Duke Research Blog

Following the people and events that make up the research community at Duke.

Category: Genetics/Genomics (Page 1 of 7)

Sean Carroll on the Evolution of Snake Venom

What’s in a snake bite?

According to University of Wisconsin-Madison evolutionary biologist Sean Carroll who visited Duke and Durham last week, a snake bite contains a full index of clues.

In his recent research, Carroll has been studying the adaptations of novelties in animal form, such as snake venom. Rattlesnakes, he explains, are the picture of novelty. With traits such as a limbless body, fangs, infrared pits, patterned skin, venom, and the iconic rattle, they represent an amazing incarnation of evolution at work.

Rattlesnakes: the picture of novelty (Photo from USGS)

Snake venoms contain a complex mixture of proteins. This mixture can differ in several ways, but the most interesting difference to Carroll is the presence or absence of neurotoxins. Neurotoxic venom has proven to be a very useful trait, because neurotoxins destroy the nervous tissue of prey, effectively paralyzing the animal’s respiratory system.

Some of today’s rattlesnake species have neurotoxic venom, but some don’t. So how did this happen? That’s what Carroll was wondering too.

Some genes within genomes, such as HOX genes, evolve very slowly from their original position among the chromosomes, and see very few changes in the sequence in millions of years.

But snake venom Pla2 genes are quite the opposite. In recent history, there has been a massive expansion of these genes in the snake genome, Carroll said. When animals evolve new functions or forms, the question always arises: are these changes the result of brand new genes or old genes taking on new functions?

Another important consideration is the concept of regulatory versus structural genes. Regulatory genes control the activity of other genes, such as structural genes, and because of this, duplicates of regulatory genes are generally not going to be a favorable adaptation. In contrast, structural gene activity doesn’t affect other genes, and duplicates are often a positive change. This means it is easier for a new structural gene to evolve than a regulatory one. Carroll explained.

Evolutionary Biologist Sean Carroll (Photo from seanbcarroll.com)

Carroll examined neurotoxic and non-neurotoxic snakes living in overlapping environments. His research showed that the most recent common ancestor of these species was a snake with neurotoxic venom. When comparing the genetic code of neurotoxic snakes to non-neurotoxic ones, he found that the two differed by the presence or absence of 16 genes in the metalloproteinase gene complex. He said this meant that non-neurotoxic venom could not evolve from neurotoxic venom.

So what is the mechanism behind this change? What could be the evolutionary explanation?

When Carroll’s lab compared another pair of neurotoxic and non-neurotoxic species in a different region of the US, they found that the two species differed in exactly the same way, with the same set of genes deleted as had been observed in the first discovery. With this new information, Carroll realized that the differences must have occurred through the mechanism of hybridization, or the interbreeding of neurotoxic and non-neurotoxic species.

Carroll’s lab is now doing the structural work to study if the genes that result in neurotoxic and  non-neurotoxic protein complexes are old genes carrying out new functions or entirely new genes. They are using venom gland organoids to look into the regulatory processes of these genes.

In addition to his research studying the evolution of novelties, Carroll teaches molecular biology and genetics at Madison and has devoted a large portion of his career to  storytelling and science education.

Drug Homing Method Helps Rethink Parkinson’s

The brain is the body’s most complex organ, and consequently the least understood. In fact, researchers like Michael Tadross, MD, PhD, wonder if the current research methods employed by neuroscientists are telling us as much as we think.

Michael Tadross is using novel approaches to tease out the causes of neuropsychiatric diseases at a cellular level.

Current methods such as gene editing and pharmacology can reveal how certain genes and drugs affect the cells in a given area of the brain, but they’re limited in that they don’t account for differences among different cell types. With his research, Tadross has tried to target specific cell types to better understand mechanisms that cause neuropsychiatric disorders.

To do this, Tadross developed a method to ensure a drug injected into a region of the brain will only affect specific cell types. Tadross genetically engineered the cell type of interest so that a special receptor protein, called HaloTag, is expressed at the cell membrane. Additionally, the drug of interest is altered so that it is tethered to the molecule that binds with the HaloTag receptor. By connecting the drug to the Halo-Tag ligand, and engineering only the cell type of interest to express the specific Halo-Tag receptor, Tadross effectively limited the cells affected by the drug to just one type. He calls this method “Drugs Acutely Restricted by Tethering,” or DART.

Tadross has been using the DART method to better understand the mechanisms underlying Parkinson’s disease. Parkinson’s is a neurological disease that affects a region of the brain called the striatum, causing tremors, slow movement, and rigid muscles, among other motor deficits.

Only cells expressing the HaloTag receptor can bind to the AMPA-repressing drug, ensuring virtually perfect cell-type specificity.

Patients with Parkinson’s show decreased levels of the neurotransmitter dopamine in the striatum. Consequently, treatments that involve restoring dopamine levels improve symptoms. For these reasons, Parkinson’s has long been regarded as a disease caused by a deficit in dopamine.

With his technique, Tadross is challenging this assumption. In addition to death of dopaminergic neurons, Parkinson’s is associated with an increase of the strength of synapses, or connections, between neurons that express AMPA receptors, which are the most common excitatory receptors in the brain.

In order to simulate the effects of Parkinson’s, Tadross and his team induced the death of dopaminergic neurons in the striatum of mice. As expected, the mice displayed significant motor impairments consistent with Parkinson’s. However, in addition to inducing the death of these neurons, Tadross engineered the AMPA-expressing cells to produce the Halo-Tag protein.

Tadross then treated the mice striatum with a common AMPA receptor blocker tethered to the Halo-Tag ligand. Amazingly, blocking the activity of these AMPA-expressing neurons, even in the absence of the dopaminergic neurons, reversed the effects of Parkinson’s so that the previously affected mice moved normally.

Tadross’s findings with the Parkinson’s mice exemplifies how little we know about cause and effect in the brain. The key to designing effective treatments for neuropsychiatric diseases, and possibly other diseases outside the nervous system, may be in teasing out the relationship of specific types of cells to symptoms and targeting the disease that way.

The ingenious work of researchers like Tadross will undoubtedly help bring us closer to understanding how the brain truly works.

Post by undergraduate blogger Sarah Haurin

Post by undergraduate blogger Sarah Haurin

 

DNA Breakage: What Doesn’t Kill You…

What doesn’t kill you makes you stronger―at least according to Kelly Clarkson’s recovery song for middle school crushes, philosopher Friedrich Nietzsche, and New York University researcher Viji Subramanian.

During the creation of sperm or eggs, DNA molecules exchange genetic material. This increases the differences between offspring and their parents and the overall species diversity and is thought to make an individual and a species stronger.

However, to trade genetic information — through a process called recombination — the DNA molecules must break at points along the chromosomes, risking permanent damage and loss of genomic integrity. In humans, errors during recombination can lead to infertility, fetal loss, and birth defects.

Subramanian, a postdoctoral researcher in the lab of Andreas Hochwagen at NYU, spoke at Duke on February 26. She studies how cells prevent excessive DNA breakage and how they regulate repair.

Subramanian uses budding yeast to study the ‘synaptonemal complex,’ a structure that forms between pairing chromosomes as shown in the above image. Over three hundred DNA breakage hotspots exist in the budding yeast’s synaptonemal complex. Normally, double-stranded DNA breaks go from none to some and then return to none.

However, when Subramanian removed the synaptonemal complex, the breaks still appeared, but they did not completely disappear by the end of the process. She  concluded that synaptonemal complex shuts down DNA break formation. The synaptonemal complex therefore is one way cells prevent excessive DNA breakage.

The formation of the synaptonemal complex

 

During DNA breakage repair, preference must occur between the pairing chromosomes in order for recombination to correctly transpire. A protein called Mek1 promotes this bias by suppressing DNA in select areas. Early in the process of DNA breakage and repair Mek1 levels are high, while synaptonemal complex density is low. Later, the synaptonemal complex increases while the Mek1 decreases.

This led to Subramanian’s conclusion that synaptonemal complex is responsible for removing Mek1, allowing in DNA repair. She then explored if the protein pch2 regulates the removal of Mek1. In pch2-mutant budding yeast cells, DNA breaks were not repaired.

Subramanian showed that at least one aspect of DNA breakage and repair occurs through the Mek1 protein suppression of repair, creating selectivity between chromosomes. The synaptonemal complex then uses pch2 to remove Mek1 allowing DNA breakage repair.

Subramanian had another question about this process though: how is breakage ensured in small chromosomes? Because there are fewer possible breaking points, the chance of recombination seems lower in small chromosomes. However, Subramanian discovered that zones of high DNA break potential exist near the chromosome ends, allowing numerous breaks to form even in smaller chromosomes. This explains why smaller chromosomes actually exhibit a higher density of DNA breaks and recombination since their end zones occupy a larger percentage of their total surface area.

In the future, Subramanian wants to continue studying the specific mechanics behind DNA breaks and repair, including how the chromosomes reorganize during and after this process. She is also curious about how Mek1 suppresses repair and has more than 200 Mek1 mutants in her current study.

Kelly Clarkson may prove that heartbreaks don’t destroy you, but Viji Subramanian proves that DNA breaks create a stronger, more unique genetic code.         

Post by Lydia Goff

        

Obesity: Do Your Cells Have a Sweet Tooth?

Obesity is a global public health crisis that has doubled since 1980. That is why Damaris N. Lorenzo, a professor of  Cell Biology and Physiology at UNC-Chapel Hill, has devoted her research to this topic.

Specifically, she examines the role of ankyrin-B variants in metabolism. Ankyrins play a role in the movement of substances such as ions into and out of the cell. One of the ways that ankyrins affect this movement is through the glucose transporter protein GLUT4 which is present in the heart, skeletal muscles, and insulin-responsive tissues. GLUT4 plays a large role in glucose levels throughout the entire body.

Through her research, Lorenzo discovered that with modern life spans and high calorie diets, ankyrin-B variants can be a risk factor for metabolic disease. She presented her work for the Duke Developmental & Stem Cell Biology department on March 7th.

Prevalence of Self-Reported Obesity Among U.S. Adults by State, 2016

GLUT4 helps remove glucose from the body’s circulation by moving it into cells. The more GLUT4, the more sugar cells absorb.

Ankyrin-B’s role in regulating GLUT4 therefore proves really important for overall health. Through experiments on mice, Lorenzo discovered that mice manipulated to have ankyrin-B mutations also had high levels of cell surface GLUT4. This led to increased uptake of glucose into cells. Ankyrin-B therefore regulates how quickly glucose enters adipocytes, cells that store fat. These ankyrin-B deficient mice end up with adipocytes that have larger lipid droplets, which are fatty acids.

Lorenzo was able to conclude that ankyrin-B deficiency leads to age-dependent obesity in mutant mice. Age-dependent because young ankyrin-B mutant mice with high fat diets are actually more likely to be affected by this change.

Obese mouse versus a regular mouse

Ankyrin-B has only recently been recognized as part of GLUT4 movement into the cell. As cell sizes grow through increased glucose uptake, not only does the risk of obesity rise but also inflammation is triggered and metabolism becomes impaired, leading to overall poor health.

With obesity becoming a greater problem due to increased calorie consumption, poor dietary habits, physical inactivity, environmental and life stressors, medical conditions, and drug treatments, understanding factors inside of the body can help. Lorenzo seeks to discover how ankyrin-B protein might play a role in the amount of sugar our cells internalize.

Post by Lydia Goff

How Earth’s Earliest Lifeforms Protected Their Genes

A colorful hot spring in Yellowstone National Park

Heat-loving thermophile bacteria may have been some of the earliest lifeforms on Earth. Researchers are studying their great great great grandchildren, like those living in Yellowstone’s Grand Prismatic Spring, to understand how these early bacteria repaired their DNA.

Think your life is hard? Imagine being a tiny bacterium trying to get a foothold on a young and desolate Earth. The earliest lifeforms on our planet endured searing heat, ultraviolet radiation and an atmosphere devoid of oxygen.

Benjamin Rousseau, a research technician in David Beratan’s lab at Duke, studies one of the molecular machines that helped these bacteria survive their harsh environment. This molecule, called photolyase, fixes DNA damaged by ultraviolet (UV) radiation — the same wavelengths of sunlight that give us sunburn and put us at greater risk of skin cancer.

“Anything under the sun — in both meanings of the phrase — has to have ways to repair itself, and photolyase proteins are one of them,” Rousseau said. “They are one of the most ancient repair proteins.”

Though these proteins have been around for billions of years, scientists are still not quite sure exactly how they work. In a new study, Rousseau and coworkers, working with Professor David Beratan and Assistant Research Professor Agostino Migliore, used computer simulations to study photolyase in thermophiles, the great great great great grandchildren of Earth’s original bacterial pioneers.

The study appeared in the Feb. 28 issue of the Journal of the American Chemical Society.

DNA is built of chains of bases — A, C, G and T — whose order encodes our genetic information. UV light can trigger two adjacent bases to react and latch onto one other, rendering these genetic instructions unreadable.

Photolyase uses a molecular antenna to capture light from the sun and convert it into an electron. It then hands the electron over to the DNA strand, sparking a reaction that splits the two bases apart and restores the genetic information.

A ribbon diagram of a photolyase protein

Photolyase proteins use a molecular antenna (green, blue and red structure on the right) to harvest light and convert it into an electron. The adenine-containing structure in the middle hands the electron to the DNA strand, splitting apart DNA bases. Credit: Benjamin Rousseau, courtesy of the Journal of the American Chemical Society.

Rousseau studied the role of a molecule called adenine in shuttling the electron  from the molecular antenna to the DNA strand. He looked at photolyase in both the heat-loving ancestors of ancient bacteria, called thermophiles, and more modern bacteria like E. Coli that thrive at moderate temperatures, called mesophiles.

He found that in thermophiles, adenine played a role in transferring the electron to the DNA. But in E. coli, the adenine was in a different position, providing mainly structural support.

The results “strongly suggest that mesophiles and thermophiles fundamentally differ in their use of adenine for this electron transfer repair mechanism,” Rousseau said.

He also found that when he cooled E. Coli down to 20 degrees Celsius — about 68 degrees Fahrenheit — the adenine shifted back in place, resuming its transport function.

“It’s like a temperature-controlled switch,” Rousseau said.

Though humans no longer use photolyase for DNA repair, the protein persists in life as diverse as bacteria, fungi and plants — and is even being studied as an ingredient in sunscreens to help repair UV-damaged skin.

Understanding exactly how photolyase works may also help researchers design proteins with a variety of new functions, Rousseau said.

“Photolyase does all of the work on its own — it harvests the light, it transfers the electron over a huge distance to the other site, and then it cleaves the DNA bases,” Rousseau said. “Proteins with that kind of plethora of functions tend to be an attractive target for protein engineering.”

Post by Kara Manke

MyD88: Villain of Allergies and Asthma

Even if you don’t have allergies yourself, I guarantee you can list at least three people you know who have allergies. Asthma, a respiratory disorder commonly associated with allergies, afflicts over 300 million individuals worldwide.

Seddon Y. Thomas, PhD of the NIEHS

Seddon Y. Thomas, PhD of the NIEHS

Seddon Y. Thomas who works at the National Institute of Environmental Health Sciences has been exploring how sensitization to allergens occurs. The work, which she described at a recent  session of the Immunology Seminar Series, specifically focuses on the relationship between sensitization and the adaptor molecule MyD88.

MyD88 transfers signals between some of the proteins and receptors that are involved in immune responses to foreign invaders. Since allergies entail inflammation caused by an immune response, Thomas recognized that MyD88 played a role in the immune system’s sensitization to inhaled allergens.

Her research aims to discover how MyD88 alters conventional dendritic cells (cDCs) which are innate immune cells that drive allergic inflammation. MyD88 signaling in cDCs sometimes preserves open chromatin — the availability of DNA for rapid replication — which allows gene changes to happen quickly and in turn causes allergic sensitization. Open chromatin regions permit the DNA manipulation that can lead to allergies and asthma. 

Florescence microscopy image of mouse dendritic cells with mRNA-loaded blood cells.

To conduct her experiments, Thomas examines what happens in mice when she deletes MyD88 from lung epithelial cells and from antigen-presenting cells. Lung epithelial cells form a protective tissue where inhaled air meets the lung and protects from foreign invaders. But sometimes it takes its job a little too seriously and reacts strongly to allergens.

Similarly, antigen-presenting cells are involved in the immune system’s mission to protect the body, but can become confused about who the enemy is. When the signaling adaptor MyD88 is removed from lung epithelial cells, the number of eosinophils, inflammatory white blood cells, decreases. When it is removed from antigen-presenting cells, another type of white blood cell, neutrophils, also decreases.

Thomas said this shows that MyD88 is necessary for the inflammation in the lungs that causes asthma and allergies.

In her future research, Thomas wishes to explore dendritic cell gene expression, the molecular pathways controlling gene expression, and how specific types of lung epithelial cells adjust immune responses. Because MyD88 plays a role in the genetic changes, it makes sense to continue research on the genetic side.    

Post by Lydia Goff            

Rare Cancers and Precision Medicine in Southeast Asia

Data collected through genomics research is revolutionizing the way we treat cancer. But a large population of cancer patients are being denied the benefits of this research.

Patrick Tan MD, PhD is a professor of cancer and stem cell biology at Duke-NUS Medical School in Singapore.

In 2016, less than one percent of all the existing genomic data came from the 60% of the world population living outside of the US, Europe, and Japan. Furthermore, 70% of patients who die from cancer this year will come from Asia, Africa and Central and South America.

Patrick Tan, M.D., Ph.D., and the Duke-National University of Singapore (Duke-NUS) Medical School are key players in an effort to rectify this discrepancy, specifically as it exists in Southeast Asia.

In his talk, sponsored by the Duke Center for Applied Genomics and Precision Medicine, Tan focused specifically on his work in northeast Thailand with cholangiocarcinoma (CCA), or bile duct cancer.

Liver fluke

Liver flukes like this are parasites of fish that migrate to human hosts who eat the fish raw, leading to a form of bile duct cancer.

While CCA is rare in most of the world, it appears at 100 times the global rate in the region of Thailand where Tan and his colleagues work. Additionally, CCA in this region is of a separate and distinct nature.

CCA in this region is linked with a parasitic infection of the bile ducts called a liver fluke.  Residents of this area in Thailand have a diet consisting largely of raw fish, which can be infected by the liver fluke and transmitted to the person who eats the fish.

Because of the poverty in this area, encouraging people to avoid eating raw fish has proven ineffective. Furthermore, healthcare is not readily available, so by the time most patients are diagnosed, the disease has progressed into its later and deadly stage.

The life cycle of liver flukes. (Graphic U.S. Centers for Disease Control)

Tan’s genomic research has discovered certain factors at the gene level that make liver-fluke positive CCA different from other CCA. Thus genomic data specific to this population is vital to improve the outcomes of patients with CCA.

Duke-NUS Precision Medicine (PRISM) has partnered up with the National Heart Research Institute Singapore (NHRIS) in SPECTRA, a program designed to create a database of genomic data from the healthy Asian population. SPECTRA is sequencing the genomes of 5,000 healthy Asians in order to create a baseline to which they can compare the genomes of unhealthy individuals.

These and other programs are part of a larger effort to make precision medicine, or healthcare tailored to an individual based on factors like family history and genomic markers, accessible throughout southeast Asia.

By Sarah Haurin

 

Captive Lemurs Get a Genetic Health Checkup

DURHAM, N.C. — Careful matchmaking can restore genetic diversity for endangered lemurs in captivity, researchers report.

Ring-tailed lemurs born at the Duke Lemur Center have seen a recent infusion of new genetic material at key genes involved in the immune response, finds a new study.

Thanks to a long-term collaborative breeding program that transfers animals between institutions to preserve genetic diversity, genetic variation at one region was restored to levels seen in the wild.

The findings, published in the journal Ecology and Evolution, are important for the ability of future generations to fight disease.

Baby lemur twins Nemesis and Narcissa were the product of a breeding program developed by the American Association of Zoos and Aquariums to preserve the future genetic health of North America’s captive ring-tailed lemurs. Their mother Sophia was among 62 ring-tailed lemurs recommended for breeding across 20 institutions nationwide in 2016. Photo by David Haring, Duke Lemur Center.

Baby lemur twins Nemesis and Narcissa were the product of a breeding program developed by the American Association of Zoos and Aquariums to preserve the future genetic health of North America’s captive ring-tailed lemurs. Their mother Sophia was among 62 ring-tailed lemurs recommended for breeding across 20 institutions nationwide in 2016. Photo by David Haring, Duke Lemur Center.

Distant primate cousins with long black-and-white striped tails, ring-tailed lemurs live on the African island of Madagascar and nowhere else except in zoos and other captive facilities.

Some studies suggest that as few as 2,500 ring-tailed lemurs live in the wild today. Habit loss, hunting and the illegal pet trade have reduced their numbers by at least 50 percent in recent decades.

An additional estimated 2,500 ring-tailed lemurs live in zoos around the world, where experts work to maintain their genetic health in captivity.

The researchers studied DNA sequence variation at a region of the major histocompatibility complex, or MHC, a part of the genome that helps the immune system identify disease-causing bacteria, viruses and parasites.

Because different MHC gene variants recognize different types of pathogens, greater MHC diversity means animals are able to fend off a wider array of invaders.

The researchers estimated the number of MHC variants in 121 captive individuals born at the Duke Lemur Center and the Indianapolis and Cincinnati Zoos between 1980 and 2010.

They also compared them with 180 wild individuals from southwestern Madadgascar at the Bezà Mahafaly Special Reserve, where the animals regularly interbreed with lemurs from nearby forests.

Not surprisingly, MHC diversity was lower in captivity than in the wild.

Today’s captive ringtails came from a small group of ancestors that carried only a small fraction of the total genetic variation found in the larger wild population. Since their establishment, gene flow between captive populations and wild lemurs has been restricted.

Overall, the researchers found 20 unique MHC variants in the captive population, fewer than half the number in their wild counterparts.

However, efforts to identify good genetic matches across dozens of institutions have helped to preserve and even improve upon the diversity that is left.

For infants born at the Duke Lemur Center, MHC gene diversity remained low but stable for three decades from 1980 to 2010, then increased significantly from 2010 to 2013, researchers found.

Genetic contributions from several transplants contributed to the comeback.

An arranged marriage between ring-tailed lemurs at the Duke Lemur Center in North Carolina produced healthy twins Griselda and Hedwig in 2016. The infants are among 40 to 60 ring-tailed lemur infants born in North American zoos and other facilities each year. Photo by David Haring, Duke Lemur Center.

An arranged marriage between ring-tailed lemurs at the Duke Lemur Center in North Carolina produced healthy twins Griselda and Hedwig in 2016. The infants are among 40 to 60 ring-tailed lemur infants born in North American zoos and other facilities each year. Photo by David Haring, Duke Lemur Center.

The American Association of Zoos and Aquariums (AZA) tries to maintain a genetically healthy population by moving animals between institutions as potential mates. A team of experts uses computer software to help pick the best pairs for breeding.

Between 1980 and 2013, more than 1,160 ring-tailed lemurs were transferred between 217 institutions in North America alone.

In 2009, a male named Randy was transferred from the Saint Louis Zoo to the Duke Lemur Center for pairing with Sprite, a resident female. Experts also brought a mother-daughter pair, Schroeder and Leisl from the Zoo at Chehaw in Georgia, as potential mates for a resident male named Aracus.

“They saw an immediate improvement in the diversity of the offspring that were born,” said lead author Kathleen Grogan, who conducted the study while working on a doctorate with co-author Christine Drea at Duke University.

Grogan and colleagues are now examining whether MHC gene diversity helps the animals live longer or produce more offspring, as has been shown for other species.

“Not only do these lemurs serve as an assurance against extinction of their Malagasy counterparts, but maintaining as many variations of genes is important for keeping the individual lemurs, as well as the population healthy for any future challenges it may face,” said AZA Species Survival Plan Coordinator Gina Ferrie, a population biologist at Disney’s Animal Kingdom.

Conserving genetic diversity in captive populations over multiple generations is challenging due to their small size and relative isolation, but careful breeding can stem the loss, said Grogan, now a postdoctoral fellow at Pennsylvania State University.

Other authors include Michelle Sauther of the University of Colorado-Boulder and Frank Cuozzo at LaJuma Research Centre in South Africa.

This research was supported by Duke University, the International Primatological Society, Primate Conservation Inc., the University of Colorado-Boulder, the University of North Dakota, the National Science Foundation (BCS 0922465, BCS-1232570, IOS-071900), the Margot Marsh Biodiversity Foundation, the St. Louis Zoo and the American Society of Primatologists.

CITATION:  “Genetic Wealth, Population Health: Major Histocompatibility Complex Variation in Captive and Wild Ring-Tailed Lemurs (Lemur Catta),” Kathleen Grogan, Michelle Sauther, Frank Cuozzo and Christine Drea. Ecology and Evolution, Date. DOI: 10.1002/ece3.3317

Duke Scientists Visit Raleigh to Share Their Work

This post by graduate student Dan Keeley originally appeared on Regeneration NEXT. It is a followup to one of our earlier posts.

As a scientist, it is easy to get caught up in the day-to-day workflow of research and lose sight of the bigger picture. We are often so focused on generating and reporting solid, exciting data that we neglect another major aspect of our job; sharing our work and its impacts with the broader community. On Tuesday May 23rd, a group of graduate students from Duke went to the North Carolina legislative building to do just that.

L-R: Andrew George, Representative Marcia Morey (Durham County), Senator Terry Van Duyn (Buncombe County), Sharlini Sankaran, Dan Keeley, and Will Barclay at the NC legislative building.

Dr. Sharlini Sankaran, Executive Director of Duke’s Regeneration Next Initiative, organized a group of graduate students to attend the North Carolina Hospital Associations (NCHA) “Partnering for a Healthier Tomorrow!” advocacy day at the state legislature in Raleigh. The event gave representatives from various hospital systems an opportunity to interact with state legislators about the work they do and issues affecting healthcare in the state. Andrew George, a graduate student in the McClay Lab, Will Barclay, a graduate student in the Shinohara Lab, and I joined Dr. Sankaran to share some of the great tissue regeneration-related research going on at Duke.

Our morning was busy as elected officials, legislative staff, executive branch agency officials, and staff from other hospital systems stopped by our booth to hear what Regeneration Next is all about. We talked about the focus on harnessing Duke’s strengths in fundamental research on molecular mechanisms underlying regeneration and development, then pairing that with the expertise of our engineers and clinicians. We discussed topics including spine and heart regeneration mechanisms from the Poss Lab, advances in engineering skeletal muscle from the Bursac Lab, and clinical trials of bioengineered blood vessels for patients undergoing dialysis from Duke faculty Dr. Jeffrey Lawson.

It was remarkable to hear how engaged everyone was, we got great questions like ‘what is a zebrafish and why do you use them?’ and ‘why would a bioengineered ligament be better than one from an animal model or cadaver?’.  Every person who stopped by was supportive and many had a personal story to share about a health issue experienced by friends, family, or even themselves. As a graduate student who does basic research, it really underscored how important these personal connections are to our work, even though it may be far removed from the clinic.

Communicating our research to legislators and others at NCHA advocacy day was a great and encouraging experience. Health issues affect all of us. Our visit to the legislature on Tuesday was a reminder that there is support for the work that we do in hopes it will help lead to a healthier tomorrow.

Guest post by Dan Keeley, graduate student in BiologyDan Keeley

Scientists Engineer Disease-Resistant Rice Without Sacrificing Yield

Researchers have developed a way to make rice more resistant to bacterial blight and other diseases without reducing yield. Photo by Max Pixel.

Researchers have successfully developed a novel method that allows for increased disease resistance in rice without decreasing yield. A team at Duke University, working in collaboration with scientists at Huazhong Agricultural University in China, describe the findings in a paper published May 17, 2017 in the journal Nature.

Rice is one of the most important staple crops, responsible for providing over one-fifth of the calories consumed by humans worldwide. Diseases caused by bacterial or fungal pathogens present a significant problem, and can result in the loss of 80 percent or more of a rice crop.

Decades of research into the plant immune response have identified components that can be used to engineer disease-resistant plants. However, their practical application to crops is limited due to the decreased yield associated with a constantly active defense response.

“Immunity is a double-edged sword, ” said study co-author Xinnian Dong, professor of biology at Duke and lead investigator of the study. “There is often a tradeoff between growth and defense because defense proteins are not only toxic to pathogens but also harmful to self when overexpressed,” Dong said. “This is a major challenge in engineering disease resistance for agricultural use because the ultimate goal is to protect the yield.”

Previous studies have focused on altering the coding sequence or upstream DNA sequence elements of a gene. These upstream DNA elements are known as promoters, and they act as switches that turn on or off a gene’s expression. This is the first step of a gene’s synthesis into its protein product, known as transcription.

By attaching a promoter that gives an “on” signal to a defense gene, a plant can be engineered to be highly resistant to pathogens, though at a cost to growth and yield. These costs can be partially alleviated by attaching the defense gene to a “pathogen specific” promoter that turns on in the presence of pathogen attack.

To further alleviate the negative effects of active defense, the Dong group sought to add an additional layer of control. They turned newly discovered sequence elements, called upstream open reading frames (uORFs), to help address this problem. These sequence elements act on the intermediate of a gene, or messenger (RNA, a molecule similar to DNA) to govern its “translation” into the final protein product. A recent study by the Dong lab in an accompanying paper in Nature has identified many of these elements that respond in a pathogen-inducible manner.

The Dong group hypothesized that adding this pathogen-inducible translational regulation would result in a tighter control of defense protein expression and minimize the lost yield associated with enhanced disease resistance.

To test this hypothesis, the researchers started with Arabidopsis, a flowering plant commonly used in laboratory research. They created a DNA sequence that contains both the transcriptional and translational elements (uORFs) and fused them upstream of the potent “immune activator” gene called snc1. This hybrid sequence was called a “transcriptional/translational cassette” and was inserted into Arabidopsis plants.

When plants have snc1 constitutively active, they are highly resistant to pathogens, but have severely stunted growth. Strikingly, plants with the transcriptional/translational cassette not only have increased resistance, but they also lacked growth defects and resembled healthy wild-type plants. These results show the benefits of adding translational control in engineering plants that have increased resistance without significant costs.

The Dong group then sought to apply these findings to engineer disease-resistant rice, as it is one of the world’s most important crops. They created transgenic rice lines containing the transcriptional/translational cassette driving expression of another potent “immune activator” gene called AtNPR1. This gene was chosen as it has been found to confer broad spectrum pathogen resistance in a wide variety of crop species, including rice, citrus, apple and wheat.

The dry yellowish leaves on these rice plants are a classic symptom of bacterial blight, a devastating disease that affects rice fields worldwide. Photo by Meng Yuan.

The transgenic rice lines containing the transcriptional/translational cassette were infected with bacterial/fungal pathogens that cause three major rice diseases — rice  blight, leaf streak, and fungal blast. These showed high resistance to all three pathogens, indicating broad spectrum resistance could be achieved. Importantly, when grown in the field, their yield — both in terms of grain quantity and quality per plant — was almost unaffected. These results indicate a great potential for agricultural applications.

This strategy is the first known use of adding translational control for the engineering of disease-resistant crops with minimal yield costs. It has many advantages, as it is broadly applicable to a variety of crop species against many pathogens. Since this strategy involves activating the plants’ endogenous defenses, it may also reduce the use of pesticides on crops and hence protect the environment.

Additionally, these findings may be broadly applicable to other systems as well. These upstream elements (uORFs) are widely present in organisms from yeast to humans, with nearly half of all human transcripts containing them. “The great potential in using these elements in controlling protein translation during specific biological processes has yet to be realized,” Dong said.

Corresponding author Xinnian Dong can be reached at xdong@duke.edu or (919) 613-8176.

CITATION:  “uORF-Mediated Translation Allows Engineered Plant Disease Resistance Without Fitness Costs,” Guoyong Xu, Meng Yuan,   Chaoren Ai, Lijing Liu, Edward Zhuang, Sargis Karapetyan, Shiping Wang and Xinnian Dong. Nature, May 17, 2017. DOI: 10.1038/nature22372

 

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