GN/ Researchers develop first method to study microRNA activity in single cells
Genetics biweekly vol.45, 20th September — 6th October
TL;DR
- Researchers have developed the first method to uncover the tasks that microRNAs perform in single cells. This is a huge improvement over existing state-of-the-art methods that require millions of cells and will for the first time allow researchers to study microRNAs in complex tissues such as brains.
- To make a gene-editing tool more precise and easier to control, engineers split it into two pieces that only come back together when a third molecule is added.
- Researchers have developed a novel genome editing technique known as NICER, which results in significantly fewer off-target mutations than CRISPR/Cas9 editing. The technique uses a different type of enzyme that makes single-stranded ‘nicks’ in the DNA. Repair of these nicks is more efficient and accurate than repair of double-strand breaks caused by the current CRISPR/Cas9 editing. This technique represents a novel approach for the treatment of genetic diseases caused by heterozygous mutations.
- Scientists have made a significant finding in determining the genetic background of dilated cardiomyopathy in Dobermanns. This research helps us understand the genetic risk factors related to fatal diseases of the heart muscle and the mechanisms underlying the disease, and offers new tools for their prevention.
- Living things from plants to humans must constantly adjust the chemical soup of proteins — the workhorse molecules of life — inside their cells to adapt to stress or changing conditions. Now, researchers have identified a previously unknown molecular mechanism that helps explain how they do it. A team now reveals hairpin-like structures of mRNA that, by zipping and unzipping, help cells change the mix of proteins they produce when under stress.
- Researchers have developed a method that lets them genetically modify each cell differently in animals. This allows them to study in a single experiment what used to require many animal experiments. Using the new method, the researchers have discovered genes that are relevant for a severe rare genetic disorder.
- Biologists have discovered why an enzyme is important for the survival of fruit flies, even though it can shorten their lives under certain conditions.
- A new study shows the isolation and sequencing of more than a century-old RNA molecules from a Tasmanian tiger specimen preserved at room temperature in a museum collection. This resulted in the reconstruction of skin and skeletal muscle transcriptomes from an extinct species for the first time. The researchers note that their findings have relevant implications for international efforts to resurrect extinct species, including both the Tasmanian tiger and the woolly mammoth, as well as for studying pandemic RNA viruses.
- Researchers have developed a way of detecting the early onset of deadly infectious diseases using a test so ultrasensitive that it could someday revolutionize medical approaches to epidemics. The test is an electronic sensor contained within a computer chip. It employs nanoballs — microscopic spherical clumps made of tinier particles of genetic material — and combines that technology with advanced electronics.
- A study sheds new light on the evolution of neurons, focusing on the placozoans, a millimetre-sized marine animal. Researchers find evidence that specialized secretory cells found in these unique and ancient creatures may have given rise to neurons in more complex animals.
- 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
Detection of transcriptome-wide microRNA–target interactions in single cells with agoTRIBE
by Vaishnovi Sekar, Emilio Mármol-Sánchez, Panagiotis Kalogeropoulos, Laura Stanicek, Eduardo A. Sagredo, Albin Widmark, Evangelos Doukoumopoulos, Franziska Bonath, Inna Biryukova, Marc R. Friedländer in Nature Biotechnology
Researchers at Stockholm University and SciLifeLab have developed the first method to uncover the tasks that microRNAs perform in single cells. This is a huge improvement over existing state-of-the-art methods that require millions of cells and will for the first time allow researchers to study microRNAs in complex tissues such as brains.
MicroRNAs are small molecules that regulate gene activity by binding to and destroying RNAs produced by the genes. More than 60% of all human genes are estimated to be regulated by microRNAs, therefore it is not surprising that these small molecules are involved in many biological processes including diseases such as cancer. To discover the function of a microRNA, it is necessary to find out exactly which RNAs are targeted by it. While such methods exist, they require a lot of material typically in order of millions of cells, to work.
Now researchers at Stockholm University and SciLifeLab have developed a new method to detect microRNA targets at the level of single cells. Such cells are each around one-hundredth millimeter in diameter and weigh less than a billionth gram, and comprise the basic building blocks of living organisms. With their new sensitive method, the researchers can follow microRNA targeting of thousands of RNAs during biological processes such as the cell cycle or differentiation into red blood cells. In these processes, the researchers find that microRNAs — surprisingly — perform quite different tasks in each cell. In the future, it will be possible to also apply this method to study microRNA targeting in whole tissues, to find out exactly what is happening in each of the many cell types that comprise complex organs such as brains.
Marc Friedländer, associate professor at Stockholm University, says: “In our research team, we want to understand and ultimately make mathematical models of gene regulation at the level of the single cell. Our new method is a huge leap towards making this possible.”
The work was spearheaded by Dr. Inna Biryukova, who took a leading role in developing the laboratory method, and by PhD student Vaishnovi Sekar, who performed the bulk of the advanced computational analyses. Vaishnovi Sekar highlights the challenges of the project: “In terms of complexity of the computational work, this is uncharted territory, and we lacked reference points and thresholds. We had to explore a myriad of approaches to devise a methodology that not only works but also yields biologically meaningful observations.”
A split and inducible adenine base editor for precise in vivo base editing
by Hongzhi Zeng, Qichen Yuan, Fei Peng, Dacheng Ma, Ananya Lingineni, Kelly Chee, Peretz Gilberd, Emmanuel C. Osikpa, Zheng Sun, Xue Gao in Nature Communications
To make a gene-editing tool more precise and easier to control, Rice University engineers split it into two pieces that only come back together when a third small molecule is added.
Researchers in the lab of chemical and biomolecular engineer Xue Sherry Gao created a CRISPR-based gene editor designed to target adenine ⎯ one of the four main DNA building blocks ⎯ that remains inactive when disassembled but kicks into gear once the binding molecule is added.
Compared to the intact original, the split editor is more precise and stays active for a narrower window of time, which is important for avoiding off-target edits. Moreover, the activating small molecule used to bind the two pieces of the tool together is already being used as an anticancer and immunosuppressive drug.
According to a study, the tool developed by Gao and collaborators performed well both in human cell cultures and in living mice, where it accurately edited a single base pair on a target gene. Given that single base-pair mutations ⎯ also known as point mutations ⎯ are responsible for thousands of diseases, the split editor could have broad therapeutic applications.
“This tool has the potential to correct nearly half of the disease-causing point mutations in our genome,” said Hongzhi Zeng, the lead author of the study and a graduate student in the Gao lab. “However, current adenine base editors are in a constant ‘on’ state, which could lead to unwanted genome changes alongside the desired correction in the host genome.
“Our team set out to create a much improved version that can be turned on or off as needed, providing an unparalleled level of safety and accuracy.”
To install an ‘on/off’ switch, the researchers broke the adenine base editor into two separate proteins that remain inactive until sirolimus (previously known as rapamycin) is added ⎯ a molecule discovered in 1972 in soil bacteria on Easter Island that is approved by the U.S. Food and Drug Administration for use in cancer therapies and other medical procedures.
“Upon introduction of this small molecule, the two separate inactive fragments of the adenine base editor are glued together and rendered active,” Zeng said. “As the body metabolizes the rapamycin, the two fragments disjoin, deactivating the system.”
The researchers found some additional benefits to splitting the gene editor in two.
“Compared to an intact editor, our version reduces overall off-target edits by over 70% and increases the accuracy of on-target edits,” Zeng said.
In collaboration with Zheng Sun, associate professor in the Department of Molecular and Cellular Biology and in the endocrinology, diabetes and metabolism division of the Department of Medicine at Baylor College of Medicine, researchers targeted the PCSK9 gene, which serves as the blueprint for a protein that helps regulate blood cholesterol levels.
“We hope to see the eventual application of our split genome-editing tool with higher precision to address human health-related questions in a much safer way,” said Gao, the Ted N. Law Assistant Professor of Chemical and Biomolecular Engineering.
Inducing multiple nicks promotes interhomolog homologous recombination to correct heterozygous mutations in somatic cells
by Akiko Tomita, Hiroyuki Sasanuma, Tomoo Owa, Yuka Nakazawa, Mayuko Shimada, Takahiro Fukuoka, Tomoo Ogi, Shinichiro Nakada in Nature Communications
The gene editing technique CRISPR/Cas9 has allowed researchers to make precise and impactful changes to an organism’s DNA to fix mutations that cause genetic disease. However, the CRISPR/Cas9 method can also result in unintended DNA mutations that may have negative effects. Recently, researchers in Japan have developed a new gene editing technique that is as effective as CRISPR/Cas9 while significantly reducing these unintended mutations.
In a new study, researchers led by Osaka University introduced a novel technique called NICER, which is based on the creation of multiple small cuts in single DNA strands by an enzyme called a nickase.
Traditional CRISPR/Cas9 editing uses small pieces of genetic code called guide RNAs and an enzyme called Cas9. The guide RNAs target a specific section of the DNA and the Cas9 enzyme initiates a break in the double-stranded DNA structure at this location. This double-strand break is key for initiating changes to the DNA. However, cellular repair of double-strand breaks can lead to unintended DNA mutations, as well as the integration of exogenous DNA to the human genome, which raises safety concerns for clinical applications of CRISPR/Cas9 technology. To minimize these unintended mutations, the Osaka University-led research team investigated the use of Cas9 nickase, which creates single-strand breaks or “nicks” in DNA that are typically repaired without causing mutations.
“Each chromosome in the genome has a ‘homologous’ copy,” says lead author of the study Akiko Tomita. “Using the NICER technique, heterozygous mutations — in which a mutation appears in one chromosome but not its homologous copy — are repaired using the unmutated homologous chromosome as a template.”
For their initial experiments, the research team used human lymphoblast cells with a known heterozygous mutation in a gene called TK1. When these cells were treated with nickase to induce a single cut in the TK1 region, TK1 activity was recovered at a low rate. However, when the nickase induced multiple nicks in this region on both homologous chromosomes, gene correction efficiency was enhanced approximately seventeen-fold via activation of a cellular repair mechanism.
“Further genomic analysis showed that the NICER technique rarely induced off-target mutations,” says senior author Shinichiro Nakada. “We were also pleased to find that NICER was able to restore the expression of disease-causing genes in cells derived from genetic diseases involving compound heterozygous mutations.”
Because the NICER method does not involve DNA double-strand breaks or the use of exogenous DNA, this technique appears to be a safe alternative to conventional CRISPR/Cas9 methods. NICER may represent a novel approach for the treatment of genetic diseases caused by heterozygous mutations.
Identification of novel genetic risk factors of dilated cardiomyopathy: from canine to human
by Julia E. Niskanen, Åsa Ohlsson, Ingrid Ljungvall, et al in Genome Medicine
Researchers have made a significant finding in determining the genetic background of dilated cardiomyopathy in Dobermanns. This research helps us understand the genetic risk factors related to fatal diseases of the heart muscle and the mechanisms underlying the disease, and offers new tools for their prevention.
Researchers from the University of Helsinki and the Folkhälsan Research Center, together with their international partners, have identified the genetic background of dilated cardiomyopathy, a disease that enlarges the heart muscle, in dogs and humans. Based on a dataset encompassing more than 500 Dobermanns, the disease was associated with two nearby genomic loci, where changes were identified in genes that affect the functioning, energy metabolism and structure of the heart muscle. The study revealed that these same risk genes cause heart muscle disease in human patients. A variety of factors can cause cardiomyopathy, but genetics play a significant role. Although dozens of genes underlying cardiomyopathy in humans have been identified, the hereditary nature and genetic background of the disease in dogs have remained unclear.
“The situation with Dobermanns is serious in terms of both their health and breeding. The disease has been studied from various angles for decades without significant gene discoveries. Better diagnostic tools are needed, particularly in early diagnostics. Our new research might improve the situation,” says Professor Hannes Lohi, the principal investigator in the project.
The study has significant implications for veterinary medicine, providing a basis for developing a new genetic test for early diagnostics and breeding. Various research data collected over decades on more than 500 Dobermanns from across Europe were combined for the research. The dogs in the study cohort were categorised into five different groups:
- Dogs with only dilated cardiomyopathy
- Dogs with only arrhythmia
- Dogs with dilated cardiomyopathy and arrhythmia
- Dogs with congestive heart failure
- Healthy dogs aged at least six years as a control subcohort
With the help of genetic mapping, two adjacent gene loci in chromosome 5 were associated with dilated cardiomyopathy. Among the numerous genes in the loci, two, namely RNF207 and PRKAA2, demonstrated structural variation, which could have a detrimental effect on the functioning of the genes and cause heart failure.
“The genetic mapping we conducted produced important observations. Until now, it has been unclear whether Dobermanns with differing symptoms have the same disease. The genes we identified are only associated with a dilated heart and affected cardiac function. Arrhythmia appears to be a genetically distinct disease. Our dataset was insufficient to identify genes causing arrhythmia only. We also observed that several genes affect cardiac function and identified a model of two genes that increase the disease risk,” explains Professor Lohi.
The significance of the gene discovery in dogs was investigated in human patients diagnosed with dilated cardiomyopathy using Dutch, English (UK Biobank) and Finnish (FinnGen) cohorts. Fifteen potentially harmful and predisposing variants in the same RNF207 and PRKAA2 genes, which had been identified in dogs, were discovered in humans.
“The identical genetic background suggests that, to a degree, similar problems with the functioning of the heart muscle lead to dilated cardiomyopathy in both humans and dogs. A deeper understanding of the pathogenetic mechanisms is important, and Dobermanns represent a natural model organism for further research,” Lohi states.
The DNA markers associated with the disease found in the study may be a step toward a genetic test, but it is important to confirm its clinical significance before such tests are offered.
“We discovered how the variants of the two genes together increase the disease risk. However, a pilot is needed to combine genetic and health data to monitor how frequently individuals who belong to the at-risk group develop the disease for varying genetic reasons. Then, we can obtain a more accurate estimate of how the gene discoveries should be ideally interpreted and utilised. In any case, this is a hope-inspiring finding because, in the past, we lacked such tools,” Lohi describes.
For the consistent synchronised pumping of the heart, the heart muscle cells must interact with each other. Unlike in skeletal muscles, in the cell membrane of the heart muscle are finger-like discs that conduct the undulation required for pumping.
“Our study revealed that the RNF207 gene is expressed exactly in these discs. Earlier research has shown that RNF207 plays an important role in heart muscle contraction. The absence of these discs has also earlier been linked with cardiomyopathy. The other gene identified, PKAA2, serves as an energy sensor in the heart muscle, and its malfunction can reduce cardiac efficiency. Further research is required to understand the pathogenic mechanism, but we are in a good position to continue. A while ago, the disease was a total mystery, but now we have opened a view to its cellular-level secrets,” Professor Lohi concludes.
Pervasive downstream RNA hairpins dynamically dictate start-codon selection
by Yezi Xiang, Wenze Huang, Lianmei Tan, Tianyuan Chen, Yang He, Patrick S. Irving, Kevin M. Weeks, Qiangfeng Cliff Zhang, Xinnian Dong in Nature
Living things from bacteria to plants to humans must constantly adjust the chemical soup of proteins — the workhorse molecules of life — inside their cells to adapt to stress or changing conditions, such as when nutrients are scarce, or when a pathogen attacks. Now, researchers have identified a previously unknown molecular mechanism that helps explain how they do it.
Studying a spindly plant called Arabidopsis thaliana, a Duke University-led team discovered short snippets of folded RNA that, under normal conditions, keep levels of defense proteins low to avoid harming the plants themselves. But when the plants detect a pathogen, these folded RNA structures are unzipped, enabling plant cells to make defense proteins to fight infection.
“It’s another tool in our toolkit” to control protein production, said Duke biology professor Xinnian Dong, senior author of the study.
In the soupy interior of every cell in the body, millions of protein molecules carry out the tasks of life: They are the cellular equivalent of bricks and beams, providing structure and support. They’re also the cell’s chemical messengers, sending and receiving signals. And they’re the defenders, deployed in response to foreign invaders.
To build a protein, sections of the DNA blueprint packed inside the cell’s nucleus are transcribed into messenger molecules called mRNA, which are instructions for making proteins. These instructions are carried out to the rest of the cell, where decoding devices called ribosomes translate the mRNA’s message to assemble a chain of amino acids, the building blocks of a protein.
Normally, ribosomes scan along the mRNA molecule until they find a special three-letter sequence that says, “start here to make a protein.” But in the new study, Dong and Yezi Xiang, a Ph.D. student in Dong’s Lab, found that, when an Arabidopsis seedling detects a potential pathogen, the plant’s ribosomes bypass the usual ‘start’ signal for protein synthesis and begin translating the mRNA further downstream, building a completely different chain of amino acids — and thus a different protein — required for fighting infection.
Dong and her team wanted to know: how do cells make the switch from one start site to another? To better understand this rapid cellular decision-making that takes place when a plant detects an invader, the researchers turned to a technique, called SHAPE-MaP, that allows them to detect changes in mRNA folding within living cells. Near the usual ‘green light’ that sets protein synthesis in motion, the researchers discovered short stretches of mRNA that fold back upon themselves to form double-stranded “hairpin” structures. Under normal conditions, these hairpins act as brakes, preventing ribosomes from making defense proteins whose instructions lie further downstream.
But when Arabidopsis seedlings sense they’re under attack, special enzymes called RNA helicases are produced that unzip the hairpins so the ribosomes can pass through and continue scanning along the mRNA molecule.
“With these stop signs removed, the ribosomes don’t stop there, but go further down to translate defense proteins,” Dong said.
Though the team did the bulk of their experiments in Arabidopsis plants, similar RNA helicases and hairpin structures have been found in other organisms, from yeast to humans, suggesting that this mechanism for reprogramming protein synthesis may be widespread. In follow-up experiments, the researchers used machine learning to come up with a design for a lab-made mRNA hairpin and added it to human genes. The synthetic hairpins worked to alter protein production in human cells, too.
The team has filed for a provisional patent on the discovery. Dong says the findings could lead to new ways to engineer crops that are “not only resistant to pathogens, but also to environmental stresses like heat, cold, and drought.” In the future, Dong said, it might also be possible to design mRNA hairpins for genome editing to help fight infections or treat diseases in people.
“The goal is to help cells produce the right amount of protein at the right time and the right place,” Dong said. “This is a step towards that goal.”
Transcriptional linkage analysis with in vivo AAV-Perturb-seq
by Antonio J. Santinha, Esther Klingler, Maria Kuhn, Rick Farouni, Sandra Lagler, Georgios Kalamakis, Ulrike Lischetti, Denis Jabaudon, Randall J. Platt in Nature
Researchers at ETH Zurich have developed a method that lets them genetically modify each cell differently in animals. This allows them to study in a single experiment what used to require many animal experiments. Using the new method, the researchers have discovered genes that are relevant for a severe rare genetic disorder.
One proven method for tracking down the genetic causes of diseases is to knock out a single gene in animals and study the consequences this has for the organism. The problem is that for many diseases, the pathology is determined by multiple genes. This makes it extremely difficult for scientists to determine the extent to which any one of the genes is involved in the disease. To do this, they would have to perform many animal experiments — one for each desired gene modification.
Researchers led by Randall Platt, Professor of Biological Engineering at the Department of Biosystems Science and Engineering at ETH Zurich in Basel, have now developed a method that will greatly simplify and speed up research with laboratory animals: using the CRISPR-Cas gene scissors, they simultaneously make several dozen gene changes in the cells of a single animal, much like a mosaic. While no more than one gene is altered in each cell, the various cells within an organ are altered in different ways. Individual cells can then be precisely analysed. This enables researchers to study the ramifications of many different gene changes in a single experiment.
For the first time, the ETH Zurich researchers have now successfully applied this approach in living animals — specifically, in adult mice — as they report in the current issue of Nature. Other scientists had previously developed a similar approach for cells in culture or animal embryos.
To “inform” the mice’s cells as to which genes the CRISPR-Cas gene scissors should destroy, the researchers used the adeno-associated virus (AAV), a delivery strategy that can target any organ. They prepared the viruses so that each virus particle carried the information to destroy a particular gene, then infected the mice with a mixture of viruses carrying different instructions for gene destruction. In this way, they were able to switch off different genes in the cells of one organ. For this study, they chose the brain.
Using this method, the researchers from ETH Zurich, together with colleagues from the University of Geneva, obtained new clues to a rare genetic disorder in humans, known as 22q11.2 deletion syndrome. Patients affected by the disease show many different symptoms, typically diagnosed with other conditions such as schizophrenia and autism spectrum disorder. Before now, it was known that a chromosomal region containing 106 genes is responsible for this disease. It was also known that the disease was associated with multiple genes, however, it was not known which of the genes played which part in the disease.
For their study in mice, the researchers focused on 29 genes of this chromosomal region that are also active in the mouse brain. In each individual mouse brain cell, they modified one of these 29 genes and then analysed the RNA profiles of those brain cells. The scientists were able to show that three of these genes are largely responsible for the dysfunction of brain cells. In addition, they found patterns in the mouse cells that are reminiscent of schizophrenia and autism spectrum disorders. Among the three genes, one was already known, but the other two had not previously been the focus of much scientific attention.
“If we know which genes in a disease have abnormal activity, we can try to develop drugs that compensate for that abnormality,” says António Santinha, a doctoral student in Platt’s group and lead author of the study.
The method would also be suitable for use in studying other genetic disorders. “In many congenital diseases, multiple genes play a role, not just one, Santinha says. “This is also the case with mental illnesses such as schizophrenia. Our technique now lets us study such diseases and their genetic causes directly in fully grown animals.” The number of modified genes could be increased from the current 29 to several hundred genes per experiment.
“It’s a big advantage that we can now do these analyses in living organisms, because cells behave differently in culture to how they do as part of a living body,” Santinha says. Another advantage is that the scientists can simply inject the AAVs into the animals’ bloodstreams. There are various different AAVs with different functional properties. In this study, researchers used a virus that enters the animals’ brains. “Depending on what you’re trying to investigate, though, you could also use AAVs that target other organs,” Santinha says.
ETH Zurich has applied for a patent on the technology. The researchers now want to use it as part of a spin-off they are establishing. The technique presented here is one of a series of new genetic editing methods used to alter the genome of cells in a mosaic-like manner. CRISPR perturbation is the technical term for this research approach that involves the perturbation of the genome using CRISPR-Cas gene scissors. This approach is currently revolutionising research in the life sciences. It makes it possible to obtain a great deal of information from a single scientific experiment. As a result, the approach has the potential to accelerate biomedical research, such as in the search for the molecular causes of genetically complex diseases.
The fruit fly acetyltransferase chameau promotes starvation resilience at the expense of longevity
by Anuroop Venkateswaran Venkatasubramani, Toshiharu Ichinose, Mai Kanno, Ignasi Forne, Hiromu Tanimoto, Shahaf Peleg, Axel Imhof in EMBO reports
A team led by biologist Axel Imhof has discovered why an enzyme is important for the survival of fruit flies, even though it can shorten their lives under certain conditions.
Nothing comes for free — this saying applies very well to the work of chameau. As researchers led by Axel Imhof from LMU’s Biomedical Center Munich (BMC) have demonstrated, the enzyme plays an important role in helping fruit flies to survive periods when food is in short supply. However, this comes at a cost: when there is plenty of food available, chameau has a life-shortening effect.
Chameau is an enzyme that chemically modifies proteins, by which it also plays a part in gene regulation. In earlier studies, Imhof and his team had discovered that well-nourished fruit flies live longer if their chameau levels are lowered through mutations. Other researchers had reported a similar effect in mice, when the production of a protein analogous to chameau was reduced.
From an evolutionary perspective, a longer fertile life should lead to more offspring and therefore be favored by evolution. So why does chameau occur in potentially life-shortening quantities? To answer this question, the scientists investigated which mechanisms are influenced by the enzyme.
“The complete absence of chameau results in fatal development defects,” says Imhof. “However, we managed to lower the enzyme levels to around a fifth of its normal levels without affecting viability under normal conditions to study it’s role in stress response.”
Their results show that flies with low chameau levels are less well able to deal with hunger. “When flies are starving, their bodies break down storage molecules such as glycogen and fats. With a defect of the enzyme chameau, this process no longer works as effectively,” says Imhof. “Flies with normal enzyme levels survived without food up to 40 percent longer. In the wild, this gives them an advantage, because they can fly around and search for food during this extra time period.”
Furthermore, the scientists observed that flies with very low chameau levels were much thinner than wild types even when fed well. “This means that these flies not only have a problem with the expenditure, but also with the storage of energy,” explains Imhof. Comprehensive molecular analyses revealed that chameau mutations result in a failure to properly regulate the genes and proteins required for energy storage and expenditure. “Based on research with the mouse chameau gene, we believe that similar mechanisms may also exists in vertebrates such as mice and humans” says Imhof. However, this would require further studies.
That thin flies live longer under normal conditions but are more sensitive to hunger are essentially two sides of the same coin. As long as there are enough nutrients available, the effects of chameau deficits resemble that of caloric restrictions: “Studies on various organisms have shown that reducing caloric intake can lead to a longer lifespan,” says Imhof. When conditions become unfavorable, however, the flies are unable to respond appropriately.
From their findings, the researchers conclude that chameau is important for coping with changing environmental conditions. “As this ability is likely a stronger evolutionary driver than the ability to live a long life, the enzyme has been preserved despite its disadvantages,” says Imhof.
Historical RNA expression profiles from the extinct Tasmanian tiger
by Emilio Mármol-Sánchez, Bastian Fromm, Nikolay Oskolkov, Zoé Pochon, Panagiotis Kalogeropoulos, Eli Eriksson, Inna Biryukova, Vaishnovi Sekar, Erik Ersmark, Björn Andersson, Love Dalén, Marc R. Friedländer in Genome Research
A new study shows the isolation and sequencing of more than a century-old RNA molecules from a Tasmanian tiger specimen preserved at room temperature in a museum collection. This resulted in the reconstruction of skin and skeletal muscle transcriptomes from an extinct species for the first time. The researchers note that their findings have relevant implications for international efforts to resurrect extinct species, including both the Tasmanian tiger and the woolly mammoth, as well as for studying pandemic RNA viruses.
The Tasmanian tiger, also known as the thylacine, was a remarkable apex carnivorous marsupial that was once distributed all across the Australian continent and the island of Tasmania. This extraordinary species found its final demise after European colonization, when it was declared as an agricultural pest and a bounty of £1 per each full-grown animal killed was set by 1888. The last known living Tasmanian tiger died in captivity in 1936 at the Beaumaris Zoo in Hobart, Tasmania.
Recent efforts in de-extinction have focused on the Tasmanian tiger, as its natural habitat in Tasmania is still mostly preserved, and its reintroduction could help recovering past ecosystem equilibriums lost after its final disappearance. However, reconstructing a functional living Tasmanian tiger not only requires a comprehensive knowledge of its genome (DNA) but also of tissue-specific gene expression dynamics and how gene regulation worked, which are only attainable by studying its transcriptome (RNA).
“Resurrecting the Tasmanian tiger or the woolly mammoth is not a trivial task, and will require a deep knowledge of both the genome and transcriptome regulation of such renowned species, something that only now is starting to be revealed,” says Emilio Mármol, the lead author of a study recently published in the Genome Research journal by researchers at SciLifeLab in collaboration with the Centre for Palaeogenetics, a joint venture between the Swedish Museum of Natural History and Stockholm University.
The researchers behind this study have sequenced, for the first time, the transcriptome of the skin and skeletal muscle tissues from a 130-year-old desiccated Tasmanian tiger specimen preserved at room temperature in the Swedish Museum of Natural History in Stockholm. This led to the identification of tissue-specific gene expression signatures that resemble those from living extant marsupial and placental mammals. The recovered transcriptomes were of such good quality that it was possible to identify muscle- and skin-specific protein coding RNAs, and led to the annotation of missing ribosomal RNA and microRNA genes, the later following MirGeneDB recommendations.
“This is the first time that we have had a glimpse into the existence of thylacine-specific regulatory genes, such as microRNAs, that got extinct more than one century ago,” says Marc R. Friedländer, Associate Professor at the Department of Molecular Biosciences, The Wenner-Gren Institute at Stockholm University and SciLifeLab.
This pioneering study opens up new exciting opportunities and implications for exploring the vast collections of specimens and tissues stored at museums across the globe, where RNA molecules might await to be uncovered and sequenced.
“In the future, we may be able to recover RNA not only from extinct animals, but also RNA virus genomes such as SARS-CoV2 and their evolutionary precursors from the skins of bats and other host organisms held in museum collections,” says Love Dalén, Professor of evolutionary genomics at Stockholm University and the Centre for Palaeogenetics.
The authors of the study say they are excited for future holistic research developments integrating both genomics and transcriptomics towards a new era in palaeogenetics beyond DNA.
Digital assay for rapid electronic quantification of clinical pathogens using DNA nanoballs
by Muhammad Tayyab, Donal Barrett, Gijs van Riel, Shujing Liu, Björn Reinius, Curt Scharfe, Peter Griffin, Lars M. Steinmetz, Mehdi Javanmard, Vicent Pelechano in Science Advances
Rutgers researchers have developed a way of detecting the early onset of deadly infectious diseases using a test so ultrasensitive that it could someday revolutionize medical approaches to epidemics.
The test is an electronic sensor contained within a computer chip. It employs nanoballs — microscopic spherical clumps made of tinier particles of genetic material, each of those with diameters 1,000 times smaller than the width of a human hair — and combines that technology with advanced electronics.
“During the COVID pandemic, one of the things that didn’t exist but could have stemmed the spread of the virus was a low-cost diagnostic that could flag people known as the ‘quiet infected’ — patients who don’t know they are infected because they are not exhibiting symptoms,” said Mehdi Javanmard, a professor in the Department of Electrical and Computer Engineering in the Rutgers School of Engineering and an author of the study. “In a pandemic, pinpointing an infection early with accuracy is the Holy Grail. Because once a person is showing symptoms — sneezing and coughing — it’s too late. That person has probably infected 20 people.”
For the past 20 years, Javanmard has been developing biosensors — devices that monitor and transmit information about a life process. During the COVID-19 pandemic, he became disheartened about the extent of infections and the extreme loss of life. He believed there had to be a way of using biosensors as a test to detect illness earlier.
Working with Muhammad Tayyab, a Rutgers doctoral student and co-author of the study, Javanmard and research colleagues at the Karolinska Institute in Sweden and Stanford and Yale universities started brainstorming.
“We thought: How is there a way where we can leverage our individual expertise to build something new?” Javanmard said.
The biosensor developed by the team works through a series of steps. First, it zeroes in on a virus’ characteristic sequence of nucleic acids — naturally occurring chemical compounds that serve as the primary information-carrying molecules in a cell. Next, because it amplifies any nucleic acid sequence found in the sample, it makes many more copies, as many as 10,000. Then, it clumps those thousands of specks of nucleic acids into nanoballs that are “large” enough to be detected.
The nanoballs are identified electrically when they are directed individually through minute channels containing electrodes on opposite sides. The process is akin to people walking single file through an airport security gate and being X-rayed one by one.
“Our method involves taking the viral nucleic acid material and rolling it up into a ball of DNA that is large enough to be detected by a cell measurement device known as an electronic cytometer,” Javanmard said. “As a result, we can flag the infection at its earliest stages when the concentration is still very low.”
The approach works in samples taken from blood and saliva and has been shown to detect early infections by several viruses, including the rhinovirus causing the common cold, and even bacterial infections such as tuberculosis. The technology, which has been miniaturized and is contained within a computer chip, is small enough to be portable and wearable.
The most immediate use, once commercialized, is expected to be the test’s efficacy in flagging viral infections at early stages. Ultimately, the technology also may be used to test bacteria-based illnesses as well as bacterial contamination found in food and water supplies, Javanmard said.
“These pathogens can be devastating to the economy, to people’s livelihoods, and to people’s health,” Javanmard said. “We should not stop preparing for this. We need to be creating the next generation of tools to stop them. And that’s really what this technology is.”
Stepwise emergence of the neuronal gene expression program in early animal evolution
by Sebastián R. Najle, Xavier Grau-Bové, Anamaria Elek, et al in Cell
A study sheds new light on the evolution of neurons, focusing on the placozoans, a millimetre-sized marine animal. Researchers at the Centre for Genomic Regulation in Barcelona find evidence that specialized secretory cells found in these unique and ancient creatures may have given rise to neurons in more complex animals.
Placozoans are tiny animals, around the size of a large grain of sand, which graze on algae and microbes living on the surface of rocks and other substrates found in shallow, warm seas. The blob-like and pancake-shaped creatures are so simple that they live without any body parts or organs. These animals, thought to have first appeared on Earth around 800 million years ago, are one of the five main lineages of animals alongside Ctenophora (comb jellies), Porifera (sponges), Cnidaria (corals, sea anemones and jellyfish) and Bilateria (all other animals).
The sea creatures coordinate their behaviour thanks to peptidergic cells, special types of cells that release small peptides which can direct the animal’s movement or feeding. Driven by the intrigue of the origin of these cells, the authors of the study employed an array of molecular techniques and computational models to understand how placozoan cell types evolved and piece together how our ancient ancestors might have looked and functioned.
The researchers first made a map of all the different placozoan cell types, annotating their characteristics across four different species. Each cell type has a specialised role which comes from certain sets of genes. The maps or ‘cell atlases’ allowed researchers to chart clusters or ‘modules’ of these genes. They then created a map of the regulatory regions in DNA that control these gene modules, revealing a clear picture about what each cell does and how they work together. Finally, they carried out cross-species comparisons to reconstruct how the cell types evolved.
The research showed that the main nine cell types in placozoans appear to be connected by many “in-between” cell types which change from one type to another. The cells grow and divide, maintaining the delicate balance of cell types required for the animal to move and eat. The researchers also found fourteen different types of peptidergic cells, but these were different to all other cells, showing no in-between types or any signs of growth or division.
Surprisingly, the peptidergic cells shared many similarities to neurons — a cell type which didn’t appear until many millions of years later in more advanced animals such as and bilateria. Cross-species analyses revealed these similarities are unique to placozoans and do not appear in other early-branching animals such as sponges or comb jellies (ctenophores).
The similarities between peptidergic cells and neurons were threefold. First, the researchers found that these placozoan cells differentiate from a population of progenitor epithelial cells via developmental signals that resemble neurogenesis, the process by which new neurons are formed, in cnidaria and bilateria.
Second, they found that peptidergic cells have many gene modules required to build the part of a neuron which can send out a message (the pre-synaptic scaffold). However, these cells are far from being a true neuron, as they lack the components for the receiving end of a neuronal message (post-synaptic) or the components required for conducting electrical signals.
Finally, the authors used deep learning techniques to show that placozoan cell types communicate with each other using a system in cells where specific proteins, called GPCRs (G-protein coupled receptors), detect outside signals and start a series of reactions inside the cell. These outside signals are mediated by neuropeptides, chemical messengers used by neurons in many different physiological processes.
“We were astounded by the parallels,” says Dr. Sebastián R. Najle, co-first author of the study and postdoctoral researcher at the Centre for Genomic Regulation. “The placozoan peptidergic cells have many similarities to primitive neuronal cells, even if they aren’t quite there yet. It’s like looking at an evolutionary stepping stone.”
The study demonstrates that the building blocks of the neuron were forming 800 million years ago in ancestral animals grazing inconspicuously in the shallow seas of ancient Earth. From an evolutionary point of view, early neurons might have started as something like the peptidergic secretory cells of today’s placozoans. These cells communicated using neuropeptides, but eventually gained new gene modules which enabled cells to create post-synaptic scaffolds, form axons and dendrites and create ion channels that generate fast electrical signals — innovations which were critical for the dawn of the neuron around one hundred million years after the ancestors of placozoans first appeared on Earth.
However, the complete evolutionary story of nerve systems is still to be told. The first modern neuron is thought to have originated in the common ancestor of cnidarians and bilaterians around 650 million years ago. And yet, neuronal-like cells exist in ctenophores, although they have important structural differences and lack the expression of most genes found in modern neurons. The presence of some of these neuronal genes in the cells of placozoans and their absence in ctenophores raises fresh questions about the evolutionary trajectory of neurons.
“Placozoans lack neurons, but we’ve now found striking molecular similarities with our neural cells. Ctenophores have neural nets, with key differences and similarities with our own. Did neurons evolve once and then diverge, or more than once, in parallel? Are they a mosaic, where each piece has a different origin? These are open questions that remain to be addressed,” says Dr. Xavier Grau-Bové, co-first author of the study and postdoctoral researcher at the Centre for Genomic Regulation.
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