each other. They also don’t know how it all results in gene expression or how different waves interact with each other and work together.

“There’s a lot to learn because we know that if we just look at this from the narrow window of the ROS wave, we know that it’s important,” Mittler said. “If you don’t have the ROS wave, the plant cannot acclimate properly.”

Though Mittler lab researchers have been hard at work for the past few years dissecting this process, it’s unlikely they’ll stop anytime soon.

“In a way, it’s harder to publish new ideas, but it’s more exciting,” Fichman said. “We are grabbed by our curiosity and we want to find the answer. We want to find the big answers.”

Find more details about Mittler’s current research in the most recent publication found in the Plant Cell journal.

Unknown Origins

November 9, 2020 By Lauren Hines in Research Tags: NASA, RNA

$5 million grant awarded to study RNA’s place in start of life on Earth

In his lab at Bond LSC, Donald Burke-Agüero examines his model of the RNA protein structure. | Photo by Lauren Hines, Bond LSC

By Lauren Hines | Bond LSC

The search for life on other planets may seem quite literally out of reach, but the search actually starts here, on Earth.

“Something that guides people in thinking about how life might come about on other worlds is trying to understand how life might have started here on this planet,” said Donald Burke-Agüero, principal investigator at Bond Life Sciences Center and professor of Molecular Microbiology & Immunology in the MU School of Medicine.

The Burke lab recently won a $5 million Interdisciplinary Consortium for Astrobiology Research (ICAR) award from NASA in partnership with eight other institutions. The award will fund their proposal to study RNA as a model for the origins of life.

Scientists hypothesize that RNA, a nucleic acid, was at the heart of molecular processes when life started around 4 billion years ago. RNA is like DNA in that it is a set of instructions and can store genetic information about basic building blocks for life, but it can also perform chemical reactions like enzymes. These two functions help speed up reactions to make important molecules needed for life.

“Becoming part of this has been over 20 years in the making for me,” Burke said. “It’s a theme that I care about tremendously, which is providing a really solid experimental foundation for some of the speculation about how life may have started.”

Burke found out about the grant while he was in another zoom meeting.

“I went back to the meeting and said, ‘Guys, I can’t stay. I’m not paying attention to anything you’re saying right now. Have fun. I’m bouncing off the walls,’” Burke said.

The predecessor to the ICAR program was the NASA Astrobiology Institute, which first called for members in 1998. Burke was starting his first faculty position back then. He always wanted to be a part of it, but it never quite worked out until now.

The “RNA World Hypothesis” has intrigued and driven scientists since the 1960s but has been increasingly explored in the past three decades as technological advances have allowed scientists to look for evidence to support it.

“Part of being in the origin of life team is working with RNA and to expand its known limitations,” said Jordyn Lucas, a Ph.D. student in the Burke lab. “We also like to explore how various modifications could make RNA better at doing a specific job, or how can it make it more durable under certain conditions.”

While RNA can make some reactions, it is possible that a certain set of modifications, tools or conditions could push RNA to perform more like an enzyme. This grant will help the Burke lab find those conditions if there are any.

“It’s kind of like a tool,” Lucas said. “It’s like if I gave you a piece of paper, asked you to write an essay, but I didn’t give you a pen. You can’t do it. It’s not because you don’t have the ability to do it. You just don’t have the right tools.”

Those tools can be hard even for humans to access.

A tube the size of your fingertip can contain quadrillions of RNA molecules and therefore quadrillions of sequences. That’s not an exaggeration. You can imagine how difficult it would be to sift through all that by testing them one by one. It wasn’t until in vitro evolution — a method that screens through large amounts of nucleic acid sequences — in the 1990s that scientists could explore many questions about RNA. Even with that, today’s technology is limited.

“Part of what this grant is going to do is attack that problem to try to specifically do the things that have been technologically very difficult,” Burke said.

The aim of the proposal is to push the limits of what RNA is capable of doing. These findings could bring the RNA World theory closer to being an established fact. This would help NASA know what conditions to look for life on other planets.

The technological advances from this project will also have implications for synthetic biology applications like biomedicines and biofuels. The goal is to find ways in which parts of modern biology can be replaced with RNA that can do something similar.

“Obviously, understanding our place in the universe and how we came about is a big deal and would be a societal benefit, but also a tangible thing that helps somebody be healthier,” Burke said. “That’s important too.”

Further exploring RNA brings us one step closer to a model of the origins of life on Earth and even life on other planets.

“We have no idea how likely life is,” Burke said. “If we can help demonstrate some of the ways that life comes into existence, it will really help to understand how many neighbors we might have on other worlds. And whether we’re likely to have any neighbors at all.”

Institutions involved in the proposal are the University of Missouri in Columbia, St. Louis University, Purdue University, University of Southern Mississippi, University of California in Irvine, University of California in San Diego, University of Washington in Seattle and University of California in Los Angeles.

NASA Selects New Science Teams for Astrobiology Research

Shining Light on Plant Reaction

November 5, 2020 By Lauren Hines in Research Tags: plant signaling pathways, reactive oxygen species

Arabidopsis grows in Ron Mittler’s lab. | photo by Becca Wolf, Bond LSC

By Lauren Hines | Bond LSC

Daylight might not seem dangerous, but for plants, too much daylight can cause hazards similar to a nasty sunburn or a human scalding themselves.

When you jerk your hand back from a boiling pan or a hot faucet, in a millisecond electrical signals are sent up your arm to tell your brain to move away. Like this reaction, plants have their own protective reactions to too much light.

“I think what you need to understand is that plants can’t survive without light,” said Ron Mittler, principal investigator at the Bond Life Sciences Center. “Light is the master of the plant basically.”

Mittler’s lab discovered that light receptor phytochrome B is responsible for triggering the signals for light danger that reverberate from one cell to another and not chloroplasts as previously thought. Understanding how plants respond can help build a picture of a process called systemic signaling to help researchers create genetically modified crops to withstand these stresses. Their results were recently published in the Plant Physiology journal.

The scientific community thought if light was put on a plant, the organelles that conduct photosynthesis, called chloroplasts, would trigger systemic signaling and the ROS wave.

“It’s really about having this spreadsheet of knowledge and not knowing where the pieces of the puzzle from high light-reactive action to chloroplast or to stomatal closure, where all those pieces fit,” said Mannie Liscum, collaborator and Mizzou professor of biological sciences.

The ROS wave, named by Mittler in 2009, is the process where reactive oxygen triggers a cell reaction that subsequently triggers the cell next to it and so on creating a wave of activation. Reactive oxygen is a tricky molecule that reacts very easily. It can cause destruction in cells with these reactions or use its reactivity to send a message in a proactive way between many cells.

“What we found is that if we use a phytochrome mutant — phytochrome B in particular — we completely eliminate this ROS production by high light stress,” Mittler said. “To me, it was really amazing because everybody else thought that the light would impact the chloroplast.”

While chloroplasts absorb light to turn into energy, phytochromes can take certain types of light and turn genes on. Depending on the type of phytochrome, they can take in different wavelengths of light and help plants understand light intensity and light quality. In turn, this helps a leaf or a stem figure out where to grow and if light is coming through shade.

The Mittler and Liscum labs used the sensitivity of the light sensor to their advantage.

“By using different wavelengths, different colors of light, we started narrowing down which of the sensory systems in the plant were actually doing the job,” Liscum said.

Eventually, they found responses to red light. This meant that phytochromes, especially phytochrome B, was responsible.

However, just like how we all learn to not touch the hot stove after our first burn, researchers also found that the ROS wave triggered a systemic stress memory mechanism. That meant the plant became familiar with the stress and its response was stronger.

“Evolution has realized that these high light intensities don’t just show up and instantaneously leave, but that they might stay around for a while,” Liscum said. “Evolution is adapted to have some system for the chemistry in the plant to change for some period of time before it goes back to what it was before it saw the stress.”

While the paper revealed much about phytochrome B and the systemic response to light stress, the Mittler and Liscum labs want to further investigate the memory and mechanistic aspects of the process.

“The next stage is to figure out how the sensing of the light stress by the receptors and not by the chloroplast triggers this response,” Mittler said.

Seed size matters: searching for a gene to make a bigger soybean

November 2, 2020 By Becca Wolf in Research Tags: CRISPR, gene expression, soybean

Bing Stacey | photo by Mariah Cox, Bond LSC

By Becca Wolf | Bond LSC

Patience is a virtue, at least it is for Bing Stacey.

Stacey recently completed a project that took her a total of eight years. It took her five years to develop a fast neutron mutant population and it took an additional three years to screen the population to identify a mutant that showed increase soybean seed size and then identifying the causative gene.

This gene, GmKIX8-1, and the seed size QTL, qSW17-1, can potentially be exploited for increasing yield in soybeans. Being able to alter these to increase seed size is important to improve the economic traits of soybeans such as yield and seed quality.

“The most important thing for farmers is the yield,” said Stacey, assistant professor of plant sciences at Bond Life Sciences Center. “For farmers, they plant soybeans, and the value or the profit they get is based on yield.”

There are two components of yield: seed size and seed quality. Stacey went to work finding what genes contribute to these components.

Mutants are very important for gene study because they have a new DNA sequence for genes. While it sounds like a sci-fi term, mutants are just plants manipulated using chemicals, radiations, or genetic modifications to change their characteristic traits in comparison to the norm. And the DNA changes to the GmKIX8-1 gene Stacey found is exactly the mutation she was looking for.

When she began this research, there were not many mutants for soybeans regarding seed size, so she had to develop her own mutant population using fast neutron irradiation.

After taking several years to develop this population, Stacey eventually found several mutants showing altered seed traits, including one showing increased seed size.

“So, we have the mutant, and then we utilized a fast and cost-effective way to genotype for changes in the DNA sequence of the mutant that is associated with the increased seed size. The genotyping method is called Comparative Genome Hybridization (CGH) which can be completed within one week and specifically detects missing DNA sequences in the mutant genome,” Stacey said.

Next, Stacey used CRISPR/Cas9 mutagenesis, which is a very powerful way to cut out specific DNA sequences in targeted genes in an organism. Before CRISPR, plant scientists could only create random mutations in plants. For example, using processes involving mutagenic chemicals and radiations, which meant one had to look at a very large number of mutants to screen for induced changes in the DNA sequence of specific genes.

Using fast neutron mutagenesis which creates deletions in bases, combined with CRISPR/Cas9, Stacey was able to characterize the role of the GmKIX8-1 gene in controlling seed size in soybeans.

“In a way, we got lucky because there is a QTL overlapping GmKIX8-1,” Stacey said. “And once you know the genetics behind a trait, then it is possible to define a possible mechanism of how the gene works, for example, how it makes soybean plants produce bigger seeds.” GmKIX8-1 controls the ability of cells to multiply. It also increases the size of the organs and the seeds of soybeans, which is exactly what Stacey was looking for.

An additional new and important thing Stacey found in her work was that GmKIX8-1 controls seed size, but not leaf size, in a dosage-dependent manner indicating that this gene is a major player in regulating seed size but not leaf size.

“This gene dosage effect on seed size and the identification of GmKIX8-1 as the causative gene behind a major seed size QTL are the novel aspects of this work,” Stacey said.

These novel aspects have been rewarding for Stacey.

“I’m quite happy, and it’s more fulfilling discovering something novel, something that hasn’t been reported before in other plants, instead of confirming what has been already reported,” Stacey said, “For example, based on what is already well-known in maize, or in rice, or in Arabidopsis, I can ask the question, ‘Does this gene also work in soybean?’ Answering this question is still worthwhile to me because maybe the gene will not work at all, or maybe it works in a different way, and that would also be novel, but to discover new genes or new explanations on how a gene works is more fulfilling.”

For more information on Bing Stacey’s work, check out the September 2020 edition of the New Phytologist Foundation article, “GmKIX8‐1 regulates organ size in soybean and is the causative gene for the major seed weight QTL qSw17‐1”

Using improved technology to create an Immune Cell Atlas

October 28, 2020 By Becca Wolf in Research Tags: agriculture, gene expression, RNA

Ashley Meyer | photo by Mariah Cox, Bond LSC

By Becca Wolf | Bond LSC

When given an opportunity to use the newest technology, one has to take it.

Ashley Meyer, the lab supervisor of the Wes Warren lab at Bond Life Sciences Center, recently started using the improved technology of single cell RNA sequencing to create an Immune Cell Atlas for chickens. This search browser of sorts allows people to search for cells and gene expression in chickens.

“It would be a platform for researchers to have a baseline of what should be normal, and then be able to run comparative experiments to say ‘Okay, if we challenge this chicken with this disease, what biological response are we seeing?’ Which can help you then understand the disease better,” Meyer said. “Like, ‘How is it being transmitted? How is this chicken reacting to it?’”

Poultry is one of the largest agricultural industries in Missouri, so it is important to keep chickens healthy.

“The Immune Cell Atlas will help us analyze tissues and will help us perform the single cell RNA sequencing on them to ask what’s normal and what’s diseased so that we can see what’s happening,” Meyer said. “It will also allow us to see which chickens are more susceptible than others or how the progression diseases happen.”

Meyer and the Warren lab are some of the first researchers to apply this technology to chicken based research.

Single cell RNA sequencing (RNAseq) allows Meyer to sequence the RNA of an individual cell which provides in-depth data. This can be done using single cell RNAseq which uses the entire cell, or it can be done using single nuclei RNAseq which uses just the nucleus of a cell. This profiles gene expression in cells that make up a tissue to get data on a cellular level to understand the composition of cells.

It is important to target the RNA because it is the messenger that brings instructions from the DNA for controlling the synthesis of proteins. RNA is commonly synthesized in the nucleus of a cell.

“We’re some of the first people to get our hands on some avian single cell data, specifically looking at nuclei. My job is a lot of optimizing these protocols to isolate these nuclei,” Meyer said.

Using nuclei allows Meyer to use samples that have been frozen, collected in other locations, etc., which provides more flexibility.

Isolating the nuclei through RNAseq allows Meyer to identify different cell types based on genes being expressed in a cell in an organ such as a spleen or lung. This information can then be added to the Immune Cell Atlas.

Meyer is a part of a team working on this project including two master’s students, a bioinformatics analyst, and principal investigator, Wes Warren, at Bond LSC, as well as a bioinformatics analyst who works remotely from Santa Cruz, California.

While the team is working from around the country, they are all connected by a common goal.

“Things have been pretty hectic, the break with the COVID quarantine kind of paused everything and then when we got back it just kind of took off, we’ve generated a couple data sets just in the last month or so,” Meyer said.

Having started this research about six months ago, they still have a way to go. Meyer and her collaborators are currently focused on analyzing five to six immune system organs, and plan on expanding in the future.

Since single cell RNAseq is so new, it can be challenging being one of the first labs to use it at MU and in the avian community.

“Since it’s still newer, everyone’s still figuring out how it works,” Meyer said. “Everybody’s determining their own way of analyzing the data. We’re working through the hiccups of everything, and not having years and years of research to guide us can be challenging; however, also exciting because then we can find what works and what doesn’t work which is the whole point of science.”

While challenging, it is rewarding. Meyer hopes that in the future once they have created the chicken Immune Cell Atlas, they are able to expand to other animals.

“It’s exciting to be part of something that’s newer and having something that could be beneficial for other researchers,” Meyer said. “I am a huge advocate for sharing science and teaching science so creating something to aid other research is super empowering for me.”

Learning From History

October 26, 2020 By Lauren Hines in Research Tags: extracellular ATP, plant signaling pathways

Sung-Hwan Cho, a research scientist in the Gary Stacey lab, checks in on the arabidopsis plants that he uses in his purinergic signaling experiments. | photo by Lauren Hines, Bond LSC.

By Lauren Hines | Bond LSC

Until the 1990s, the presence and significance of extracellular ATP, a nucleotide that normally provides energy to a cell, in animal cells was highly contested for decades. Now, the Gary Stacey lab at Bond Life Sciences Center is using that history lesson to explore ATP’s role in plant cells.

Let’s say you have $100 in your pocket. As you’re walking down the street, you throw dollar bills over your shoulder. Sounds ridiculous, right?

Much like these dollar bills, ATP is the currency of cells. When the idea of animal cells dispensing ATP outside the cell to trigger danger signals came out in the 1970s, the scientific community thought it was wasteful and therefore the idea was ridiculed.

“The idea of throwing money over your shoulder is a very good analogy because that’s a good way to influence people,” said Stacey, a principal investigator at Bond LSC. “Give them a financial incentive. It’s the same thing with ATP. The idea is that this is the currency of the cell, but they’re putting it outside the cell in order to signal and influence interactions with their environment or other cells.”

This process of ATP receptors on the cellular membrane bonding with molecules to send signals and communicate between cells is called purinergic signaling.

The term was coined by retired neurobiologist Geoffrey Burnstock in the 1970s when he discovered it in animals; however, Burnstock’s findings were met with skepticism. It wasn’t until 20 years later when the first ATP receptor in animal cells was discovered that his idea gained acceptance.

Today, animal cell purinergic signaling has been used to make drugs that target diseases and develop drug therapies. As for purinergic signaling in plant cells, the research is just beginning. Much like what Burnstock faced, this new research is met with skepticism.

“It was clear to me that if we really wanted to establish this so that people would believe it, we had to get the receptor,” Stacey said.

In 2014, the Stacey lab reported its first ATP receptor called P2K1. Earlier this year, they found another one called P2K2.

“I think more and more people are working on purinergic signaling in plants,” Stacey said. “I think we will see a big explosion of people working in this area. Just as is the case with the animal field, the plant field is really going to take off now that we actually know what the receptors are.”

Currently, the Stacey lab is trying to understand how these receptors work. Research scientist Sung-Hwan Cho from the Stacey lab took over the project regarding P2K1.

“The ATP study in animals is just half a piece of the puzzle,” Cho said. “You need to find the other half of the piece. For 40 years, many labs have already studied this, but in plants, there’s just been a few.”

Research done by the Stacey lab has shown that although not much is known about purinergic signaling in plants compared to in animals, purinergic signaling can impact plant growth, development, defense against diseases and stress tolerance. Cho is currently studying how P2K1 impacts certain metabolic pathways in plants.

In another experiment done by the lab, they found that mutants without the ATP receptor that experienced stress did not grow smaller while plants that did have the receptor and experienced stress grew smaller.

“That says the purinergic signaling is what’s mediating the stress response,” Stacey said. “That is affecting the development of the plant in response to stress… So, if we understand this better then maybe we can make plants that are more drought and stress-tolerant and grow better.”

Four months in, Baker lab hits the ground running and won’t be stopping anytime soon

October 19, 2020 By Lauren Hines in Research Tags: cancer, cell sheets, copper

Post doctorate Harim Tavares from the Baker lab works in “the hood,” which is a space used to prevent researchers and other outside factors from contaminating cells.

By Lauren Hines | Bond LSC

Saliva is often something people take for granted.

It helps break down food, maintain teeth and keep the oral cavity feeling comfortable. But head and neck cancer patients lose those benefits when chemotherapy treatments damage their salivary glands.

Olga Baker set out to research tissue regeneration for these cancer patients by collaborating with the Michael Petris and Gary Weisman labs after setting up shop four months ago as a new principal investigator at Bond Life Sciences Center.

Together, the Petris and Baker labs are looking at the role of copper in salivary gland damage.

“Nobody has asked what happens to copper metabolism during and after head and neck radiation,” Baker said. “Is copper related to the fibrosis that is developing in those patients, so they don’t make saliva? We don’t know. So, we are answering different questions…. We don’t have any data yet, but what we have is immunofluorescence studies showing that those copper transporters are present in salivary glands from mice and humans.”

If the two labs can learn how copper connects to salivary gland tissue regeneration, Baker can get a step closer to helping patients.

The Baker and Petris labs just published a paper with help from the Proteomics Center and the MU Molecular Interactions Core in the journal of Dental Research. Post doctorate Harim Tavares from the Baker lab is the leading author.

The study used synthetic networks of crosslinked polymer chains that can take in large amounts of water called, smart hydrogels. The smart hydrogels were used in the same mouse model as the cell sheets and promoted tissue regeneration on mouse salivary glands. The paper explored how different materials like smart hydrogels can improve tissue regeneration.

Baker’s other focus takes on cancer and tissue regeneration from a different angle in her collaboration with the Weisman lab.

“Dr. Weisman’s group has studied the mechanisms of salivary gland dysfunction for over 30 years, so between these two groups the tools are in place to provide a translational approach to understand and treat salivary gland disorders,” said Jean Camden, a senior research associate in the Weisman lab.

The two labs just submitted a paper on mouse submandibular glands and cell sheets, which are plastic-like sheets where cells grow and detach once it reaches a certain temperature. Kihoon Nam from the Baker lab is the corresponding author. The study transplanted submandibular gland cell sheets into a wounded mouse to see if cells could be transferred.

“We believe that is a good proof of concept for future use in radiation studies,” Baker said.

The Baker lab uses cell sheets with the Weisman lab to study the role of the P2Y2 receptor in salivary gland regeneration. P2Y2 is a plasma membrane receptor that binds the molecules outside of cells and serves as signals to trigger an action.

The Weisman lab previously showed that P2Y2 receptor activation encourages salivary gland cells to migrate together and form a unit capable of secreting saliva.

“At this time, we don’t know what the role of P2Y2R is in the cell sheets,” Camden said. “That is an aim we proposed in a recent TRIUMPH proposal.”

The TRIUMPH initiative — The Translational Research Informing Useful and Meaningful Precision Health initiative — is a pilot grant funding opportunity through the Missouri School of Medicine.

As if two collaborations weren’t enough, Baker is also working on saliva substitutes. The lab just submitted a grant to fund the project.

“If we get the money, we will be able to make a far better saliva substitute as compared to the Biotine, or Aquoral, or Oasis,” Baker said. “Those are the three leading brands. So, hopefully, we can make something better here at MU and commercialize the product.”

Regulators Classify Gene-Edited Rice Varieties with Disease Resistance as Equivalent to Conventional Varieties

October 14, 2020 By Roger Meissen in Research Tags: CRISPR, Danforth Plant Science Center, rice

Crops resist bacterial leaf blight; ruling clears path to provide smallholder farmers with a safe, affordable option for preventing destructive disease

Columbia and St. Louis, MO, October 14, 2020 – The Healthy Crops team, with support from the Bill & Melinda Gates Foundation, have used gene editing tools to develop new varieties of disease-resistant rice that regulators in the United States and Colombia have determined are equivalent to what could be accomplished with conventional breeding. Bacterial blight can reduce rice yields by up to 70 percent, with the heaviest losses typically experienced by smallholder rice growers in low and middle-income countries. This has a profound impact on farmer productivity and economic mobility. The Healthy Crops team turned to gene editing to develop disease-resistant varieties as a way to provide farmers with a safe, affordable, effective solution.

“We first set about to understand the gene the bacteria use to make the plant vulnerable to its disease,” said Bing Yang, PhD, a researcher with the University of Missouri Bond Life Sciences Center professor, Division of Plant Sciences and member, Donald Danforth Plant Science Center in St. Louis. “We then used our CRISPR technology precisely to remove the element in the gene to avoid the pathway the pathogen takes that makes the plants susceptible to blight.”

The team used gene editing to create rice lines in elite varieties that are comparable to naturally occurring variants. These lines can resist infection by bacterial leaf blight, which leads to major losses for one of the world’s most important food crops. The rulings from the United States Department of Agriculture (USDA) and the corresponding authority in Colombia, the Instituto Colombiano Agropecuario (ICA), clear the way for field tests to select the best material for distribution to breeders in the U.S. and Colombia.

The improvements were accomplished via gene editing, which did not introduce any DNA into the plants and focused on “promoter regions” in three genes that are targeted by the causative agent of rice blight, the bacterium Xanthomonas oryzae pathovar oryzae. The research was described in an article in Nature Biotechnology in 2019.

Yang is just one member of the research consortium, headed by Humboldt Professor Wolf B. Frommer from Heinrich Heine University Düsseldorf (HHU), that has worked more than four years on this research. Six research institutions on three continents were involved including the University of Missouri, Donald Danforth Plant Science Center, University of Florida, the Alliance of Bioversity International and the International Center for Tropical Agriculture (CIAT) in Colombia, the Institut de Recherche pour le Développement (IRD) in France and the International Rice Research Institute (IRRI) in the Philippines.

In the wake of the ruling from U.S. and Colombian officials, the new blight-resistant varieties can now be used to introduce the resistance trait into many different types of rice via standard breeding strategies. Additional testing and breeding work is expected to take place in multiple locations that are favorable for growing tropical rice varieties.

“It’s exciting to use science and technology to do to help farmers protect and improve their rice production,” Yang said. “We hope to work closely with the local institutions in the next phase to introduce these into the varieties of rice small farmers use.”

The Healthy Crops Team has no commercial interest in its work. Its goal is to ensure disease- resistant rice varieties are accessible and affordable, especially for smallholder farmers who depend on rice production to support their families.

About The Donald Danforth Plant Science Center
Founded in 1998, the Donald Danforth Plant Science Center is a not-for-profit research institute with a mission to improve the human condition through plant science. Research, education and outreach aim to have impact at the nexus of food security and the environment, and position the St. Louis region as a world center for plant science. The Center’s work is funded through competitive grants from many sources, including the National Institutes of Health, U.S. Department of Energy, National Science Foundation, and the Bill & Melinda Gates Foundation. Follow us on Twitter at @DanforthCenter.

About Bond Life Sciences Center

Founded in 2004, the Christopher S. Bond Life Sciences Center was designed with teamwork in mind, fostering collaborations between scientists of diverse disciplines and backgrounds. From cancer and HIV to plant science and informatics, our researchers work together to move basic science forward and lay the groundwork for a better world. Learn more at bondlsc.missouri.edu.

Finding a Link between Xenoestrogens and Autism

October 12, 2020 By Becca Wolf in Research Tags: Autism Spectrum Disorder, biochemistry, mice

California mice | photo by Roger Meissen, Bond LSC

By Becca Wolf | Bond LSC

It’s hard to see a family member treated differently because of a behavior disorder, but those with relatives diagnosed with Autism Spectrum Disorder (ASD) know its impact. Since it affects how people act, communicate, and learn, people with ASD often get bullied or feel left out because of these behaviors.

Sarabjit Kaur, a former undergraduate researcher in the Cheryl Rosenfeld lab at the Bond Life Sciences Center, witnessed this stigma firsthand growing up with a brother who is autistic. Going to college, she knew she wanted to do something that made a difference in understanding ASD.

In Kaur’s final year at MU, she conducted a correlational study to see the effect of xenoestrogens on the microbiome-gut-brain axis and autistic behavior.

“The dramatic rise in ASD suggests that there’s an environmental component to this because genetics don’t change that fast,” Kaur said, “We know that steroid hormones play an important role in brain development. That’s why we suspect that endocrine disruptors such as the xenoestrogens, BPA [bisphenol A] and genistein, might be an important contributor to this disease.”

Xenoestrogens like BPA are often called “forever chemicals” because they don’t readily break down in the environment. Since we use millions of tons of manufacturing plastics, as food can linings and in things like receipts, our exposure can add up. While genistein is naturally found in plant products such as soybeans and coffee, both are common in our environment and affect the body in some way.

To start, Kaur tested the effects of the chemicals on California mice. She found that mice exposed to genistein were more likely to exhibit repetitive behaviors and socio-communicative disturbances. Mice with a low dose of BPA resulted in more frequent repetitive behaviors compared to those with a higher dose. These social communicative deficits were dose dependent autistic-like behaviors, showing that there is a possible correlation between these xenoestrogens and ASD.

In this study, Kaur also found that there are sex dependent effects of these xenoestrogens. BPA and genistein affected female and male mice differently. Exposed female mice resulted in different types of metabolites relative to control such as amino acids or glucose involved in carbohydrate metabolism, but in males the protein metabolism was affected by BPA and genistein relative to the control group.

These processes occur in the microbiome-gut-brain axis, which is the feedback loop where microbes communicate to the central nervous system through channels involving nervous, endocrine, and immune signaling mechanisms.

Exposure to xenoestrogens may be inducing epigenetic changes through altering gene/protein expression, without changing the DNA sequence itself.

“The results get super complicated because there has been dose dependent effects, both sex dependent effects and treatment dependent effects,” Kaur said.

Scientists use California mice as a model for behavior research because of their relationship similarities to humans. They are social creatures, both parents care for their young, and the male and female mate for life. This gives insight as to how BPA and genistein may impact humans.

“We wanted to do this in an animal model because we could help inform our epidemiology colleagues,” Kaur said, “So those that do work with humans, this could help them see what they are looking for as in terms of a diagnosis tool.”

In the future, doctors could look at the bacterial population or the metabolites in the microbiome-gut-brain axis to determine if a child is autistic.

“I just hope, this research will help my brother and people like him. That’s what research is all about, to help advance. Hopefully, we could give people a fresh, healthy start in the beginning of their lives,” Kaur said.

“It would mean a lot to many if in the future people could help their friends, family members, or their siblings that go through ASD,” Kaur said, “It would really help even if we could alleviate the symptoms or give people a fresh start.”

For more information, check out the December 2020 edition of Scientific Reports article, “Developmental exposure of California mice to endocrine disrupting chemicals and potential effects on the microbiome-gut-brain axis at adulthood.”

Plant vs. Pathogen

October 7, 2020 By Lauren Hines in Research Tags: plant immune system

Research scientist Jianbin Su studies the immune system in lettuce on the third floor of the Bond Life Science Center.

By Lauren Hines | Bond LSC

Scientist Jianbin Su’s research lately took him outside to look at patches of grass and cracks in sidewalks around Mizzou’s campus, searching for a subject in the wild. He found his subject in the form of wild lettuce.

That search can potentially help Su — a research scientist in the Walter Gassmann lab at Bond Life Sciences Center — better understand the immune system of lettuce and therefore protect it against diseases like Downy Mildew and Xanthomonas, which would increase lettuce crop yield.

Su’s paper under review about how the pathogen effector protein, AvrRps4, interacts with lettuce’s immune system will hopefully soon be accepted in the journal Molecular Plant-Microbe Interactions.

“Scientists in Florida are still focusing on lettuce resistance to Xanthomonas,” Su said. “If we can find a mechanism where lettuce recognizes our protein, this finding can be applied to make lettuce immune to Xanthomonas.”

Xanthomonas causes bacterial leaf spots and is a serious seed-borne disease that results in significant losses for the lettuce industry

To infect a plant successfully, the bacterium must inject 30 to 40 protein effectors, like AvrRps4, in the cell and interact with multiple plant cell functions at once to overcome the host. Effectors for a pathogen can bind to proteins that affect the biological activities of the plant, letting the disease invade the plant, suppress its immune system and allow Xanthomonas to survive.

However, when the plant cell’s receptors recognize the pathogen, it can trigger its immune response. To stop the pathogen from growing, the plant cell initiates a hypersensitive response by killing itself. If the plant can’t stop the pathogen, it will multiply and affect much larger areas of the leaf. This will ultimately destroy the plant tissue and cause the characteristic round, brown dead spots on the lettuce leaves.

“Plants can afford the hypersensitive response because their developmental program isn’t set in stone like in most animals,” said Walter Gassmann, interim director for Bond Life Sciences Center. “They can adjust their growth in real-time…so losing a leaf is not a big deal to a plant, not like us, losing a limb or something like that.”

Wild Lettuce Found on Campus — While studying lettuce immunity, research scientist Jianbin Su took photos of wild lettuce growing on campus. | photo contributed by Jianbin Su.

Once AvRrps4 enters the plant cell, it splits up into two parts: AvRrps4-C and AvRrps4-N. Some plants like flowers, tomatoes and melons only recognize the C part, but lettuce only recognizes the N part of the effector.

“The C part of AvrRps4 has been studied by many labs, and they’ve already found some mechanisms,” Su said. “The C part attacks a group of important proteins that are required for plant immunity. That’s the function of the C part, but the function of the N part is not clear. That’s what I’m focusing on in lettuce.”

Recognition is important because, without it, plant cells can’t respond to fight off the pathogen.

Su is studying lettuce not only because it hasn’t been studied as much, but so far, it’s the only plant that requires recognition of both parts to initiate immunity.

This means lettuce evolved to recognize and attack AvRrps4-C. In turn, the effector evolved by adding a second part to prevent recognition by plants.

This evolutionary choice, if you will, is odd. Instead of getting around the immune response by getting rid of the protein that is recognized, the protein added a second part to itself. This means AvRrps4-N, the original part, must possess a vital function for the pathogen worth keeping.

“That’s why we think this effector arose in nature,” Gassmann said. “It’s because the first part triggered a very strong immune response, something else got tacked on to it to weaken that immune response to give the pathogen a leg up.”

This evolutionary battle can be seen in Mizzou’s very own backyard. Su found five different types of wild lettuce. After testing if the types were able to recognize the N part of the effector, he found two of the five did.

Since some did and others did not, Su thinks the receptor for recognizing the effector has been lost in some wild lettuce types due to evolution and the absence of the pathogen in certain areas.

“We haven’t found the lettuce receptor but based on this information we can narrow it down,” Su said. “Right now, we have 36 candidates. We are already sure that among the 36, there’s 99.999% that there is one receptor that recognizes our protein.”

Su still has much work left to do but, for now, he resubmitted his paper Oct. 5.

“Once we identify the receptors that recognize our protein, our research will go into the next direction, and maybe have some application purpose for our research,” Su said.

«‹ 8 9 10 11 ›»