reactive oxygen species

#IAmScience Kulbir Sandhu

May 14, 2021 By Lauren Hines in #IAmScience Tags: CRISPR, reactive oxygen species

By Lauren Hines | Bond LSC

Kulbir Sandhu’s curiosity had guided him from place to place, but it was his fascination with plant science that has stayed the same.

While Sandhu has been a postdoctoral fellow in the Bing Yang lab at Bond Life Sciences Center for the past six months, his path towards plant science began when he was 18 years old in his home country of India.

In high school, Sandhu was drawn to the biology route because of helpful and enthusiastic science teachers. He grew to like it as time went on and found that the subject came easy to him.

“I always had, you could say, a ‘scientific’ attitude,” Sandhu said. “Even when I had little understanding of the process of science, I always had an attitude that suited this field.”

Sandhu also received support and inspiration regarding science from his father, who was an engineer.

“When I was young, my dad used to help me with school homework,” Sandhu said. “His favorite subject was maths, and so he always insisted that I do well in maths and science. In this way, it became natural for me to develop an inclination for these two subjects. In India, most students interested in science choose either maths or biology streams after 10th grade. Initially, I wanted to be a doctor, so I chose biology, and it was more of a happenstance that I ended up becoming a plant ‘doctor.’”

Years later, Sandhu received his Master’s in plant breeding from the Punjab Agricultural University in Ludhiana, India in 2003, and in 2013 he received his Ph.D. at Washington University.

Sandhu first met Bing Yang four years later as a postdoctoral fellow at Iowa State University. Since they worked on a previous project together, Sandhu became a great addition to the Yang lab when he joined in November last year.

While Sandhu is working on a few projects, his main one involves using the gene-editing tool, CRISPR/CAS9, to target genes in Arabidopsis that code for reactive oxygen species. ROS helps plants with signaling, development, stress responses and other processes.

ROS is also part of the plant’s innate defense system against pathogens. Understanding how pathogens overcome this primary defense system of plants is necessary to breed better resistant crops and reduce environmental impact due to chemical control.

By causing these gene mutations, he prevents ROS from being formed in cells. That way, they can compare the mutant plant to a wild type and see the difference in basal-defense responses.

“Now this is exciting again because we are working in CRISPR in field crops as well as in basic science,” Sandhu said. “So, I get to do both things.”

First-year researcher Jack Ogilvy has been working on this project with Sandhu for the past three months as part of the Freshmen Research in Plants program.

“This is my first time … mentoring someone, and by this experience, I have realized that it is equally beneficial to me,” Sandhu said. “I mean … talking about scientific concepts helps create a deeper understanding, and both parties gain from this interaction.”

Together, the two are learning more about ROS and the Yang lab.

“He cares more about just being a mentor in terms of science,” Ogilvy said. “He also is just as interested in my personal life … We’ve formed a relationship between the two of us where it’s not just like, he tells me what to do in the lab. It’s like we are working together, essentially.”

Ogilvy appreciates Sandhu’s curiosity and advice.

“He’s always telling me to try to find the answer on my own before I go for help to gain that skill … just because it’s such an important skill to have to be somewhat self-reliant,” Ogilvy said. “But that being said, he’s always there if I get stuck or if I need help.”

Sandhu found a place for himself in the Yang lab. In a few weeks, he plans on focusing more on his own projects.

Bond LSC Researchers Named Most Highly Cited for 2020

January 26, 2021 By Lauren Hines in Research Tags: Highly Cited Researchers, plant immune system, reactive oxygen species

Bing Yang and Ron Mittler — Bond Life Sciences Center principal investigators Bing Yang (left) and Ron Mittler (right) are pictured above. | photos by Erica Overfelt, Bond LSC.

By Lauren Hines | Bond LSC

Building onto previous knowledge is a pillar of the scientific process, and citations in research do just that.

This makes recognition of Bond Life Sciences Center principal investigators Ron Mittler and Bing Yang, as well as Mizzou biochemistry professor Shuqun Zhang in the Highly Cited Researchers list for 2020, an important acknowledgment.

“I’m glad to have this, and this is the second year [I have been on the list],” Yang said. “If you have your paper widely read by other researchers and actually cited, it’s relevant to the rest of the field, or to the community. We always talk about the impact of your research to the community and the impact of the research on science.”

Yang studies the resilience and susceptibility of disease in crops, specifically rice. The Bing Yang lab also strives to develop genome editing technology to discover and improve traits in rice, like disease resistance.

“It’s significant because we create some useful tools and develop technology, so then other people or the community can use it,” Yang said. “We also work on the bacteria and disease, which can help the small farmer, help the grower of the rice to fight the disease and make the rice more productive.”

On the other hand, Mittler’s research focuses more on how reactive oxygen species (ROS) helps regulate systemic signaling in plants. ROS are reactive molecules that perform chemical reactions easily. While these molecules destroy cells with these reactions, they can also send widespread messages and regulate many essential processes.

Understanding ROS – and building upon the knowledge of ROS – can help researchers genetically modify crops to withstand more environmental stresses in the future.

With some luck and most of 2021 ahead, Mittler, Yang, Zhang and others will continue their pursuit of discovery and expand the shared knowledge base of science for the future.

To learn more, read some of Bing Yang and Ron Mittler’s most cited work. The Highly Cited Researchers List is compiled by Clarivate’s Web of Science.

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.

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.

A NEET Connection

March 2, 2020 By Becca Wolf in Research Tags: Arabidopsis, cancer, reactive oxygen species

Protein is important in balancing iron and reactive oxygen in plant and cancer cells

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

By Becca Wolf | Bond LSC

You might tend to think durability is more of an issue in building a car or engineering a building, but environmental stress makes resilience vital for plants, too.

In the world of understanding and engineering more durable crops, scientists recently identified a protein that’s key to some of that resilience in a neat way. Actually, it’s literally a NEET protein.

Ron Mittler, a Bond Life Sciences Center principal investigator and professor of plant sciences and surgery, is probing the use of the NEET protein to balance iron and reactive oxygen species (ROS) in the chloroplast to make plants stronger and healthier, and it plays a similar role in cancer cells. These proteins belong to a unique family of iron-sulfur (Fe-S) proteins ideally maintaining the proper balance of the heavy metal in and outside of plant cells. Along with several other scientists, Mittler published his most recent plant findings in The Plant Journal last October.

Since NEET proteins regulate both iron and ROS, Mittler teamed up with heavy metal expert David Mendoza, fellow Bond LSC primary investigator and associate professor of plant sciences. Through a Bond LSC Grant for Innovative Collaborative Research, the two started work at the intersection of two labs.

“The main philosophy of Bond LSC is to establish collaborations between researchers to advance the field of whatever sciences that you do,” Mendoza said.

Mittler and Mendoza looked at how NEET impacts iron and ROS, working together to figure out how to make plants more durable. Doing so, they have made a lot of progress in the field and believe that their collaboration has been a good one. Says Mendoza, “It is a very productive collaboration.”

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

A Balancing Act

But the work doesn’t stop with plants. Mittler found NEET proteins also play a similar role in cancer cells, regulating the level of iron and potentially toxic ROS compounds. Humans and plants need iron and ROS, but too much of either can be destructive. NEET proteins allow cells to grow and spread quickly which is beneficial for plants, but detrimental for humans when cancers grow out of control. In humans, NEET proteins also play a significant role metabolism related to diabetes and progression of neurodegenerative diseases.

Despite needing both iron and ROS to function, there can be too much of a good thing. To keep iron levels balanced, oxygen is needed. While oxygen on its own is a fairly stable molecule, adding electrons or exciting the present electrons can make the molecule want to react with things around it creating oxidants. Electrons are the negatively charged particles of an atom that keep balance within the cell. When balanced, iron and ROS helps cells grow, but when placed together, the iron turns ROS into hydroxyl radical, an ion that reacts with anything and destroys everything. If not controlled, this desire to bond with things nearby can damage cells and lead to cancer.

Cancer cells like ROS and iron because it enables them to spread quickly and take over more of the body. To stay balanced, NEET proteins are needed.

“These proteins apparently enable the cells to keep high levels of both iron and ROS and to survive,” says Mittler, “So it looks like cancer needs these proteins because if you suppress these proteins or if you target them with drugs, the cancer cells die.”

Sara Zandalinas holding a dying Arabidopsis seedling and a living seedling in Ron Mittler’s lab. | photo by Becca Wolf, Bond LSC

Poisoning the cell

Knowing NEET proteins make cancer and plant cells stronger, Mittler wanted to look at “what happens if they’re not functioning well,” in hopes to understand their function. To test this, Mittler and his collaborators found a way to make the NEET proteins not function. Mittler called this “poisoning the cell.”

“If there is no protein, there’s no life,” he states. “We were able to control when we introduce this protein, and being able to introduce this protein enabled us to actually find out what happened to the cell if this protein goes bad.”

It was found that if the function of this protein is disrupted, the chloroplast is destroyed. Mittler described it as “completely blown to pieces.” The destruction of the chloroplast is caused by too much iron due to a broken NEET protein. Similarly in cancer cells, if there are no NEET proteins, the cells do not spread as quickly and die.

Mittler elaborated, “What we found is that we can target those proteins in cancer, so then we basically destroy the cancer cells.” Currently, NEET proteins are used as markers in the cancer field. Doctors and scientists look for them in order to help diagnose patients and to figure out life expectancy rates. But in the future, doctors hope to use targets these proteins to kill off cancer cells, a treatment that would be less toxic than chemotherapy.

In plants, Mittler has concluded that the NEET protein would make them grow bigger and faster, leading plants to be “more tolerant to stresses because it has the ability to modulate the levels of iron and reactive oxygen.” Findings like these may one day help plants deal with stress like drought and temperature extremes. With humans continuing to contaminate the environment and climate change escalating conditions, Mittler hopes his findings on iron, ROS, and NEET proteins will make plants able to withstand changes better.

“If I can improve the way plants balance reactive oxygen and iron, I can make plants tolerate much more of these bad combinations of stresses even, and maybe even make bigger, stronger, healthier plants.”

Read more about this research in the October 2019 edition of The Plant Journal article “Expression of a dominant‐negative AtNEET‐H89C protein disrupts iron–sulfur metabolism and iron homeostasis in Arabidopsis.”

The Bond LSC Grant for Innovative Collaborative Research program aims to foster inter-laboratory collaboration to make pilot projects that have potential for later federal funding.

The search for oxidative stress treatment continues

October 1, 2014 By Paige Blankenbuehler in Research Tags: Bond Life Sciences Center, drug, jeff sossamon, mark hannink, oxidative stress, reactive oxygen species, research, university of missouri science research

A yellow light indicates oxidant production in the tissue of a migrating fly larva. Source: Tobias Dick, German Cancer Research Center | Illustration by Paige Blankenbuehler Read more at: http://phys.org/news/2011-12-oxidative-stress.html#jCp — A yellow light indicates oxidant production in the tissue of a migrating fly larva. Source: Tobias Dick, German Cancer Research Center | Illustration by Paige Blankenbuehler

University of Missouri research characterizes a novel compound

By Paige Blankenbuehler | Bond LSC

Your body has an invisible enemy.

One that it creates all on it’s own called oxidative stress, long thought of as an underlying cause of some of humanity’s most insidious diseases – cancer, Alzheimer’s, Parkinson’s Disease, cardiovascular disease and diabetes.

Every day, our bodies are exposed to harmful free radicals known as reactive oxygen species as a result of our environment.

But, when something goes wrong with this energy extraction process, cells become inundated with reactive oxygen compounds that cause oxidative stress. The search for drugs to treat the problem have been ongoing, and with a complicated problem like oxidative stress, it’s all about finding the right combination.

Recent research by Bond Life Sciences Center investigator and Biochemistry Department Professor, Mark Hannink, provides a new approach for addressing the problem of oxidative stress and a starting point on developing a drug in pill form.

Mark Hannink and Kimberly Jasmer, a Ph.D. student in his lab, recently helped characterize a new molecule (called HPP-4382) that provides a novel way to treat oxidative stress. Their research was done in a partnership with High Point Pharmaceuticals, LLC, of North Carolina, where this new molecule was developed.

Oxidative stress can cause damage to the building blocks of a cell, resulting in excessive cell proliferation in the case of cancer or cell death in the case of neurodegenerative diseases like Parkinson’s.

Often, the majority of stressors are actually created inside our own cells as a byproduct of how our cells extract energy from the food that we eat and the air that we breathe.

Specimen of protein are prepared for an experiment in a lab at the Bond Life Sciences Center at the University of Missouri.

Understanding oxidative stress

Most simply, oxidative stress is an imbalance that happens when the body uses oxygen to produce energy.

Superoxide, a “promiscuous and nonspecific” compound produced as a byproduct of this process, is a highly reactive molecule that can damage DNA and other cellular components, Hannink said.

The superoxide molecule is a “free radical.” That means it’s especially promiscuous and reacts with many different types of cellular molecules, leaving destruction in its wake, he said.

That damage can lead to a long list of problems, including cancer or neurodegenerative diseases like Parkinson’s disease.

In response to oxidative stress, the cell produces protective “anti-oxidant” proteins, which help remove the harmful reactive oxygen species and minimize damage.

But a heavy anti-oxidant response could be dangerous, too. The bottom line: it’s about maintaining a fine balance between “oxidants” and “anti-oxidants”.

Search for right combination continues

A drug that corrects the imbalance of oxidative stress could one day have wide applicability.

Jasmer developed a test to measure how specific compounds altered gene expression. The genetic response to oxidative stress has both an “ON” switch and an “OFF” switch.

Using this test, Jasmer determined how each compound affected specific genetic switches and, in turn, how the response to oxidative stress is regulated.

This test helped identify which molecules might be promising candidates for treating oxidative stress, leading them to one in particular that seemed to have the desired properties: HPP-4382.

But creating effective drugs is a long process of trial and error. Once molecules have been identified that show efficacy in lab-based assays, scientists try different combinations to increase their potency and drug-like properties, and High Point is currently testing other molecules that behave like HPP-4382.

The compound serves as a good starting point for researchers who are interested in understanding how oxidative stress affects cellular processes, such as cell proliferation or cell death.“Now we have a better understanding of what this compound is doing,” Hannink said. “This compound can be used to test different ideas of how the balance between oxidants and anti-oxidants is achieved in healthy cells and how perturbation of this balance can lead to different diseases.”

This research was published in PLoS One in July and was funded by High Point Pharmaceuticals, LLC.