New compound allows bacterial communication to be controlled by light

Scientists from the University of Groningen have succeeded in incorporating a light-controlled switch into a molecule used by bacteria for quorum sensing -- a process by which bacteria communicate and subsequently control different cellular processes. With the molecule described, it is possible to either inhibit or stimulate communication. This makes it a very useful tool for further research into bacterial communication and its influence on different genetic pathways. The results were published on 15 April in the journal Chem.

 

Irradiation setup to switch the photoswitchable modulator of bacterial communication from the trans-isomer to the more active cis-isomer.

Credit: Dusan Kolarski, University of Groningen

 

In order to respond to their environment, bacteria 'talk' to each other through a form of chemical communication called quorum sensing. The cells secrete a signal molecule and at the same time monitor its concentration. As more cells secrete the signal molecule, it can exceed a threshold concentration and activate certain genetic pathways, for example, to produce toxins or form a protective biofilm.
Light-sensitive switch
'If we would be able to influence quorum sensing, we might be able to use it to treat serious infections,' says University of Groningen organic chemist Mickel Hansen. 'And it would also be useful to investigate how quorum sensing exactly works.' To do this, it would be useful to have a modulator of quorum sensing that could be externally controlled. That is why Hansen and colleagues in the synthetic organic chemistry group led by Professor Ben Feringa set out to build a light-sensitive switch into a molecule used by bacteria as a signal for quorum sensing.
The molecule is made up of a head and a flexible carbon-based tail, connected via a β-keto-amide linker. The plan was to incorporate a switch into the tail. 'This meant we had to connect the modified tail to the head via β-keto-amide linkage. However, the synthetic process to obtain this linkage produces a very unstable intermediate, which made it almost impossible to synthesize the molecule.'
Library
Building on the extensive experience of the synthetic organic chemistry group at the Stratingh Institute of Chemistry at the University of Groningen, the researchers came up with a solution in the form of a new coupling reaction with a stabilized intermediate. Using this intermediate, they were able to synthesize photoswitchable derivatives in a fast and straightforward way.
Hansen, together with Master's student Jacques Hille, produced a 'library' of 16 different compounds that had the potential to act as agonists or antagonists of quorum sensing. All were fitted with a light-operated switch. All compounds were based on a molecule that is used in one particular quorum sensing system in Pseudomonas aeruginosa, which has about five of these quorum sensing systems. In collaboration with molecular biologists from the lab of Professor of Molecular Microbiology Arnold Driessen, also at the University of Groningen, the genes for one of these systems were transferred to an E. coli reporter strain, allowing any effect of the newly synthesized compounds to be tested without the interference of other quorum sensing mechanisms.
Toxin production
Bioactivity tests on the compounds obtained showed which parts of the molecule were crucial to controlling quorum sensing. The optimum number of carbon atoms making up the tail appeared to be four. Flipping the switch with light caused the tail to bend. Remarkably, the straight tail had no effect, whereas the bent tail induced the quorum sensing signal. Hansen: 'Overall, it appears that small changes in the molecule can have a large effect on its activity, but we don't yet know exactly why.'
They did find one compound that was able to strongly inhibit the quorum sensing signal and -- after irradiation with light, leading to the bending of the tail -- to also strongly stimulate it. The difference in activity was more than 700-fold, which is huge. 'Such a large difference has, to our knowledge, never been shown before for light-switched bioactive molecules.' This particular molecule will be a very useful tool for investigating how bacteria communicate. 'In the study, we showed that we could light-control toxin production in a Pseudomonas strain with our switchable modulator. This will be a powerful tool for both clinical and fundamental research into the mechanism of quorum sensing.'
 

Journal Reference:

1. Mickel J. Hansen, Jacques I.C. Hille, Wiktor Szymanski, Arnold J.M. Driessen, Ben L. Feringa. Easily Accessible, Highly Potent, Photocontrolled Modulators of Bacterial Communication. Chem, 2019; DOI: 10.1016/j.chempr.2019.03.005 

Courtesy: ScienceDaily
 

Engineers tap DNA to create 'lifelike' machines

Tapping into the unique nature of DNA, Cornell engineers have created simple machines constructed of biomaterials with properties of living things.

 

Cornell professor of biological and environmental engineering Dan Luo and research associate Shogo Hamada have created a DNA material capable of metabolism, in addition to self-assembly and organization.

Credit: John Munson/Cornell University

Using what they call DASH (DNA-based Assembly and Synthesis of Hierarchical) materials, engineers constructed a DNA material with capabilities of metabolism, in addition to self-assembly and organization -- three key traits of life.
"We are introducing a brand-new, lifelike material concept powered by its very own artificial metabolism. We are not making something that's alive, but we are creating materials that are much more lifelike than have ever been seen before," said Dan Luo, professor of biological and environmental engineering.
The paper published in Science Robotics.
For any living organism to maintain itself, there must be a system to manage change. New cells must be generated; old cells and waste must be swept away. Biosynthesis and biodegradation are key elements of self-sustainability and require metabolism to maintain its form and functions.
Through this system, DNA molecules are synthesized and assembled into patterns in a hierarchical way, resulting in something that can perpetuate a dynamic, autonomous process of growth and decay.
Using DASH, the Cornell engineers created a biomaterial that can autonomously emerge from its nanoscale building blocks and arrange itself -- first into polymers and eventually mesoscale shapes. Starting from a 55-nucleotide base seed sequence, the DNA molecules were multiplied hundreds of thousands times, creating chains of repeating DNA a few millimeters in size. The reaction solution was then injected in a microfluidic device that provided a liquid flow of energy and the necessary building blocks for biosynthesis.
As the flow washed over the material, the DNA synthesized its own new strands, with the front end of the material growing and the tail end degrading in optimized balance. In this way, it made its own locomotion, creeping forward, against the flow, in a way similar to how slime molds move.
The locomotive ability allowed the researchers to pit sets of the material against one another in competitive races. Due to randomness in the environment, one body would eventually gain an advantage over the other, allowing one to cross a finish line first.
"The designs are still primitive, but they showed a new route to create dynamic machines from biomolecules. We are at a first step of building lifelike robots by artificial metabolism," said Shogo Hamada, lecturer and research associate in the Luo lab, and lead and co-corresponding author of the paper. "Even from a simple design, we were able to create sophisticated behaviors like racing. Artificial metabolism could open a new frontier in robotics."
 

Journal Reference:

1. Shogo Hamada, Kenneth Gene Yancey, Yehudah Pardo, Mingzhe Gan, Max Vanatta, Duo An, Yue Hu, Thomas L. Derrien, Roanna Ruiz, Peifeng Liu, Jenny Sabin, Dan Luo. Dynamic DNA material with emergent locomotion behavior powered by artificial metabolism. Science Robotics, 2019; 4 (29): eaaw3512 DOI: 10.1126/scirobotics.aaw3512 

Courtesy: ScienceDaily
 

Scientists print first 3D heart using patient's biological materials

In a major medical breakthrough, Tel Aviv University researchers have "printed" the world's first 3D vascularised engineered heart using a patient's own cells and biological materials. Their findings were published on April 15 in a study in Advanced Science.

 

A 3D-printed, small-scaled human heart engineered from the patient's own materials and cells.

Credit: Advanced Science. © 2019 The Authors.

 

Until now, scientists in regenerative medicine -- a field positioned at the crossroads of biology and technology -- have been successful in printing only simple tissues without blood vessels.

"This is the first time anyone anywhere has successfully engineered and printed an entire heart replete with cells, blood vessels, ventricles and chambers," says Prof. Tal Dvir of TAU's School of Molecular Cell Biology and Biotechnology, Department of Materials Science and Engineering, Center for Nanoscience and Nanotechnology and Sagol Center for Regenerative Biotechnology, who led the research for the study.

Heart disease is the leading cause of death among both men and women in the United States. Heart transplantation is currently the only treatment available to patients with end-stage heart failure. Given the dire shortage of heart donors, the need to develop new approaches to regenerate the diseased heart is urgent.

"This heart is made from human cells and patient-specific biological materials. In our process these materials serve as the bioinks, substances made of sugars and proteins that can be used for 3D printing of complex tissue models," Prof. Dvir says. "People have managed to 3D-print the structure of a heart in the past, but not with cells or with blood vessels. Our results demonstrate the potential of our approach for engineering personalized tissue and organ replacement in the future."

Research for the study was conducted jointly by Prof. Dvir, Dr. Assaf Shapira of TAU's Faculty of Life Sciences and Nadav Moor, a doctoral student in Prof. Dvir's lab.

"At this stage, our 3D heart is small, the size of a rabbit's heart," explains Prof. Dvir. "But larger human hearts require the same technology."

For the research, a biopsy of fatty tissue was taken from patients. The cellular and a-cellular materials of the tissue were then separated. While the cells were reprogrammed to become pluripotent stem cells, the extracellular matrix (ECM), a three-dimensional network of extracellular macromolecules such as collagen and glycoproteins, were processed into a personalized hydrogel that served as the printing "ink."

After being mixed with the hydrogel, the cells were efficiently differentiated to cardiac or endothelial cells to create patient-specific, immune-compatible cardiac patches with blood vessels and, subsequently, an entire heart.

According to Prof. Dvir, the use of "native" patient-specific materials is crucial to successfully engineering tissues and organs.

"The biocompatibility of engineered materials is crucial to eliminating the risk of implant rejection, which jeopardizes the success of such treatments," Prof. Dvir says. "Ideally, the biomaterial should possess the same biochemical, mechanical and topographical properties of the patient's own tissues. Here, we can report a simple approach to 3D-printed thick, vascularized and perfusable cardiac tissues that completely match the immunological, cellular, biochemical and anatomical properties of the patient."

The researchers are now planning on culturing the printed hearts in the lab and "teaching them to behave" like hearts, Prof. Dvir says. They then plan to transplant the 3D-printed heart in animal models.

"We need to develop the printed heart further," he concludes. "The cells need to form a pumping ability; they can currently contract, but we need them to work together. Our hope is that we will succeed and prove our method's efficacy and usefulness.

"Maybe, in ten years, there will be organ printers in the finest hospitals around the world, and these procedures will be conducted routinely."

Journal Reference:

1. Nadav Noor, Assaf Shapira, Reuven Edri, Idan Gal, Lior Wertheim, Tal Dvir. 3D Printing of Personalized Thick and Perfusable Cardiac Patches and Hearts. Advanced Science, 2019; 1900344 DOI: 10.1002/advs.201900344 

Courtesy: 

ScenceDaily

Biophysicists use machine learning to understand, predict dynamics of worm behavior

Caenorhabditis elegans (stock image).

Credit: © heitipaves / Fotolia

Biophysicists have used an automated method to model a living system -- the dynamics of a worm perceiving and escaping pain. The Proceedings of the National Academy of Sciences (PNAS) published the results, which worked with data from experiments on the C. elegans roundworm.

"Our method is one of the first to use machine-learning tools on experimental data to derive simple, interpretable equations of motion for a living system," says Ilya Nemenman, senior author of the paper and a professor of physics and biology at Emory University. "We now have proof of principle that it can be done. The next step is to see if we can apply our method to a more complicated system."

The model makes accurate predictions about the dynamics of the worm behavior, and these predictions are biologically interpretable and have been experimentally verified.

Collaborators on the paper include first author Bryan Daniels, a theorist from Arizona State University, and co-author William Ryu, an experimentalist from the University of Toronto.

The researchers used an algorithm, developed in 2015 by Daniels and Nemenman, that teaches a computer how to efficiently search for the laws that underlie natural dynamical systems, including complex biological ones. They dubbed the algorithm "Sir Isaac," after one of the most famous scientists of all time -- Sir Isaac Newton. Their long-term goal is to develop the algorithm into a "robot scientist," to automate and speed up the scientific method of forming quantitative hypotheses, then testing them by looking at data and experiments.

While Newton's Three Laws of Motion can be used to predict dynamics for mechanical systems, the biophysicists want to develop similar predictive dynamical approaches that can be applied to living systems.

For the PNAS paper, they focused on the decision-making involved when C. elegans responds to a sensory stimulus. The data on C. elegans had been previously gathered by the Ryu lab, which develops methods to measure and analyze behavioral responses of the roundworm at the holistic level, from basic motor gestures to long-term behavioral programs.

C. elegans is a well-established laboratory animal model system. Most C. elegans have only 302 neurons, few muscles and a limited repertoire of motion. A sequence of experiments involved interrupting the forward movement of individual C. elegans with a laser strike to the head. When the laser strikes a worm, it withdraws, briefly accelerating backwards and eventually returning to forward motion, usually in a different direction. Individual worms respond differently. Some, for instance, immediately reverse direction upon laser stimulus, while others pause briefly before responding. Another variable in the experiments is the intensity of the laser: Worms respond faster to hotter and more rapidly rising temperatures.

The researchers fed the Sir Isaac platform the motion data from the first few seconds of the experiments -- before and shortly after the laser strikes a worm and it initially reacts. From this limited data, the algorithm was able to capture the average responses that matched the experimental results and also to predict the motion of the worm well beyond these initial few seconds, generalizing from the limited knowledge. The prediction left only 10 percent of the variability in the worm motion that can be attributed to the laser stimulus unexplained. This was twice as good as the best prior models, which were not aided by automated inference.

"Predicting a worm's decision about when and how to move in response to a stimulus is a lot more complicated than just calculating how a ball will move when you kick it," Nemenman says. "Our algorithm had to account for the complexities of sensory processing in the worms, the neural activity in response to the stimuli, followed by the activation of muscles and the forces that the activated muscles generate. It summed all this up into a simple and elegant mathematical description."

The model derived by Sir Isaac was well-matched to the biology of C. elegans, providing interpretable results for both the sensory processing and the motor response, hinting at the potential of artificial intelligence to aid in discovery of accurate and interpretable models of more complex systems.

"It's a big step from making predictions about the behavior of a worm to that of a human," Nemenman says, "but we hope that the worm can serve as a kind of sandbox for testing out methods of automated inference, such that Sir Isaac might one day directly benefit human health. Much of science is about guessing the laws that govern natural systems and then verifying those guesses through experiments. If we can figure out how to use modern machine learning tools to help with the guessing, that could greatly speed up research breakthroughs."

Journal Reference:

1. Bryan C. Daniels, William S. Ryu, Ilya Nemenman. Automated, predictive, and interpretable inference of Caenorhabditis elegans escape dynamics. Proceedings of the National Academy of Sciences, 2019; 201816531 DOI: 10.1073/pnas.1816531116 

Courtesy: ScienceDaily

Seeds inherit memories from their mother

 

This is a seed of Arabidopsis thaliana at the beginning of germination.

Credit: © UNIGE / Sylvain Loubéry

Seeds remain in a dormant state -- a temporary blockage of their germination -- as long as environmental conditions are not ideal for germination. The depth of this sleep, which is influenced by various factors, is inherited from their mother, as researchers from the University of Geneva (UNIGE), Switzerland, had previously shown. Today, they reveal in the journal eLife how this maternal imprint is transmitted through small fragments of so-called 'interfering' RNAs, which inactivate certain genes. The biologists also reveal that a similar mechanism enables to transmit another imprint, that of the temperatures present during the development of the seed. The lower this temperature was, the higher the seed's dormancy level will be. This mechanism allows the seed to optimize the timing of its germination. The information is then erased in the germinated embryo, so that the next generation can store new data on its environment.

Dormancy is implemented during seed development in the mother plant. This property allows the seeds to germinate during the appropriate season, to prevent all the offspring of a plant from developing in the same place and competing for limited resources, and to promote plant dispersal. Seeds also lose their dormancy at variable times. "Subspecies of the same plant can have different levels of dormancy depending on the latitudes at which they are produced, and we wanted to understand why," explains Luis Lopez-Molina, Professor at the Department of Botany and Plant Biology of the UNIGE Faculty of Science.

The paternal gene is silenced

Like all organisms with sexual reproduction, the seed receives two versions of each gene, a maternal and a paternal allele, which may have different levels of expression. The UNIGE biologists had shown in 2016 that the dormancy levels of Arabidopsis thaliana, a model organism used in laboratories, are inherited from the mother. Indeed, in the seed, the level of expression of a dormancy regulating gene called allantoinase (ALN) is the same as that of the maternal allele. This implies that it is the maternal allele of ALN that is mainly expressed, to the detriment of the paternal allele.

In the current study, the researchers show that this maternal imprint is transmitted by an epigenetic mechanism, which influences the expression of certain genes without altering their sequence. The paternal allele of ALN is 'silenced' by biochemical modifications called methylations, which are carried out in the promoter region of the gene in order to inactivate it.

"These methylations are themselves the result of a process in which different enzymatic and factor complexes are involved, as well as small fragments of so-called 'interfering' RNA. This is a unique example of genomic imprinting, because it is made in the absence of the enzyme usually responsible for methylation," says Mayumi Iwasaki, researcher in the Geneva group and the first author of the article.

The imprint of past cold prevents the seed from awakening

The environmental conditions present during the seed formation also leave their mark, as its dormancy level increases with decreasing temperatures. "We have discovered that, in this case, both alleles of the ALN gene are strongly repressed in the seed. This is due to a similar epigenetic mechanism, but not all of the actors are the same as those used to silence the paternal allele," says Luis Lopez-Molina.

This imprint of the cold enables the seed to keep information on past temperatures, in order to include them in the choice of the optimal time of germination. After germination, the ALN gene is reactivated in the embryo. The memory of the cold will then be cleared, allowing the counters to be reset for the next generation.

"Studying how maternal and environmental factors cause dormant seeds to awaken is of crucial importance for agriculture, especially to prevent early germination in an environment subject to climate change," concludes Mayumi Iwasaki. The ecological stakes are also high, because increasing temperatures could reduce the dormancy of the seed bank and thus modify the distribution of plant species under a given latitude. This would have multiple consequences, both direct and indirect, for native animal and plant species.

Journal Reference:

1. Mayumi Iwasaki, Lena Hyvärinen, Urszula Piskurewicz, Luis Lopez-Molina. Non-canonical RNA-directed DNA methylation participates in maternal and environmental control of seed dormancy. eLife, 2019; 8 DOI: 10.7554/eLife.37434

 Courtesy: ScienceDaily

Woman with novel gene mutation lives almost pain-free

A woman in Scotland can feel virtually no pain due to a mutation in a previously-unidentified gene, according to a research paper co-led by UCL.

She also experiences very little anxiety and fear, and may have enhanced wound healing due to the mutation, which the researchers say could help guide new treatments for a range of conditions, they report in the British Journal of Anaesthesia.

"We found this woman has a particular genotype that reduces activity of a gene already considered to be a possible target for pain and anxiety treatments," said one of the study's lead researchers, Dr James Cox (UCL Medicine).

"Now that we are uncovering how this newly-identified gene works, we hope to make further progress on new treatment targets."

At age 65, the woman sought treatment for an issue with her hip, which turned out to involve severe joint degeneration despite her experiencing no pain. At age 66, she underwent surgery on her hand, which is normally very painful, and yet she reported no pain after the surgery. Her pain insensitivity was diagnosed by Dr Devjit Srivastava, Consultant in Anaesthesia and Pain Medicine at an NHS hospital in the north of Scotland and co-lead author of the paper.

The woman tells researchers she has never needed painkillers after surgery such as dental procedures.

She was referred to pain geneticists at UCL and the University of Oxford, who conducted genetic analyses and found two notable mutations. One was a microdeletion in a pseudogene, previously only briefly annotated in medical literature, which the researchers have described for the first time and dubbed FAAH-OUT. She also had a mutation in the neighbouring gene that controls the FAAH enzyme.

Further tests by collaborators at the University of Calgary, Canada, revealed elevated blood levels of neurotransmitters that are normally degraded by FAAH, further evidence for a loss of FAAH function.

The FAAH gene is well-known to pain researchers, as it is involved in endocannabinoid signalling central to pain sensation, mood and memory. The gene now called FAAH-OUT was previously assumed to be a 'junk' gene that was not functional. The researchers found there was more to it than previously believed, as it likely mediates FAAH expression.

Mice that do not have the FAAH gene have reduced pain sensation, accelerated wound healing, enhanced fear-extinction memory and reduced anxiety.

The woman in Scotland experiences similar traits. She notes that in her lifelong history of cuts and burns (sometimes unnoticed until she can smell burning flesh), the injuries tend to heal very quickly. She is an optimist who was given the lowest score on a common anxiety scale, and reports never panicking even in dangerous situations such as a recent traffic incident. She also reports memory lapses throughout life such as forgetting words or keys, which has previously been associated with enhanced endocannabinoid signalling.

The researchers say that it's possible there are more people with the same mutation, given that this woman was unaware of her condition until her 60s.

"People with rare insensitivity to pain can be valuable to medical research as we learn how their genetic mutations impact how they experience pain, so we would encourage anyone who does not experience pain to come forward," said Dr Cox.

The research team is continuing to work with the woman in Scotland, and are conducting further tests in cell samples, in order to better understand the novel pseudogene.

"We hope that with time, our findings might contribute to clinical research for post-operative pain and anxiety, and potentially chronic pain, PTSD and wound healing, perhaps involving gene therapy techniques," said Dr Cox.

"The implications for these findings are immense," said Dr Srivastava.

"One out of two patients after surgery today still experiences moderate to severe pain, despite all advances in pain killer medications and techniques since the use of ether in 1846 to first 'annul' the pain of surgery. There have already been unsuccessful clinical trials targeting the FAAH protein -- while we hope the FAAH-OUT gene could change things particularly for post-surgical pain, it remains to be seen if any new treatments could be developed based on our findings."

"The findings point towards a novel pain killer discovery that could potentially offer post-surgical pain relief and also accelerate wound healing. We hope this could help the 330 million patients who undergo surgery globally every year," Dr Srivastava said.

"I would be elated if any research into my own genetics could help other people who are suffering," the woman in Scotland commented.

"I had no idea until a few years ago that there was anything that unusual about how little pain I feel -- I just thought it was normal. Learning about it now fascinates me as much as it does anyone else."

Lead funding for the study came from the Medical Research Council and Wellcome.

Journal Reference:

1. Abdella M. Habib, Andrei L. Okorokov, Matthew N. Hill, Jose T. Bras, Man-Cheung Lee, Shengnan Li, Samuel J. Gossage, Marie van Drimmelen, Maria Morena, Henry Houlden, Juan D. Ramirez, David L.H. Bennett, Devjit Srivastava, James J. Cox. Microdeletion in a pseudogene identified in a patient with high anandamide concentrations and pain insensitivity. British Journal of Anaesthesia, 2019; DOI: 10.1016/j.bja.2019.02.019

 Courtesy: ScienceDaily