GN/ Structures considered key to gene expression are surprisingly fleeting
Genetics biweekly vol.26, 7th April — 20th April
TL;DR
- Scientists find that loops in the genome may be much rarer and shorter-lived than previously thought, lasting only tens of minutes, which suggests current theories of how loops influence gene expression may need to be revised.
- Gut microbiota by-products circulate in the bloodstream, regulating host physiological processes including immunity, metabolism and brain functions. Scientists have discovered that hypothalamic neurons in an animal model directly detect variations in bacterial activity and adapt appetite and body temperature accordingly. These findings demonstrate that a direct dialog occurs between the gut microbiota and the brain, a discovery that could lead to new therapeutic approaches for tackling metabolic disorders such as diabetes and obesity.
- A new study describes a breakthrough in the quest to improve photosynthesis in certain crops, a step toward adapting plants to rapid climate changes and increasing yields to feed a projected 9 billion people by 2050.
- Current antibody treatments block SARS-CoV-2 by binding to one of three binding sites on the spike protein. A new protein-based antiviral binds to all three sites on the spike protein, making it more effective than current therapies. The new therapy also is low-cost, easy to manufacture, does not require complicated supply chains with extreme refrigeration and potentially could be self-administered.
- Researchers have found that Zika virus can mutate to become more infective — and potentially break through pre-existing immunity.
- Given enough time and energy, the body will heal, but when doctors or engineers intervene, the processes do not always proceed as planned because chemicals that control and facilitate the healing process are missing. Now, an international team of engineers is bioprinting bone along with two growth factor encoding genes that help incorporate the cells and heal defects in the skulls of rats.
- New research breakthrough bridges a complexity gap between chemistry and biology and provides a new methodology that uses designed mixtures to engineer adaptive properties that are normally only associated with living systems.
- Researchers have engineered a strain of bacteria that can help protect the natural flora of the human digestive tract from antibiotics and curb the emergence of antimicrobial resistance.
- A new study compares the accumulation of mutations across many animal species and has shed new light on decades-old questions about the role of these genetic changes in ageing and cancer. Researchers found that despite huge variation in lifespan and size, different animal species end their natural life with similar numbers of genetic changes.
- Decoy nanoparticles mimic cells, attracting viruses to bind to them rather than infecting healthy cells. Researchers tested the strategy against the novel coronavirus and five of its variants, finding it was consistently effective.
- And more!
Overview
Genetic technology is defined as the term which includes a range of activities concerned with the understanding of gene expression, advantages of natural genetic variation, modifying genes and transferring genes to new hosts. Genes are found in all living organisms and are transferred from one generation to the next. Gene technology encompasses several techniques including marker-assisted breeding, RNAi and genetic modification. Only some gene technologies produce genetically modified organisms.
Modern genetic technologies like genome editing would not be possible without all the previous generations of genetic technologies that have enabled scientists to discover what genes are, what they do and how DNA can be modified to add, remove or replace genes. You can find major genetic technologies development milestones via the link.
Gene Technology Market
- The valuation of the genetic engineering market is projected to escalate to USD 6.90 MN by the end of 2027.
- Global Genetic Engineering Market is projected to grow at 12.48% CAGR during the assessment period (2017–2027).
- North America holds the largest share in the global genetic engineering market, followed by Europe and the Asia Pacific, respectively.
- Another research provider, MarketsandMarkets, forecasts the genome editing, genome engineering market to grow from USD 3.19 billion in 2017 to USD 6.28 billion by 2022, at a compounded annual growth rate (CAGR) of 14.5% during the forecast period. The key factors propelling market growth are rising government funding and growth in the number of genomics projects, high prevalence of infectious diseases (like COVID-19) and cancer, technological advancements, increasing production of genetically modified (GM) crops, and growing application areas of genomics.
Latest News & Research
Dynamics of CTCF- and cohesin-mediated chromatin looping revealed by live-cell imaging
by Michele Gabriele, Hugo B. Brandão, Simon Grosse-Holz, Asmita Jha, et al in Science
In human chromosomes, DNA is coated by proteins to form an exceedingly long beaded string. This “string” is folded into numerous loops, which are believed to help cells control gene expression and facilitate DNA repair, among other functions. A new study from MIT suggests that these loops are very dynamic and shorter-lived than previously thought.
In the new study, the researchers were able to monitor the movement of one stretch of the genome in a living cell for about two hours. They saw that this stretch was fully looped for only 3 to 6 percent of the time, with the loop lasting for only about 10 to 30 minutes. The findings suggest that scientists’ current understanding of how loops influence gene expression may need to be revised, the researchers say.
“Many models in the field have been these pictures of static loops regulating these processes. What our new paper shows is that this picture is not really correct,” says Anders Sejr Hansen, the Underwood-Prescott Career Development Assistant Professor of Biological Engineering at MIT. “We suggest that the functional state of these domains is much more dynamic.”
Hansen is one of the senior authors of the new study, along with Leonid Mirny, a professor in MIT’s Institute for Medical Engineering and Science and the Department of Physics, and Christoph Zechner, a group leader at the Max Planck Institute of Molecular Cell Biology and Genetics in Dresden, Germany, and the Center for Systems Biology Dresden. MIT postdoc Michele Gabriele, recent Harvard University PhD recipient Hugo Brandão, and MIT graduate student Simon Grosse-Holz are the lead authors of the paper.
Using computer simulations and experimental data, scientists including Mirny’s group at MIT have shown that loops in the genome are formed by a process called extrusion, in which a molecular motor promotes the growth of progressively larger loops. The motor stops each time it encounters a “stop sign” on DNA. The motor that extrudes such loops is a protein complex called cohesin, while the DNA-bound protein CTCF serves as the stop sign. These cohesin-mediated loops between CTCF sites were seen in previous experiments. However, those experiments only offered a snapshot of a moment in time, with no information on how the loops change over time. In their new study, the researchers developed techniques that allowed them to fluorescently label CTCF DNA sites so they could image the DNA loops over several hours. They also created a new computational method that can infer the looping events from the imaging data.
“This method was crucial for us to distinguish signal from noise in our experimental data and quantify looping,” Zechner says. “We believe that such approaches will become increasingly important for biology as we continue to push the limits of detection with experiments.”
The researchers used their method to image a stretch of the genome in mouse embryonic stem cells. “If we put our data in the context of one cell division cycle, which lasts about 12 hours, the fully formed loop only actually exists for about 20 to 45 minutes, or about 3 to 6 percent of the time,” Grosse-Holz says.
“If the loop is only present for such a tiny period of the cell cycle and very short-lived, we shouldn’t think of this fully looped state as being the primary regulator of gene expression,” Hansen says. “We think we need new models for how the 3D structure of the genome regulates gene expression, DNA repair, and other functional downstream processes.”
While fully formed loops were rare, the researchers found that partially extruded loops were present about 92 percent of the time. These smaller loops have been difficult to observe with the previous methods of detecting loops in the genome.
“In this study, by integrating our experimental data with polymer simulations, we have now been able to quantify the relative extents of the unlooped, partially extruded, and fully looped states,” Brandão says.
“Since these interactions are very short, but very frequent, the previous methodologies were not able to fully capture their dynamics,” Gabriele adds. “With our new technique, we can start to resolve transitions between fully looped and unlooped states.”
The researchers hypothesize that these partial loops may play more important roles in gene regulation than fully formed loops. Strands of DNA run along each other as loops begin to form and then fall apart, and these interactions may help regulatory elements such as enhancers and gene promoters find each other.
“More than 90 percent of the time, there are some transient loops, and presumably what’s important is having those loops that are being perpetually extruded,” Mirny says. “The process of extrusion itself may be more important than the fully looped state that only occurs for a short period of time.”
Since most of the other loops in the genome are weaker than the one the researchers studied in this paper, they suspect that many other loops will also prove to be highly transient. They now plan to use their new technique study some of those other loops, in a variety of cell types.
“There are about 10,000 of these loops, and we’ve looked at one,” Hansen says. “We have a lot of indirect evidence to suggest that the results would be generalizable, but we haven’t demonstrated that. Using the technology platform we’ve set up, which combines new experimental and computational methods, we can begin to approach other loops in the genome.”
The researchers also plan to investigate the role of specific loops in disease. Many diseases, including a neurodevelopmental disorder called FOXG1 syndrome, could be linked to faulty loop dynamics. The researchers are now studying how both the normal and mutated form of the FOXG1 gene, as well as the cancer-causing gene MYC, are affected by genome loop formation.
Bacterial sensing via neuronal Nod2 regulates appetite and body temperature
by Ilana Gabanyi, Gabriel Lepousez, Richard Wheeler, et al in Science
Gut microbiota by-products circulate in the bloodstream, regulating host physiological processes including immunity, metabolism and brain functions. Scientists from the Institut Pasteur (a partner research organization of Université Paris Cité), Inserm and the CNRS have discovered that hypothalamic neurons in an animal model directly detect variations in bacterial activity and adapt appetite and body temperature accordingly. These findings demonstrate that a direct dialog occurs between the gut microbiota and the brain, a discovery that could lead to new therapeutic approaches for tackling metabolic disorders such as diabetes and obesity.
The gut is the body’s largest reservoir of bacteria. A growing body of evidence reveals the degree of interdependence between hosts and their gut microbiota, and emphasizes the importance of the gut-brain axis. At the Institut Pasteur, neurobiologists from the Perception and Memory Unit (Institut Pasteur/CNRS), immunobiologists from the Microenvironment and Immunity Unit (Institut Pasteur/Inserm), and microbiologists from the Biology and Genetics of the Bacterial Cell Wall Unit (Institut Pasteur/CNRS/Inserm) have shared their expertise to investigate how bacteria in the gut directly control the activity of particular neurons in the brain.
Food consumption induces expansion of the intestinal microbiota. This expansion is followed by an increase in muropeptide release from the gut bacteria. When they reach the brain, these muropeptides target a subset of inhibitory hypothalamic neurons. In older females, activation of neuronal Nod2 receptors by muropeptides decreases neuronal activity, which in turn helps to regulate satiety and body temperature.
The scientists focused on the NOD2 (nucleotide oligomerization domain) receptor which is found inside of mostly immune cells. This receptor detects the presence of muropeptides, which are the building blocks of the bacterial cell wall. Moreover, it has previously been established that variants of the gene coding for the NOD2 receptor are associated with digestive disorders, including Crohn’s disease, as well as neurological diseases and mood disorders. However, these data were insufficient to demonstrate a direct relationship between neuronal activity in the brain and bacterial activity in the gut. This was revealed by the consortium of scientists in the new study.
Using brain imaging techniques, the scientists initially observed that the NOD2 receptor in mice is expressed by neurons in different regions of the brain, and in particular, in a region known as the hypothalamus. They subsequently discovered that these neurons’ electrical activity is suppressed when they come into contact with bacterial muropeptides from the gut. “Muropeptides in the gut, blood and brain are considered to be markers of bacterial proliferation,” explains Ivo G. Boneca, Head of the Biology and Genetics of the Bacterial Cell Wall Unit at the Institut Pasteur (CNRS/Inserm). Conversely, if the NOD2 receptor is absent, these neurons are no longer suppressed by muropeptides. Consequently, the brain loses control of food intake and body temperature. The mice gain weight and are more susceptible to developing type 2 diabetes, particularly in older females.
In this study, the scientists have demonstrated the astonishing fact that neurons perceive bacterial muropeptides directly, while this task was thought to be primarily assigned to immune cells. “It is extraordinary to discover that bacterial fragments act directly on a brain center as strategic as the hypothalamus, which is known to manage vital functions such as body temperature, reproduction, hunger and thirst,” comments Pierre-Marie Lledo, CNRS scientist and Head of the Institut Pasteur’s Perception and Memory Unit.
The neurons thus appear to detect bacterial activity (proliferation and death) as a direct gauge of the impact of food intake on the intestinal ecosystem.
“Excessive intake of a specific food may stimulate the disproportionate growth of certain bacteria or pathogens, thus jeopardizing intestinal balance,” says Gérard Eberl, Head of the Microenvironment and Immunity Unit at the Institut Pasteur (Inserm).
The impact of muropeptides on hypothalamic neurons and metabolism raises questions on their potential role in other brain functions, and may help us understand the link between certain brain diseases and genetic variants of NOD2. This discovery paves the way for new interdisciplinary projects at the frontier between neurosciences, immunology and microbiology, and ultimately, for new therapeutic approaches to brain diseases and metabolic disorders such as diabetes and obesity.
Improving the efficiency of Rubisco by resurrecting its ancestors in the family Solanaceae
by Myat T. Lin, Heidi Salihovic, Frances K. Clark, Maureen R. Hanson in Science Advances
A Cornell University study describes a breakthrough in the quest to improve photosynthesis in certain crops, a step toward adapting plants to rapid climate changes and increasing yields to feed a projected 9 billion people by 2050.
The authors developed a computational technique to predict favorable gene sequences that make Rubisco, a key plant enzyme for photosynthesis. The technique allowed the scientists to identify promising candidate enzymes that could be engineered into modern crops and, ultimately, make photosynthesis more efficient and increase crop yields.The senior author of the study is Maureen Hanson, the Liberty Hyde Bailey Professor of Plant Molecular Biology in the College of Agriculture and Life Sciences. First author Myat Lin is a postdoctoral research associate in Hanson’s lab.
Their method relied on evolutionary history, where the researchers predicted Rubisco genes from 20–30 million years ago, when Earth’s carbon dioxide (CO2) levels were higher than they are today and the Rubisco enzymes in plants were adapted to those levels. By resurrecting ancient Rubisco, early results show promise for developing faster, more efficient Rubisco enzymes to incorporate into crops and help them adapt to hot, dry future conditions, as human activities are increasing heat-trapping CO2 gas concentrations in Earth’s atmosphere.
The study describes predictions of 98 Rubisco enzymes at key moments in the evolutionary history of plants in the Solanaceae family, which include tomato, pepper, potato, eggplant and tobacco. Researchers use tobacco as the experimental model for their studies of Rubisco.
“We were able to identify predicted ancestral enzymes that do have superior qualities compared to current-day enzymes,” Hanson said. Lin developed the new technique for identifying predicted ancient Rubisco enzymes.
Scientists have known that they can increase crop yields by accelerating photosynthesis, where plants convert CO2, water and light into oxygen and sugars that plants use for energy and for building new tissues. For many years, researchers have focused on Rubisco, a slow enzyme that pulls (or fixes) carbon from CO2 to create sugars. Aside from being slow, Rubisco also sometimes catalyzes a reaction with oxygen in the air; by so doing, it creates a toxic byproduct, wastes energy and makes photosynthesis inefficient.
Hanson’s lab had previously tried to use Rubisco from cyanobacteria (blue-green algae), which is faster but also reacts readily with oxygen, forcing the researchers to try to create micro-compartments to protect the enzyme from oxygen, with mixed results. Other researchers have tried to engineer more optimal Rubisco by making changes in the enzyme’s amino acids, though little was known about which changes would lead to desired results.
In this study, Lin reconstructed a phylogeny — a tree-like diagram showing evolutionary relatedness among groups of organisms — of Rubisco, using Solanaceae plants.
“By getting a lot of [genetic] sequences of Rubisco in existing plants, a phylogenetic tree could be constructed to figure out which Rubiscos likely existed 20 to 30 million years ago,” Hanson said.
The advantage of identifying potential ancient Rubisco sequences is that carbon dioxide levels were possibly as high as 500 to 800 parts per million (ppm) in the atmosphere 25 million to 50 million years ago. Today, heat-trapping CO2 levels are rising sharply due to many human activities, with current measurements at around 420 ppm, after staying relatively constant under 300 ppm for hundreds of millennia until the 1950s. Lin, Hanson and colleagues then used an experimental system developed for tobacco in Hanson’s lab, and described in a 2020 Nature Plants paper, which employs E. coli bacteria to test in a single day the efficacy of different versions of Rubisco. Similar tests done in plants take months to verify. The team found that ancient Rubisco enzymes predicted from modern-day Solanaceae plants showed real promise for being more efficient.
“For the next step, we want to replace the genes for the existing Rubisco enzyme in tobacco with these ancestral sequences using CRISPR [gene-editing] technology, and then measure how it affects the production of biomass,” Hanson said. “We certainly hope that our experiments will show that by adapting Rubisco to present day conditions, we will have plants that will give greater yields.”
If their method proves successful, these efficient Rubisco sequences could be transferred into crops such as tomatoes, as well as those from other plant families, such as soybeans and rice.
Multivalent designed proteins neutralize SARS-CoV-2 variants of concern and confer protection against infection in mice
by Andrew C. Hunt, James Brett Case, Young-Jun Park, et al in Science Translational Medicine
A new protein-based antiviral nasal spray developed by researchers at Northwestern University, University of Washington and Washington University at St. Louis is being advanced toward Phase I human clinical trials to treat COVID-19.
Designed computationally and refined in the laboratory, the new protein therapies thwarted infection by interfering with the virus’ ability to enter cells. The top protein neutralized the virus with similar or greater potency than antibody treatments with Emergency Use Authorization status from the U.S. Food and Drug Administration (FDA). Notably, the top protein also neutralized all tested SARS-CoV-2 variants, something that many clinical antibodies have failed to do.
When researchers administered the treatment to mice as a nasal spray, they found that the best of these antiviral proteins reduced symptoms of infection — or even prevented infection outright. This work was led by Northwestern’s Michael Jewett; David Baker and David Veesler at the University of Washington School of Medicine; and Michael S. Diamond at WashU.
To begin, the team first used supercomputers to design proteins that could stick to vulnerable sites on the surface of the novel coronavirus, targeting the spike protein. In the new work, the team reengineered the proteins — called minibinders — to make them even more potent. Rather than targeting just one site of the virus’ infectious machinery, the minibinders simultaneously bind to three sites, making the drug less likely to detach.
“SARS-CoV-2’s spike protein has three binding domains, and common antibody therapies may only block one,” Jewett said. “Our minibinders sit on top of the spike protein like a tripod and block all three. The interaction between the spike protein and our antiviral is among the tightest interactions known in biology. When we put the spike protein and our antiviral therapeutic in a test tube together for a week, they stayed connected and never fell apart.”
Jewett is a professor of chemical and biological engineering at Northwestern’s McCormick School of Engineering and director of Northwestern’s Center for Synthetic Biology. Andrew C. Hunt, a graduate research fellow in Jewett’s laboratory, is the paper’s co-first author.
As the SARS-CoV-2 virus has mutated to create new variants, some treatments have become less effective in fighting the ever-evolving virus. Just last month, the FDA paused several monoclonal antibody treatments, for example, due to their failure against the BA.2 omicron subvariant. Unlike these antibody treatments, which failed to neutralize omicron, the new minibinders maintained potency against the omicron variant of concern. By blocking the virus’ spike protein, the new antiviral prevents it from binding to the human angiotensin-converting enzyme 2 (ACE2) receptor, which is the entry point for infecting the body. Because the novel coronavirus and its mutant variants cannot infect the body without binding to the ACE2 receptor, the antiviral also should work against future variants.
“To enter the body, the spike protein and ACE2 receptor engage in a handshake,” Jewett said. “Our antiviral blocks this handshake and, as a bonus, has resistance to viral escape.”
In addition to losing effectiveness, current antibody therapies also come with several problems: They are difficult to develop, expensive and require a healthcare professional to administer. They also require complicated supply chains and extreme refrigeration, which is often unavailable in low-resource settings.
The new antiviral solves all these problems. As opposed to monoclonal antibodies, which are made by cloning and culturing living mammalian cells, the new antiviral treatment is produced large-scale in microorganisms like E. coli, making them more cost-effective to manufacture. Not only is the new therapy stable in high heat, which could further streamline manufacturing and decrease the cost of goods for clinical development, it also holds promise for being self-administered as a one-time nasal spray, bypassing the need for medical professionals.
Controlled Co-delivery of pPDGF-B and pBMP-2 from intraoperatively bioprinted bone constructs improves the repair of calvarial defects in rats
by Kazim K. Moncal, R. Seda Tigli Aydın, Kevin P. Godzik, Timothy M. Acri, Dong N. Heo, Elias Rizk, Hwabok Wee, Gregory S. Lewis, Aliasger K. Salem, Ibrahim T. Ozbolat in Biomaterials
Given enough time and energy, the body will heal, but when doctors or engineers intervene, the processes do not always proceed as planned because chemicals that control and facilitate the healing process are missing. Now, an international team of engineers is bioprinting bone along with two growth factor encoding genes that help incorporate the cells and heal defects in the skulls of rats.
“Growth factors are essential for cell growth,” said Ibrahim T. Ozbolat, associate professor of engineering science and mechanics. “We use two different genes encoding two different growth factors. These growth factors help stem cells to migrate into the defect area and then help the progenitor cells to convert into bone.”
The researchers used gene encoding PDGF-B, platelet derived-growth factor, which encourages cells to multiply and to migrate, and gene encoding BMP-2, bone morphogenetic protein, which improves bone regeneration. They delivered both genes using bioprinting.
“We used a controlled co-delivery release of plasmids from a gene-activated matrix to promote bone repair,” the researchers stated.
Ozbolat and his team embedded the DNA for the protein in plasmids — ringlike loops of DNA that can transport genetic information. Once the DNA enters the progenitor cell, it begins to produce the appropriate proteins to enhance bone growth. The two genes were printed during surgery onto a hole in the skull of a rat using a device very similar to an ink-jet printer. The mixture was created to release a burst of PDGF-B encoding gene in 10 days and a continuing release of BMP-2 encoding gene for five weeks. The rats that received bioprinted genes with controlled release of BMP-2 encoding gene saw about 40% bone tissue creation and 90% bone coverage in six weeks compared to 10% new bone tissue and 25% bone coverage for rats with the same defect, but no treatment.
“This method is better than simply dumping the growth factors,” said Ozbolat. “If we do that, the amounts of proteins are finite, but if we use gene therapy, the cells continue to produce the necessary growth factors.”
A Zika Virus Mutation Enhances Transmission Potential and Confers Escape from Protective Dengue Virus Immunity
by Jose Angel Regla-Nava et al. in Cell Reports
Researchers at La Jolla Institute for Immunology (LJI) have found that Zika virus can mutate to become more infective — and potentially break through pre-existing immunity.
“The world should monitor the emergence of this Zika virus variant,” says LJI Professor Sujan Shresta, Ph.D., who co-led the study with Professor Pei-Yong Shi, Ph.D., of the University of Texas Medical Branch (UTMB).
Zika virus is carried by mosquitoes, and the symptoms of Zika infection are usually mild in adults. However, the virus can infect a developing fetus, resulting in birth defects such as microcephaly. Zika virus and dengue virus overlap in many countries worldwide. Like Zika, dengue virus is a mosquito-borne flavivirus, and thus shares many biological properties. In fact, the viruses are similar enough that the immune response sparked by prior dengue exposure can offer protection against Zika.
“In areas where Zika is prevalent, a vast majority of people have already been exposed to dengue virus and have both T cells and antibodies that cross-react,” says Shresta.
Unfortunately, both viruses are also quick to mutate. “Dengue and Zika are RNA viruses, which means they can change their genome,” explains Shresta. “When there are so many mosquitoes and so many human hosts, these viruses are constantly moving back and forth and evolving.”
To study Zika’s fast-paced evolution, the LJI team recreated infection cycles that repeatedly switched back and forth between mosquito cells and mice. This work gave the LJI scientists a window into how Zika virus naturally evolves as it encounters more hosts. The researchers found it is relatively easy for Zika virus to acquire a single amino acid change that allows the virus to make more copies of itself — and help infections take hold more easily. This mutation (called NS2B I39V/I39T mutation) boosts the virus’s ability to replicate in both mice and mosquitoes. This Zika variant also showed increased replication in human cells.
“This single mutation is sufficient to enhance Zika virus virulence,” says study first author Jose Angel Regla-Nava, Ph.D., former postdoctoral researcher at LJI and current Associate Professor at the University of Guadalajara, Mexico. “A high replication rate in either a mosquito or human host could increase viral transmission or pathogenicity — and cause a new outbreak.”
Adds Shresta, “The Zika variant that we identified had evolved to the point where the cross-protective immunity afforded by prior dengue infection was no longer effective in mice. Unfortunately for us, if this variant becomes prevalent, we may have the same issues in real life.”
So how can we prepare for this kind of variant? Shresta’s laboratory is already looking at ways to tailor Zika vaccines and treatments that counteract this dangerous mutation. She will also continue to work closely with Regla-Nava to better understand exactly how this mutation helps Zika replicate more efficiently.
“We want to understand at what point in the viral life cycle this mutation makes a difference,” says Shresta.
Tractable molecular adaptation patterns in a designed complex peptide system
by Ankit Jain, Scott A. McPhee, Tong Wang, Maya Narayanan Nair, Daniela Kroiss, Tony Z. Jia, Rein V. Ulijn in Chem
A post-doctoral researcher with the Advanced Science Research Center at the CUNY Graduate Center (CUNY ASRC) has made an important step toward understanding how complex mixtures of biomolecular building blocks form self-organized patterns.
The discovery — authored by Ankit Jain, a member of CUNY ASRC Nanoscience Initiative Director Rein Ulijn’s lab — provides new knowledge about adaptive biological functions, which could be critical in designing novel materials and technologies with similar abilities and attributes.
“All life forms start with the same conserved sets of building blocks, which includes the 20 amino acids that make up proteins,” said Jain. “Figuring out how mixtures of these molecules communicate, interact and form self-organizing patterns would enhance our understanding of how biology creates functionality. This understanding could also give rise to completely new ways of creating materials and technologies that incorporate life processes such as adapting, growing, healing and developing new properties when required.”
Jain took a new, synthetic, approach to begin uncovering how complex biomolecule mixtures interact and collectively adapt to changes in their environment. Instead of trying to disentangle molecular organization in existing systems, such as those found in biological cells, he addressed the problem in a test tube by creating mixtures with components designed to react and interact. Jain then tracked and observed the emergence of increasingly complex patterns that the biomolecules spontaneously formed in response to changes in their environment.
“Complex mixtures of interacting molecules are fundamental to life processes, but they are not commonly studied in chemistry labs, because they are messy, very complicated and difficult to study and understand,” said Ulijn. “Systematically designing mixtures and tracking their behavior allows us to make fundamental observations about how mixtures of molecules become functional collectives. We were able to detail how these chemical systems absorb changes in external conditions to form specific patterns of build-up and breakdown. We also discovered that systems with so many variables show a stochastic behavior, so while overall pattern formation looks similar when running multiple experiments, the precise details in two independent experiments are different.”
Jain’s experiment began with mixing a number of selected dipeptides, which are minimalistic protein-like compounds composed of two amino acids. These sets of dipeptides (designed based on their ability to aggregate and interact) also contained a catalyst that enabled the dipeptides to dynamically recombine and form peptides with more complex interaction patterns. The most complex system studied in this paper began with 15 different dipeptides, which reversibly combine to form 225 unique tetrapeptides. It was then possible for Jain to track the formation and breakdown of peptides of different sequence within the mixtures. He observed that their patterns of interaction were strongly dictated by environmental conditions.
Illuminating molecular self-organization through hierarchical patterns of both covalent and non-covalent interactions is key to understanding how biological functions relevant to life emerge. The new bottom-up approach enables researchers to understand, for the first time, ensemble characteristics while simultaneously providing molecular resolution of the information. The work demonstrates that mixtures of simple molecules demonstrate spontaneous sequence selection, which may provide insights into the chemical origins of biological function. Overall, the design of adaptive systems based on multi-component mixtures is likely to lead to discovery of how patterns dictate the formation of reconfigurable, functional materials that hold promise for future bioinspired technologies.
An engineered live biotherapeutic for the prevention of antibiotic-induced dysbiosis
by Cubillos-Ruiz, A., Alcantar, M.A., Donghia, N.M. et al. in Nature Biomedical Engineering
Antibiotics are life-saving drugs, but they can also harm the beneficial microbes that live in the human gut. Following antibiotic treatment, some patients are at risk of developing inflammation or opportunistic infections such as Clostridiodes difficile. Indiscriminate use of antibiotics on gut microbes can also contribute to the spread of resistance to the drugs.
In an effort to reduce those risks, MIT engineers have developed a new way to help protect the natural flora of the human digestive tract. They took a strain of bacteria that is safe for human consumption and engineered it to safely produce an enzyme that breaks down a class of antibiotics called beta-lactams. These include ampicillin, amoxicillin, and other commonly used drugs. When this “living biotherapeutic” is given along with antibiotics, it protects the microbiota in the gut but allows the levels of antibiotics circulating in the bloodstream to remain high, the researchers found in a study of mice.
“This work shows that synthetic biology can be harnessed to create a new class of engineered therapeutics for reducing the adverse effects of antibiotics,” says James Collins, the Termeer Professor of Medical Engineering and Science in MIT’s Institute for Medical Engineering and Science (IMES) and Department of Biological Engineering, and the senior author of the new study. Andres Cubillos-Ruiz PhD ’15, a research scientist at IMES and the Wyss Institute for Biologically Inspired Engineering at Harvard University, is the lead author of the paper. Other authors include MIT graduate students Miguel Alcantar and Pablo Cardenas, Wyss Institute staff scientist Nina Donghia, and Broad Institute research scientist Julian Avila-Pacheco.
Over the past two decades, research has revealed that the microbes in the human gut play important roles in not only metabolism but also immune function and nervous system function.
“Throughout your life, these gut microbes assemble into a highly diverse community that accomplishes important functions in your body,” Cubillos-Ruiz says. “The problem comes when interventions such as medications or particular kinds of diets affect the composition of the microbiota and create an altered state, called dysbiosis. Some microbial groups disappear, and the metabolic activity of others increases. This unbalance can lead to various health issues.”
One major complication that can occur is infection of C. difficile, a microbe that commonly lives in the gut but doesn’t usually cause harm. When antibiotics kill off the strains that compete with C. difficile, however, these bacteria can take over and cause diarrhea and colitis. C. difficile infects about 500,000 people every year in the United States, and causes around 15,000 deaths. Doctors sometimes prescribe probiotics (mixtures of beneficial bacteria) to people taking antibiotics, but those probiotics are usually also susceptible to antibiotics, and they don’t fully replicate the native microbiota found in the gut.
“Standard probiotics cannot compare to the diversity that the native microbes have,” Cubillos-Ruiz says. “They cannot accomplish the same functions as the native microbes that you have nurtured throughout your life.”
To protect the microbiota from antibiotics, the researchers decided to use modified bacteria. They engineered a strain of bacteria called Lactococcus lactis, which is normally used in cheese production, to deliver an enzyme that breaks down beta-lactam antibiotics. These drugs make up about 60 percent of the antibiotics prescribed in the United States. When these bacteria are delivered orally, they transiently populate the intestines, where they secrete the enzyme, which is called beta-lactamase. This enzyme then breaks down antibiotics that reach the intestinal tract. When antibiotics are given orally, the drugs enter the bloodstream primarily from the stomach, so the drugs can still circulate in the body at high levels. This approach could also be used along with antibiotics that are injected, which also end up reaching the intestine. After their job is finished, the engineered bacteria are excreted through the digestive tract.
Using engineered bacteria that degrade antibiotics poses unique safety requirements: Beta-lactamase enzymes confer antibiotic resistance to harboring cells and their genes can readily spread between different bacteria. To address this, the researchers used a synthetic biology approach to recode the way the bacterium synthetizes the enzyme. They broke up the gene for beta-lactamase into two pieces, each of which encodes a fragment of the enzyme. These gene segments are located on different pieces of DNA, making it very unlikely that both gene segments would be transferred to another bacterial cell. These beta-lactamase fragments are exported outside the cell where they reassemble, restoring the enzymatic function. Since the beta-lactamase is now free to diffuse in the surrounding environment, its activity becomes a “public good” for the gut bacterial communities. This prevents the engineered cells from gaining an advantage over the native gut microbes.
“Our biocontainment strategy enables the delivery of antibiotic-degrading enzymes to the gut without the risk of horizontal gene transfer to other bacteria or the acquisition of an added competitive advantage by the live biotherapeutic,” Cubillos-Ruiz says.
To test their approach, the researchers gave the mice two oral doses of the engineered bacteria for every injection of ampicillin. The engineered bacteria made their way to the intestine and began releasing beta-lactamase. In those mice, the researchers found that the amount of ampicillin circulating the bloodstream was as high as that in mice who did not receive the engineered bacteria. In the gut, mice that received engineered bacteria maintained a much higher level of microbial diversity compared to mice that received only antibiotics. In those mice, microbial diversity levels dropped dramatically after they received ampicillin. Furthermore, none of the mice that received the engineered bacteria developed opportunistic C. difficile infections, while all of the mice who received only antibiotics showed high levels of C. difficile in the gut.
“This is a strong demonstration that this approach can protect the gut microbiota, while preserving the efficacy of the antibiotic, as you’re not modifying the levels in the bloodstream,” Cubillos-Ruiz says.
The researchers also found that eliminating the evolutionary pressure of antibiotic treatment made it much less likely for the microbes of the gut to develop antibiotic resistance after treatment. In contrast, they did find many genes for antibiotic resistance in the microbes that survived in mice who received antibiotics but not the engineered bacteria. Those genes can be passed to harmful bacteria, worsening the problem of antibiotic resistance. The researchers now plan to begin developing a version of the treatment that could be tested in people at high risk of developing acute diseases that stem from antibiotic-induced gut dysbiosis, and they hope that eventually, it could be used to protect anyone who needs to take antibiotics for infections outside the gut.
“If the antibiotic action is not needed in the gut, then you need to protect the microbiota. This is similar to when you get an X-ray, you wear a lead apron to protect the rest of your body from the ionizing radiation,” Cubillos-Ruiz says. “No previous intervention could offer this level of protection. With our new technology we can make antibiotics safer by preserving beneficial gut microbes and by reducing the chances of emergence of new antibiotic resistant variants.”
Somatic mutation rates scale with lifespan across mammals
by Alex Cagan, Adrian Baez-Ortega, Natalia Brzozowska, et al. in Nature
The first study to compare the accumulation of mutations across many animal species has shed new light on decades-old questions about the role of these genetic changes in ageing and cancer. Researchers from the Wellcome Sanger Institute found that despite huge variation in lifespan and size, different animal species end their natural life with similar numbers of genetic changes.
The study analysed genomes from 16 species of mammal, from mice to giraffes. The authors confirmed that the longer the lifespan of a species, the slower the rate at which mutations occur, lending support to the long-standing theory that somatic mutations play a role in ageing. Genetic changes, known as somatic mutations, occur in all cells throughout the life of an organism. This is a natural process, with cells acquiring around 20 to 50 mutations per year in humans. Most of these mutations will be harmless, but some of them can start a cell on the path to cancer or impair the normal functioning of the cell.
Since the 1950s, some scientists have speculated that these mutations may play a role in ageing. But the difficulty of observing somatic mutations has made it challenging to study this possibility. In the last few years, technological advances have finally allowed genetic changes to be observed in normal tissues, raising hopes of answering this question. Another long-standing question is Peto’s paradox. Since cancers develop from single cells, species with larger bodies (and therefore more cells) should theoretically have a much higher risk of cancer. Yet cancer incidence across animals is independent of body size. Animal species with large bodies are believed to have evolved superior mechanisms to prevent cancer. Whether one such mechanism is a reduction in the accumulation of genetic changes in their tissues has remained untested.
In this study, researchers at the Wellcome Sanger Institute set out to test these theories by using new methods to measure somatic mutation in 16 mammalian species, covering a wide range of lifespans and body masses. This included species such as human, mouse, lion, giraffe, tiger, and the long-lived, highly cancer-resistant naked mole-rat, with samples provided by a number of organisations including the Zoological Society of London. Whole-genome sequences were generated from 208 intestinal crypts taken from 48 individuals, to measure mutation rates in single intestinal stem cells.
Analysis of the patterns of mutations (or mutational signatures) provided information on the processes at work. The researchers found that somatic mutations accumulated linearly over time and that they were caused by similar mechanisms across all species, including humans, despite their very different diets and life histories.Evidence of a possible role of somatic mutations in ageing was provided by the researchers’ discovery that the rate of somatic mutation decreased as the lifespan of each species increased.
Dr Alex Cagan, a first author of the study from the Wellcome Sanger Institute, said: “To find a similar pattern of genetic changes in animals as different from one another as a mouse and a tiger was surprising. But the most exciting aspect of the study has to be finding that lifespan is inversely proportional to the somatic mutation rate. This suggests that somatic mutations may play a role in ageing, although alternative explanations may be possible. Over the next few years, it will be fascinating to extend these studies into even more diverse species, such as insects or plants.” The search for an answer to Peto’s paradox goes on, however. After accounting for lifespan, the authors found no significant association between somatic mutation rate and body mass, indicating that other factors must be involved in larger animals’ ability to reduce their cancer risk relative to their size.
Dr Adrian Baez-Ortega, a first author of the study from the Wellcome Sanger Institute, said: “The fact that differences in somatic mutation rate seem to be explained by differences in lifespan, rather than body size, suggests that although adjusting the mutation rate sounds like an elegant way of controlling the incidence of cancer across species, evolution has not actually chosen this path. It is quite possible that every time a species evolves a larger size than its ancestors — as in giraffes, elephants and whales — evolution might come up with a different solution to this problem. We will need to study these species in greater detail to find out.”
Despite vast differences in lifespan and body mass between the 16 species studied, the quantity of somatic mutations acquired over each animal’s lifetime was relatively similar. On average a giraffe is 40,000 times bigger than a mouse, and a human lives 30 times longer, but the difference in the number of somatic mutations per cell at the end of lifespan between the three species only varied by around a factor of three.
Dr Simon Spiro, ZSL (Zoological Society of London) wildlife veterinary pathologist, said: “Animals often live much longer in zoos than they do in the wild, so our vets’ time is often spent dealing with conditions related to old age. The genetic changes identified in this study suggest that diseases of old age will be similar across a wide range of mammals, whether old age begins at seven months or 70 years, and will help us keep these animals happy and healthy in their later years.”
Understanding the exact causes of ageing remains an unsolved question and an area of active investigation. Ageing is likely to be caused by the accumulation of multiple types of damage to our cells and tissues throughout life, including somatic mutations, protein aggregation and epigenetic changes, among others. Comparing the rates of these processes across species with very different lifespans can shed light on their role in ageing.
Dr Inigo Martincorena, senior author of the study from the Wellcome Sanger Institute, said: “Ageing is a complex process, the result of multiple forms of molecular damage in our cells and tissues. Somatic mutations have been speculated to contribute to ageing since the 1950s, but studying them had remained difficult. With the recent advances in DNA sequencing technologies, we can finally investigate the roles that somatic mutations play in ageing and in multiple diseases. That this diverse range of mammals end their lives with a similar number of mutations in their cells is an exciting and intriguing discovery.”
Elucidating Design Principles for Engineering Cell‐Derived Vesicles to Inhibit SARS‐CoV‐2 Infection
by Taylor F. Gunnels, Devin M. Stranford, Roxana E. Mitrut, Neha P. Kamat, Joshua N. Leonard in Small
They might look like cells and act like cells. But a new potential COVID-19 treatment is actually a cleverly disguised trickster, which attracts viruses and binds them, rendering them inactive.
As the ever-evolving SARS-CoV-2 virus begins to evade once promising treatments, such as monoclonal antibody therapies, researchers have become more interested in these “decoy” nanoparticles. Mimicking regular cells, decoy nanoparticles soak up viruses like a sponge, inhibiting them from infecting the rest of the body.
In a new study, Northwestern University synthetic biologists set out to elucidate the design rules needed make decoy nanoparticles effective and resistant to viral escape. After designing and testing various iterations, the researchers identified a broad set of decoys — all manufacturable using different methods — that were incredibly effective against the original virus as well as mutant variants. In fact, decoy nanoparticles were up to 50 times more effective at inhibiting naturally occurring viral mutants, compared to traditional, protein-based inhibitor drugs. When tested against a viral mutant designed to resist such treatments, decoy nanoparticles were up to 1,500 times more effective at inhibiting infection. Although much more research and clinical evaluations are needed, the researchers believe decoy nanoparticle infusions someday could potentially be used to treat patients with severe or prolonged viral infections.
In the paper, the team tested decoy nanoparticles against the parent SARS-CoV-2 virus and five variants (including beta, delta, delta-plus and lambda) in a cellular culture.
“We showed that decoy nanoparticles are effective inhibitors of all these different viral variants,” said Northwestern’s Joshua Leonard, co-senior author of the study. “Even variants that escape other drugs did not escape our decoy nanoparticles.”
“As we were conducting the study, different variants kept popping up around the world,” added Northwestern’s Neha Kamat, co-senior author of the study. “We kept testing our decoys against the new variants, and they just kept working. It’s very effective.”
Leonard is an associate professor of chemical and biological engineering in Northwestern’s McCormick School of Engineering. Kamat is an assistant professor of biomedical engineering in McCormick. Both are key members of Northwestern’s Center for Synthetic Biology.
As the SARS-CoV-2 virus has mutated to create new variants, some treatments have become less effective in fighting the ever-evolving virus. Just last month, the U.S. Food and Drug Administration (FDA) paused several monoclonal antibody treatments, for example, due to their failure against the BA.2 omicron subvariant. But even where treatments fail, the decoy nanoparticles in the new study never lost effectiveness. Leonard said this is because the decoys put SARS-CoV-2 “between an evolutionary rock and a hard place.”
SARS-CoV-2 infects human cells by binding its infamous spike protein to the human angiotensin-converting enzyme 2 (ACE2) receptor. A protein on the surface of cells, ACE2 provides an entry point for the virus. To design decoy nanoparticles, the Northwestern team used nanosized particles (extracellular vesicles) naturally released from all cell types. They engineered cells producing these particles to overexpress the gene for ACE2, leading to many ACE2 receptors on the particles’ surfaces. When the virus came into contact with the decoy, it bonded tightly to these receptors rather than to real cells, rendering the virus unable to infect cells.
“For the virus to get into a cell, it has to bind to the ACE2 receptor,” Leonard said. “Decoy nanoparticles present an evolutionary challenge for SARS-CoV-2. The virus would have to come up with an entirely different way to enter cells in order to avoid the need to use ACE2 receptors. There is no obvious evolutionary escape route.”
In addition to being effective against drug-resistant viruses, decoy nanoparticles come with several other benefits. Because they are biological (rather than synthetic) materials, the nanoparticles are less likely to elicit an immune response, which causes inflammation and can interfere with the drug’s efficacy. They also exhibit low toxicity, making them particularly well-suited for use in sustained or repeated administration for treating severely ill patients. When the COVID-19 pandemic began, researchers and clinicians experienced an unnerving gap between discovering the virus and developing new drugs to treat it. For the next pandemic, decoy nanoparticles could provide a quick, effective treatment before vaccines are developed.
“The decoy strategy is one of the most immediate things you can try,” Leonard said. “As soon as you know the receptor that the virus uses, you can start building decoy particles with those receptors. We could potentially fast-track an approach like this to reduce severe illness and death in the crucial early stages of future viral pandemics.”
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