Sugar-powered biobattery has 10 times the energy storage of lithium: Your smartphone might soon run on enzymes

Sugar-powered biobattery has 10 times the energy storage of lithium: Your smartphone might soon run on enzymes

  7:33:00 PM    Science Lover

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As you probably know, from sucking down cans of Coke and masticating on candy, sugar — glucose, fructose, sucrose, dextrose — is an excellent source of energy. Biologically speaking, sugar molecules are energy-dense, easy to transport, and cheap to digest. There is a reason why almost every living cell on Earth generates its energy (ATP) from glucose. Now, researchers at Virginia Tech have successfully created a sugar-powered fuel cell that has an energy storage density of 596 amp-hours per kilo — or “one order of magnitude” higher than lithium-ion batteries. This fuel cell is refillable with a solution of maltodextrin, and its only by products are electricity and water. The chief researcher, Y.H. Percival Zhang, says the tech could be commercialized in as soon as three years.

Now, it’s not exactly news that sugar is an excellent energy source. As a culture we’ve probably known about it since before we were Homo sapiens. The problem is, unless you’re a living organism or some kind of incendiary device, extracting that energy is difficult. In nature, an enzymatic pathway is used — a production line of tailor-made enzymes that meddle with the glucose molecules until they become ATP. Because it’s easy enough to produce enzymes in large quantities, researchers have tried to create fuel cells that use artificial “metabolism” to break down glucose into electricity (biobatteries), but it has historically proven very hard to find the right pathway for maximum efficiency and to keep the enzymes in the right place over a long period of time.

A diagram of the enzymatic fuel cell. The little Pac-Man things are enzymes.

Now, however, Zhang and friends at Virginia Tech appear to have built a high-density fuel cell that uses an enzymatic pathway to create a lot of electricity from glucose. There doesn’t seem to be much information on how stable this biobattery is over multiple refills, but if Zhang thinks it could be commercialized in three years, that’s a very good sign. Curiously, the research paper says that the enzymes are non-immobilized — meaning Zhang found a certain battery chemistry that doesn’t require the enzymes to be kept in place… or, alternatively, that it will only work for a very short time.

The Virginia Tech biobattery uses 13 enzymes, plus air (it’s an air-breathing biobattery), to produce nearly 24 electrons from a single glucose unit. This equates to a power output of 0.8 mW/cm, current density of 6 mA/cm, and energy storage density of 596 Ah/kg. This last figure is impressive, at roughly 10 times the energy density of the lithium-ion batteries in your mobile devices. [Research paper: doi:10.1038/ncomms4026 - "A high-energy-density sugar biobattery based on a synthetic enzymatic pathway"]

If Zhang’s biobatteries pan out, you might soon be recharging your smartphone by pouring in a solution of 15% maltodextrin. That battery would not only be very safe (it produces water and electricity), but very cheap to run and very green. This seems to fit in perfectly with Zhang’s homepage, which talks about how his main goals in life are replacing crude oil with sugar, and feeding the world.

The other area in which biobatteries might be useful is powering implanted devices, such as pacemakers — or, in the future, subcutaneous sensors and computers. Such a biobattery could feed on the glucose in your bloodstream, providing an endless supply of safe electricity for the myriad implants that futuristic technocrats will surely have. 

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Brain Development Is Guided by Junk DNA That Isn't Really Junk

Brain Development Is Guided by Junk DNA That Isn't Really Junk

  8:02:00 AM    Science Lover

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Specific DNA once dismissed as junk plays an important role in brain development and might be involved in several devastating neurological diseases, UC San Francisco scientists have found.

UCSF researchers have uncovered a role in brain development and in neurological disease for little appreciated molecules called long noncoding RNA. In this image, fluorescent dyes track the presence of the RNA molecules and the genes they affect in the developing mouse brain. (Credit: Image courtesy of Alexander Ramos)

Their discovery in mice is likely to further fuel a recent scramble by researchers to identify roles for long-neglected bits of DNA within the genomes of mice and humans alike.

While researchers have been busy exploring the roles of proteins encoded by the genes identified in various genome projects, most DNA is not in genes. This so-called junk DNA has largely been pushed aside and neglected in the wake of genomic gene discoveries, the UCSF scientists said.

In their own research, the UCSF team studies molecules called long noncoding RNA (lncRNA, often pronounced as "link" RNA), which are made from DNA templates in the same way as RNA from genes.

"The function of these mysterious RNA molecules in the brain is only beginning to be discovered," said Daniel Lim, assistant professor of neurological surgery, a member of the Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research at UCSF, and the senior author of the study, published online April 11 in the journal Cell Stem Cell.

Alexander Ramos, a student enrolled in the MD/PhD program at UCSF and first author of the study, conducted extensive computational analysis to establish guilt by association, linking lncRNAs within cells to the activation of genes.

Ramos looked specifically at patterns associated with particular developmental pathways or with the progression of certain diseases. He found an association between a set of 88 long noncoding RNAs and Huntington's disease, a deadly neurodegenerative disorder. He also found weaker associations between specific groups of long noncoding RNAs and Alzheimer's disease, convulsive seizures, major depressive disorder and various cancers.

"Alex was the team member who developed this new research direction, did most of the experiments, and connected results to the lab's ongoing work," Lim said. The study was mostly funded through Lim's grant - a National Institutes of Health (NIH) Director's New Innovator Award, a competitive award for innovative projects that have the potential for unusually high impact.

Unlike messenger RNA, which is transcribed from the DNA in genes and guides the production of proteins, lncRNA molecules do not carry the blueprints for proteins. Because of this fact, they were long thought to not influence a cell's fate or actions.

Nonetheless, lncRNAs also are transcribed from DNA in the same way as messenger RNA, and they, too, consist of unique sequences of nucleic acid building blocks.

Evidence indicates that lncRNAs can tether structural proteins to the DNA-containing chromosomes, and in so doing indirectly affect gene activation and cellular physiology without altering the genetic code. In other words, within the cell, lncRNA molecules act "epigenetically" -- beyond genes -- not through changes in DNA.

The brain cells that the scientists focused on the most give rise to various cell types of the central nervous system. They are found in a region of the brain called the subventricular zone, which directly overlies the striatum. This is the part of the brain where neurons are destroyed in Huntington's disease, a condition triggered by a single genetic defect.

Ramos combined several advanced techniques for sequencing and analyzing DNA and RNA to identify where certain chemical changes happen to the chromosomes, and to identify lncRNAs on specific cell types found within the central nervous system. The research revealed roughly 2,000 such molecules that had not previously been described, out of about 9,000 thought to exist in mammals ranging from mice to humans.

In fact, the researchers generated far too much data to explore on their own. The UCSF scientists created a website through which their data can be used by others who want to study the role of lncRNAs in development and disease.

"There's enough here for several labs to work on," said Ramos, who has training grants from the California Institute for Regenerative Medicine (CIRM) and the NIH.

"It should be of interest to scientists who study long noncoding RNA, the generation of new nerve cells in the adult brain, neural stem cells and brain development, and embryonic stem cells," he said.

Other co-authors who worked on the study include UCSF postdoctoral fellows Aaron Diaz, PhD, Abhinav Nellore, PhD, Michael Oldham, PhD, Jun Song, PhD, Ki-Youb Park, PhD, and Gabriel Gonzales-Roybal, PhD; and MD/PhD student Ryan Delgado. Additional funders of the study included the Sontag Foundation and the Sandler Foundation.

UCSF is a leading university dedicated to promoting health worldwide through advanced biomedical research, graduate-level education in the life sciences and health professions, and excellence in patient care. 
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Scientists Create Nanoscale Window to Biological World

Scientists Create Nanoscale Window to Biological World

  9:13:00 AM    Science Lover

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If the key to winning battles is knowing both your enemy and yourself, then scientists are now well on their way toward becoming the Sun Tzus of medicine by taking a giant step toward a priceless advantage -- the ability to see the soldiers in action on the battlefield.

A novel microfluidics platform allowed viewing of structural 
details of rotavirus double-layered particles; the 3-D graphic 
of the virus, in purple, was reconstructed from data gathered by 
the new technique. (Credit: Virginia Tech)

Investigators at the Virginia Tech Carilion Research Institute have invented a way to directly image biological structures at their most fundamental level and in their natural habitats. The technique is a major advancement toward the ultimate goal of imaging biological processes in action at the atomic level.

"It's sort of like the difference between seeing Han Solo frozen in carbonite and watching him walk around blasting stormtroopers," said Deborah Kelly, an assistant professor at the VTC Research Institute and a lead author on the paper describing the first successful test of the new technique. "Seeing viruses, for example, in action in their natural environment is invaluable."

The technique involves taking two silicon-nitride microchips with windows etched in their centers and pressing them together until only a 150-nanometer space between them remains. The researchers then fill this pocket with a liquid resembling the natural environment of the biological structure to be imaged, creating a microfluidic chamber.

Then, because free-floating structures yield images with poor resolution, the researchers coat the microchip's interior surface with a layer of natural biological tethers, such as antibodies, which naturally grab onto a virus and hold it in place.

In a recent study in Lab on a Chip, Kelly joined Sarah McDonald, also an assistant professor at the VTC Research Institute, to prove that the technique works. McDonald provided a pure sample of rotavirus double-layered particles for the study.

"What's missing in the field of structural biology right now is dynamics -- how things move in time," said McDonald. "Debbie is developing technologies to bridge that gap, because that's clearly the next big breakthrough that structural biology needs."

Rotavirus is the most common cause of severe diarrhea among infants and children. By the age of 5, nearly every child in the world has been infected at least once. And although the disease tends to be easily managed in the developed world, in developing countries rotavirus kills more than 450,000 children a year.

At the second step in the pathogen's life cycle, rotavirus sheds its outer layer, which allows it to enter a cell, and becomes what is called a double-layered particle. Once its second layer is exposed, the virus is ready to begin using the cell's own infrastructure to produce more viruses. It was the viral structure at this stage that the researchers imaged in the new study.

Kelly and McDonald coated the interior window of the microchip with antibodies to the virus. The antibodies, in turn, latched onto the rotaviruses that were injected into the microfluidic chamber and held them in place. The researchers then used a transmission electron microscope to image the prepared slide.

The technique worked perfectly.

The experiment gave results that resembled those achieved using traditional freezing methods to prepare rotavirus for electron microscopy, proving that the new technique can deliver accurate results.

"It's the first time scientists have imaged anything on this scale in liquid," said Kelly.

The next step is to continue to develop the technique with an eye toward imaging biological structures dynamically in action. Specifically, McDonald is looking to understand how rotavirus assembles, so as to better know and develop tools to combat this particular enemy of children's health.

The researchers said their ongoing collaboration is an example of the cross-disciplinary work that is becoming a hallmark of the VTC Research Institute.

"It's an ideal collaboration because Sarah provides a phenomenal model system by which we can develop new technologies to move the field of microstructural biology forward," said Kelly.

"It's very win-win," McDonald added. "While the virus is a great tool for Debbie to develop her techniques, her technology is critical for allowing me to understand how this deadly virus assembles and changes dynamically over time."

The paper "Visualizing viral assemblies in a nanoscale biosphere" was published online and will appear in a 2013 edition of Lab on a Chip.

The authors are Brian Gilmore, a research associate at the VTC Research Institute; Shannon Showalter, a research assistant at the VTC Research Institute; Madeline Dukes, an applications scientist at Protochips; Justin Tanner, a postdoctoral associate at the VTC Research Institute; Andrew Demmert, a student at the Virginia Tech Carilion School of Medicine; McDonald, in addition to her position at the VTC Research Institute, is an assistant professor of biomedical sciences and pathobiology in the Virginia-Maryland Regional College of Veterinary Medicine; and Kelly, in addition to her position at the VTC Research Institute, is an assistant professor of biological sciences in Virginia Tech's College of Science.

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Mental time-travel in birds

Mental time-travel in birds

  10:22:00 AM    Science Lover

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Certain types of birds may track army ant swarms using sophisticated memory and the ability to plan for the future.

White Whiskered Puff bird Credit: Glenn M Duggan FZS

Some tropical birds collect their prey at army ant raids, where massive swarms of ants sweep through the forest and drive out insects. The behaviour of interest is called bivouac checking; it allows these birds to track the cyclical raid activity of army ant colonies.

Army ants have regular alternating periods of high and low raiding activity, and birds visit the ants’ temporary nest sites (bivouacs) to determine which colonies are raiding on a given day.

The new findings published today in the journal Behavioural Ecology, suggest that bivouac checking allows birds to keep track of multiple army ant colonies, keeping account of which colonies are in periods of high-raiding activity while avoiding colonies with low-raiding activity.

Recent research has discovered that birds check army ant bivouacs at the end of the day, after they have fed at the raid. They may use the information about the army ant nest location the next day to find the ants again, thus accessing a past memory (the nest location) to fulfil a future need (bird will be hungry tomorrow), also known as ‘mental time-travel’.

Two of the authors of the study Corina Logan of the University of Cambridge, and Sean O’Donnell of the University of Washington, observed bivouac checking behaviour in Monteverde, Costa Rica.

Mental time-travel consists of two elements: the ability to remember past events and the ability to anticipate and plan for future events. It has traditionally been considered a quality unique to humans. However, ever since Nicola Clayton of the University of Cambridge discovered that scrub jays (a species of large-brained crow) can remember the past and plan for the future, there have been a suite of studies showing evidence of this ability in other species as well. We now know that corvids (birds in the crow family), some primates, and possibly rats have all shown the ability to remember the past and plan for the future.

Corina Logan, said: “We suspect that future planning could be involved in bivouac-checking bird behaviour because the birds were checking bivouacs when they were not hungry, a behaviour that does not make sense until the next morning upon return to the bivouac when the bird finds the ants raiding again and encounters its next meal – a delayed benefit.”

Until recently, it has been difficult to find model systems for studying mental time travel in an ecologically relevant way. “The fact that we might have happened on a whole new system for exploring these capacities is quite exciting,” added Corina Logan.

Provided by University of Cambridge



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Editing the Genome: Scientists Unveil New Tools for Rewriting the Code of Life

Editing the Genome: Scientists Unveil New Tools for Rewriting the Code of Life

  2:50:00 AM    Science Lover

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The power to edit genes is as revolutionary, immediately useful and unlimited in its potential as was Johannes Gutenberg's printing press. And like Gutenberg's invention, most DNA editing tools are slow, expensive, and hard to use -- a brilliant technology in its infancy. Now, Harvard researchers developing genome-scale editing tools as fast and easy as word processing have rewritten the genome of living cells using the genetic equivalent of search and replace -- and combined those rewrites in novel cell strains, strikingly different from their forebears.

Researchers have unveiled genome-engineering technologies capable of fundamentally re-engineering genomes from the nucleotide to the megabase scale. Treating the chromosome as both an editable and an evolvable template, the researchers have demonstrated methods to rewrite a cell's genome through powerful new tools for biotechnology, energy and agriculture. (Credit: iStockphoto/Zmeel Photography)

"The payoff doesn't really come from making a copy of something that already exists," said George Church, a professor of genetics at Harvard Medical School who led the research effort in collaboration with Joe Jacobson, an associate professor at the Media Lab at the Massachusetts Institute of Technology. "You have to change it -- functionally and radically."

Such change, Church said, serves three goals. The first is to add functionality to a cell by encoding for useful new amino acids. The second is to introduce safeguards that prevent cross-contamination between modified organisms and the wild. A third, related aim, is to establish multi-viral resistance by rewriting code hijacked by viruses. In industries that cultivate bacteria, including pharmaceuticals and energy, such viruses affect up to 20 percent of cultures. A notable example afflicted the biotech company Genzyme, where estimates of losses due to viral contamination range from a few hundred million dollars to more than $1 billion.

In a paper scheduled for publication July 15 in Science, the researchers describe how they replaced instances of a codon -- a DNA "word" of three nucleotide letters -- in 32 strains of E. coli, and then coaxed those partially-edited strains along an evolutionary path toward a single cell line in which all 314 instances of the codon had been replaced. That many edits surpasses current methods by two orders of magnitude, said Harris Wang, a research fellow in Church's lab at the Wyss Institute for Biologically Inspired Engineering who shares lead-author credit on the paper with Farren Isaacs, an assistant professor of molecular, cellular and developmental biology at Yale University and former Harvard research fellow, and Peter Carr, a research scientist at the MIT Media Lab.



In the genetic code, most codons specify an amino acid, a protein building block. But a few codons tell the cell when to stop adding amino acids to a protein chain, and it was one of these "stop" codons that the Harvard researchers targeted. With just 314 occurrences, the TAG stop codon is the rarest word in the E. coli genome, making it a prime target for replacement. Using a platform called multiplex automated genome engineering, or MAGE, the team replaced instances of the TAG codon with another stop codon, TAA, in living E. coli cells. (Unveiled by the team in 2009, the MAGE process has been called an evolution machine for its ability to accelerate targeted genetic change in living cells.)

While MAGE, a small-scale engineering process, yielded cells in which TAA codons replaced some but not all TAG codons, the team constructed 32 strains that, taken together, included every possible TAA replacement. Then, using bacteria's innate ability to trade genes through a process called conjugation, the researchers induced the cells to transfer genes containing TAA codons at increasingly larger scales. The new method, called conjugative assembly genome engineering, or CAGE, resembles a playoff bracket -- a hierarchy that winnows 16 pairs to eight to four to two to one -- with each round's winner possessing more TAA codons and fewer TAG, explains Isaacs, who invokes "March Madness."

"We're testing decades-old theories on the conservation of the genetic code," Isaacs said. "And we're showing on a genome-wide scale that we're able to make these changes."

Eager to share their enabling technology, the team published their results as CAGE reached the semifinal round. Results suggested that the final four strains were healthy, even as the team assembled four groups of 80 engineered alterations into stretches of the chromosome surpassing 1 million DNA base pairs. "We encountered a great deal of skepticism early on that we could make so many changes and preserve the health of these cells," Carr said. "But that's what we've seen."

The researchers are confident that they will create a single strain in which TAG codons are completely eliminated. The next step, they say, is to delete the cell's machinery that reads the TAG gene -- freeing up the codon for a completely new purpose, such as encoding a novel amino acid.

"We're trying to challenge people," Wang said, "to think about the genome as something that's highly malleable, highly editable."

This research was funded by U.S. Department of Energy and the National Science Foundation.

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U of T researchers build an antenna for light

U of T researchers build an antenna for light

  9:53:00 AM    Science Lover

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University of Toronto researchers have derived inspiration from the photosynthetic apparatus in plants to engineer a new generation of nanomaterials that control and direct the energy absorbed from light.

Their findings are reported in a forthcoming issue of Nature Nanotechnology, which will be released on July 10, 2011.

The U of T researchers, led by Professors Shana Kelley and Ted Sargent, report the construction of what they term "artificial molecules."

"Nanotechnologists have for many years been captivated by quantum dots – particles of semiconductor that can absorb and emit light efficiently, and at custom-chosen wavelengths," explained co-author Kelley, a Professor at the Leslie Dan Faculty of Pharmacy, the Department of Biochemistry in the Faculty of Medicine, and the Department of Chemistry in the Faculty of Arts & Science. "What the community has lacked – until now – is a strategy to build higher-order structures, or complexes, out of multiple different types of quantum dots. This discovery fills that gap."

The team combined its expertise in DNA and in semiconductors to invent a generalized strategy to bind certain classes of nanoparticles to one another.

"The credit for this remarkable result actually goes to DNA: its high degree of specificity – its willingness to bind only to a complementary sequence – enabled us to build rationally-engineered, designer structures out of nanomaterials," said Sargent, a Professor in The Edward S. Rogers Sr. Department of Electrical & Computer Engineering at the University of Toronto, who is also the Canada Research Chair in Nanotechnology. "The amazing thing is that our antennas built themselves – we coated different classes of nanoparticles with selected sequences of DNA, combined the different families in one beaker, and nature took its course. The result is a beautiful new set of self-assembled materials with exciting properties."



Traditional antennas increase the amount of an electromagnetic wave – such as a radio frequency – that is absorbed, and then funnel that energy to a circuit. The U of T nanoantennas instead increased the amount of light that is absorbed and funneled it to a single site within their molecule-like complexes. This concept is already used in nature in light harvesting antennas, constituents of leaves that make photosynthesis efficient. "Like the antennas in radios and mobile phones, our complexes captured dispersed energy and concentrated it to a desired location. Like the light harvesting antennas in the leaves of a tree, our complexes do so using wavelengths found in sunlight," explained Sargent.

"Professors Kelley and Sargent have invented a novel class of materials with entirely new properties. Their insight and innovative research demonstrates why the University of Toronto leads in the field of nanotechnology," said Professor Henry Mann, Dean of the Leslie Dan Faculty of Pharmacy.

"This is a terrific piece of work that demonstrates our growing ability to assemble precise structures, to tailor their properties, and to build in the capability to control these properties using external stimuli," noted Paul S. Weiss, Fred Kavli Chair in NanoSystems Sciences at UCLA and Director of the California NanoSystems Institute.

Kelley explained that the concept published in today's Nature Nanotechnology paper is a broad one that goes beyond light antennas alone.

"What this work shows is that our capacity to manipulate materials at the nanoscale is limited only by human imagination. If semiconductor quantum dots are artificial atoms, then we have rationally synthesized artificial molecules from these versatile building blocks."

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The turn of the corkscrew : Structural analysis uncovers mechanisms of gene expression

The turn of the corkscrew : Structural analysis uncovers mechanisms of gene expression

  9:36:00 PM    Science Lover

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The diverse functions of living cells are all based on the information encoded in the structure of the hereditary material DNA. Gene expression must therefore be tightly controlled, and this task is accomplished by the binding of regulatory proteins to, and their removal from, specific DNA sequences. One class of large molecular machines known as Swi2/Snf2 remodelers plays a central role in modulating these processes. However, until now, it was not clear how Swi2/Snf2 remodelers actually work. A team led by Professor Karl-Peter Hopfner at the Gene Center at Ludwig-Maximilians-Universität (LMU) in Munich has clarified the structure and function of the remodeler Mot1 (Modifier of Transcription 1), which binds directly to DNA. It turns out that Mot1 acts like a molecular corkscrew that migrates along the DNA, following its helical contour. During its progress, Mot1 displaces a crucial transcription factor called TBP (for “TATA Box Binding Protein”) from the DNA. Removal of TBP from a TATA box represses transcription of the adjacent gene, and the protein encoded by that gene is no longer synthesized. At the same time, TBP is stabilized and its binding specificity is changed, which facilitates the expression of genes that lack TATA boxes and code for other proteins. (Nature 6 July 2011)

The DNA in the cells of higher organisms is tightly wrapped around protein complexes called nucleosomes. This type of structural organization not only makes it possible to package the long DNA molecules in a highly compact form, it also provides the basis for the controlled expression of genetic information. Densely packed sections of the molecule are effectively in a repressed state, and genes located in these DNA segments cannot be transcribed. Activation of repressed genes depends on the intervention of complex molecular machines, so-called Swi2/Snf2 remodelers, which reorganize condensed stretches of DNA so as to make them accessible for transcription. The precise mode of action of remodelers has so far been unclear, mainly because most of them are made up of several components and the active complexes are difficult to study. This is why Hopfner chose to study Mot1, which is a comparatively simple representative of the family that functions as an Swi2/Snf2 remodeler on its own, and can serve as a guide to understanding the more complicated members of the class. Mot1 is known to participate in the control of gene expression, but how exactly it does so is not well understood.

The first stage of the process that leads to the synthesis of a given protein is the transcription of the specific segment of DNA that codes for it into molecules of messenger RNA. This initial step requires the action of so-called transcription factors. One of the most important of these is TBP, which binds preferentially to DNA sequences called TATA boxes that are located near the beginnings of many genes. Binding of TBP introduces a kink into the DNA, and this landmark serves as a platform for the binding of further proteins, ultimately leading to the assembly of the complex necessary for the initiation of transcription. Mot1 regulates transcription by actively removing TBP from the DNA, using ATP as a source of energy. “How Mot1 dissociates the TBP-DNA complex was completely unclear up to now,” says Hopfner. With the aid of so-called hybrid methods - in which data obtained from high-resolution X-ray diffraction analysis of the crystallized protein complex with images of the same molecular complex taken with the electron microscope were combined - Hopfner’s team was able to define the three-dimensional structure of the Mot1-TBP complex for the first time. This revealed how Mot1 recognizes the surface of the DNA-bound TBP. “Once Mot1 has recognized TBP, it binds to the adjacent DNA and begins to migrate along the DNA strand, using the energy released by the hydrolysis of ATP to power its movement. This helical movement, which is reminiscent of the insertion of a corkscrew, causes TBP to detach from the DNA,” explains Dr Petra Wollmann, who is first author on the new study. The researchers were surprised to find that Mot1 contains a strikingly extended loop. After TBP has dissociated from the DNA, this loop masks TBP’s DNA binding site and prevents the protein from reoccupying it.



Previous studies had reported what appeared to be paradoxical observations, which indicated that Mot1 inhibits transcription of TATA box-containing genes while facilitating the expression of genes that lack canonical TATA boxes. “Our results suggest that Mot1 also stabilizes the DNA-free conformation of TBP, increasing the probability that it can reach, bind to and activate genes that lack TATA boxes,” explains Hopfner. In other words, Mot1 is also a redistribution factor, which enables TBP to bind to different sequences and thus controls its association with other cellular components. This combination of detachment and redistribution functions may be a common feature of remodeling complexes, and would help to explain how they mediate the large-scale redistribution of DNA-binding regulatory proteins. (göd/PH)

The project was carried out under the auspices of two Clusters of Excellence - the Center for Integrated Protein Science Munich (CiPSM) and the Munich Centre for Advanced Photonics (MAP). The work was also supported by the German Research Foundation (DFG) as part of the Collaborate Research Centers (SFB) 646 and TR5, and by the LMUexcellent Investment Fund.

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Researchers decipher protein structure of key molecule in DNA transcription system

Researchers decipher protein structure of key molecule in DNA transcription system

  11:59:00 PM    Science Lover

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The research adds an important link to discoveries that have enabled scientists to gain a deeper understanding of how cells translate genetic information into the proteins and processes of life. The findings, published in the July 3 advance online issue of the journal Nature, were reported by a research team led by Yuichiro Takagi, Ph.D., assistant professor of biochemistry and molecular biology at Indiana University School of Medicine.

Scientists have deciphered the structure of an essential 
part of Mediator, a complex molecular machine that plays 
a vital role in regulating the transcription of DNA.

The fundamental operations of all cells are controlled by the genetic information – the genes –stored in each cell's DNA, a long double-stranded chain. Information copied from sections of the DNA – through a process called transcription – leads to synthesis of messenger RNA, eventually enabling the production of proteins necessary for cellular function. Transcription is undertaken by the enzyme called RNA polymerase II.

As cellular operations proceed, signals are sent to the DNA asking that some genes be activated and others be shut down. The Mediator transcription regulator accepts and interprets those instructions, telling RNA polymerase II where and when to begin the transcription process.

Mediator is a gigantic molecular machine composed of 25 proteins organized into three modules known as the head, the middle, and the tail. Using X-ray crystallography, the Takagi team was able to describe in detail the structure of the Mediator Head module, the most important for interactions with RNA polymerase II.

"It's turned out to be extremely novel, revealing how a molecular machine is built from multiple proteins," said Takagi.

"As a molecular machine, the Mediator head module needs to have elements of both stability and flexibility in order to accommodate numerous interactions. A portion of the head we named the neck domain provides the stability by arranging the five proteins in a polymer-like structure," he said.



"We call it the alpha helical bundle," said Dr. Takagi. "People have seen structures of alpha helical bundles before but not coming from five different proteins."

A research team led by Yuichiro Takagi, Ph.D.,
Indiana University School of Medicine, has
deciphered the structure of an essential part
of Mediator, a complex molecular machine
that plays a vital role in regulating the
transcription of DNA.

"This is a completely noble structure," he said.

One immediate benefit of the research will be to provide detailed mapping of previously known mutations that affect the regulation of the transcription process, he said.

The ability to solve such complex structures will be important because multi-protein complexes such as Mediator will most likely become a new generation of drug targets for treatment of disease, he said.

Previously, the structure of RNA polymerase II was determined by Roger Kornberg of Stanford University, with whom Dr. Takagi worked prior to coming to IU School of Medicine. Kornberg received the Nobel Prize in 2006 for his discoveries. The researchers who described the structure of the ribosome, the protein production machine, were awarded the Nobel Prize in 2009. The structure of the entire Mediator has yet to be described, and thus remains the one of grand challenges in structure biology. Dr. Takagi's work on the Mediator head module structure represents a major step towards a structure determination of the entire Mediator. In addition to Dr. Takagi as a senior author, the lead author for the Nature paper was Tsuyoshi Imasaki, Ph.D., of the IU School of Medicine. Other collaborators included researchers at The Scripps Research Institute, Stanford University, Memorial Sloan-Kettering Cancer Center and the European Molecular Biology Laboratory.

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Loudest Animal Is Recorded for the First Time

Loudest Animal Is Recorded for the First Time

  7:07:00 PM    Science Lover

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Scientists have shown for the first time that the loudest animal on earth, relative to its body size, is the tiny water boatman, Micronecta scholtzi. At 99.2 decibels, this represents the equivalent of listening to an orchestra play loudly while sitting in the front row.

The water boatman (Micronecta scholtzi), shown at the top left, is only 2mm long but is the loudest animal ever to be recorded, relative to its body size, outperforming all marine and terrestrial species. (Credit: Images courtesy of Dr. Jérôme Sueur, Muséum national d'Histoire naturelle, Paris)

The frequency of the sound (around 10 kHz) is within human hearing range and Dr. James Windmill of the University of Strathclyde, explains one clue as to how loud the animals are: "Remarkably, even though 99% of sound is lost when transferring from water to air, the song is so loud that a person walking along the bank can actually hear these tiny creatures singing from the bottom of the river."

The song, used by males to attract mates, is produced by rubbing two body parts together, in a process called stridulation. In water boatmen the area used for stridulation is only about 50 micrometres across, roughly the width of a human hair. "We really don't know how they make such a loud sound using such a small area," says Dr. Windmill.

The researchers, who are presenting their work at the Society for Experimental Biology Annual Conference in Glasgow, are now keen to bring together aspects of biology and engineering to clarify how and why such a small animal makes such a loud noise, and to explore the practical applications. Dr. Windmill explains: "Biologically this work could be helpful in conservation as recordings of insect sounds could be used to monitor biodiversity. From the engineering side it could be used to inform our work in acoustics, such as in sonar systems."

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Non-coding RNA has role in inherited neurological disorder -- and maybe other brain diseases too

Non-coding RNA has role in inherited neurological disorder -- and maybe other brain diseases too

  1:26:00 AM    Science Lover

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A team of scientists, led by researchers at the University of California, San Diego School of Medicine, have uncovered a novel mechanism regulating gene expression and transcription linked to Spinocerebellar ataxia 7, an inherited neurological disorder. The discovery promises to have broad ramifications, suggesting that abundant non-coding transcripts of ribonucleic acid (RNA) may be key players in neurological development and function, and could be powerful targets for future clinical therapies.

Researchers have discovered that expression
of the ataxin‑7 gene - the cause of the
neurological disorder spinocerebellar
ataxia type 7 - has two regulators: a highly
conserved, multi‑tasking protein called
CTCF and, surprisingly, an adjacent promoter
containing non‑coding RNA.
Credit: Illustration courtesy of Christina
Takamatsu‑Butler, UC San Diego.

The research, headed by Albert La Spada, MD, PhD, chief of the division of genetics in the UCSD department of pediatrics, and professor of cellular and molecular medicine, neurosciences and biological sciences, is published in the June 22 issue of the journal Neuron.

"Our paper highlights a number of important emerging themes in our understanding of gene regulation in the brain," said La Spada, who is also associate director of the UCSD Institute for Genomic Medicine.

"With the advent of new technologies, science has learned that the vast majority of our transcripts are non-coding," said La Spada. "The challenge going forward is to determine what they do do, and if they have specific functions. It now seems increasingly likely that a multitude of these non-coding RNAs help finely tune transcription regulation in the brain, and perturbation of their work is linked to disease. If we can figure out exactly how, we should be able to gain new insights into how the brain is so precisely regulated – knowledge that may help us better understand how the brain works."



Spinocerebellar ataxia 7 is one of several types of spinocerebellar ataxia (SCA), genetic degenerative disorders characterized by atrophy in the cerebellum of the brain, progressive loss of physical coordination – and in the case of type 7 – retinal degeneration that can result in blindness. There is currently no known cure.

Many SCAs are classified as polyglutamine diseases, caused when a protein associated with the disease contains too many repeats of the amino acid glutamine. Polyglutamine diseases are also known as "CAG Triplet Repeat Disorders" because CAG is the sequence of nucleic acids that codes for glutamine.

La Spada and colleagues have long studied SCA. In 2001, they were the first to demonstrate that SCA7 retinal degeneration was the result of transcription dysregulation of ataxin-7, the protein associated with SCA7. Following up, they decided to learn how the gene that expresses ataxin-7 is itself regulated.

The researchers found not one, but two, regulators. The first is called CTCF, a highly conserved protein that regulates a variety of transcriptional processes, most notable establishing insulator domains and controlling genomic imprinting. But they also discovered an adjacent, alternative promoter dubbed intron 2 promoter (P2A) and a transcribed antisense, non-coding RNA, which they labeled SpinoCerebellarAtaxia-AntisenseNoncodingTranscript1 or SCAANT1.

Antisense RNA is single-stranded ribonucleic acid whose primary function appears to be as an inhibitor or suppressor of a gene, though sometimes it can promote gene expression instead. Most antisense RNAs are non-coding, meaning that their sequences do not provide information for making proteins. Even though non-coding RNAs do not provide instructions for the production of vital proteins, they comprise the bulk of the human genome. A major challenge for biomedical research in the 21st century is to figure what they do, and how they do it.

In their Neuron paper, La Spada and colleagues highlight one function, at least for SCAANT1. When they investigated how CTCF regulated ataxin-7 gene expression in transgenic mice, they discovered that CTCF promotes the production of SCAANT1 which in turn represses the newly discovered ataxin-7 sense promoter P2A. In mice lacking SCAANT1, sense promoter P2A is de-repressed, allowing a mutant ataxin-7 gene to be expressed, resulting in mice with a version of SCA7. The scientists found a similar lack of antisense SCAANT1 in the fibroblasts and white blood cells taken from human patients with SCA7, implicating deregulation of this pathway in the disease process.

As many inherited neurological disorders are now known to exhibit such overlapping "bidirectional" transcription, the findings in SCA7 could shed light on similar abnormalities with non-coding RNA function in a number of brain diseases.

Provided by University of California - San Diego

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New Insect Repellant May Be Thousands of Times Stronger Than DEET

New Insect Repellant May Be Thousands of Times Stronger Than DEET

  9:00:00 PM    Science Lover

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Imagine an insect repellant that not only is thousands of times more effective than DEET -- the active ingredient in most commercial mosquito repellants -- but also works against all types of insects, including flies, moths and ants.

Anopheles mosquito, which can spread malaria. 
(Credit: © Kletr / Fotolia)

That possibility has been created by the discovery of a new class of insect repellant made in the laboratory of Vanderbilt Professor of Biological Sciences and Pharmacology Laurence Zwiebel and reported this week in the online Early Edition of the Proceedings of the National Academy of Sciences.

"It wasn't something we set out to find," said David Rinker, a graduate student who performed the study in collaboration with graduate student Gregory Pask and post-doctoral fellow Patrick Jones. "It was an anomaly that we noticed in our tests."

The tests were conducted as part of a major interdisciplinary research project to develop new ways to control the spread of malaria by disrupting a mosquito's sense of smell supported by the Grand Challenges in Global Health Initiative funded by the Foundation for the NIH through a grant from the Bill & Melinda Gates Foundation.

"It's too soon to determine whether this specific compound can act as the basis of a commercial product," Zwiebel cautioned. "But it is the first of its kind and, as such, can be used to develop other similar compounds that have characteristics appropriate for commercialization."

The discovery of this new class of repellant is based on insights that scientists have gained about the basic nature of the insect's sense of smell in the last few years. Although the mosquito's olfactory system is housed in its antennae, 10 years ago biologists thought that it worked in the same way at the molecular level as it does in mammals. A family of special proteins called odorant receptors, or ORs, sits on the surface of nerve cells in the nose of mammals and in the antennae of mosquitoes. When these receptors come into contact with smelly molecules, they trigger the nerves signaling the detection of specific odors.

In the last few years, however, scientists have been surprised to learn that the olfactory system of mosquitoes and other insects is fundamentally different. In the insect system, conventional ORs do not act autonomously. Instead, they form a complex with a unique co-receptor (called Orco) that is also required to detect odorant molecules. ORs are spread all over the antennae and each responds to a different odor. To function, however, each OR must be connected to an Orco.

"Think of an OR as a microphone that can detect a single frequency," Zwiebel said. "On her antenna the mosquito has dozens of types of these microphones, each tuned to a specific frequency. Orco acts as the switch in each microphone that tells the brain when there is a signal. When a mosquito smells an odor, the microphone tuned to that smell will turn "on" its Orco switch. The other microphones remain off. However, by stimulating Orco directly we can turn them all on at once. This would effectively overload the mosquito's sense of smell and shut down her ability to find blood."

Because the researchers couldn't predict what chemicals might modulate OR-Orco complexes, they decided to "throw the kitchen sink" at the problem. Through their affiliation with Vanderbilt's Institute of Chemical Biology, they gained access to Vanderbilt's high throughput screening facility, a technology intended for the drug discovery process, not for the screening of insect ORs.

Jones used genetic engineering techniques to insert mosquito odorant receptors into the human embryonic kidney cells used in the screening process. Rinker tested these cells against a commercial library of 118,000 small molecules normally used in drug development. They expected to find, and did find, a number of compounds that triggered a response in the conventional mosquito ORs they were screening, but they were surprised to find one compound that consistently triggered OR-Orco complexes, leading them to conclude that they had discovered the first molecule that directly stimulates the Orco co-receptor. They have named the compound VUAA1.

Although it is not an odorant molecule, the researchers determined that VUAA1 activates insect OR-Orco complexes in a manner similar to a typical odorant molecule. Jones also verified that mosquitoes respond to exposure to VUAA1, a crucial step in demonstrating that VUAA1 can affect a mosquito's behavior.

"If a compound like VUAA1 can activate every mosquito OR at once, then it could overwhelm the insect's sense of smell, creating a repellant effect akin to stepping onto an elevator with someone wearing too much perfume, except this would be far worse for the mosquito," Jones said.

The researchers have just begun behavioral studies with the compound. In preliminary tests with mosquitoes, they have found that VUAA1 is thousands of times more effective than DEET.

They have also established that the compound stimulates the OR-Orco complexes of flies, moths and ants. As a result, "VUAA1 opens the door for the development of an entirely new class of agents, which could be used not only to disrupt disease vectors, but also the nuisance insects in your backyard or the agricultural pests in your crops," Jones said.

Many questions must be answered before VUAA1 can be considered for commercial applications. Zwiebel's team is currently working with researchers in Vanderbilt's Drug Discovery Program to pare away the parts of VUAA1 that don't contribute to its activity. Once that is done, they will begin testing its toxicity.

Vanderbilt University has filed for a patent on this class of compounds and is talking with potential corporate licensees interested in incorporating them into commercial products, with special focus on development of products to reduce the spread of malaria in the developing world.

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Can Traumatic Memories Be Erased?

Can Traumatic Memories Be Erased?

  1:33:00 AM    Science Lover

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Could veterans of war, rape victims and other people who have seen horrific crimes someday have the traumatic memories that haunt them weakened in their brains? In a new study, UCLA life scientists report a discovery that may make the reduction of such memories a reality.

Could veterans of war, rape victims and other people 
who have seen horrific crimes someday have the traumatic 
memories that haunt them weakened in their brains? In a 
new study, UCLA life scientists report a discovery that may 
make the reduction of such memories a reality. "I think we 
will be able to alter memories someday to reduce the trauma 
from our brains," said the study's senior author. 
(Credit: © adimas / Fotolia)

"I think we will be able to alter memories someday to reduce the trauma from our brains," said the study's senior author, David Glanzman, a UCLA professor of integrative biology and physiology and of neurobiology.

The study appears in the April 27 issue of the Journal of Neuroscience.

Glanzman, a cellular neuroscientist, and his colleagues report that they have eliminated, or at least substantially weakened, a long-term memory in both the marine snail known as Aplysia and neurons in a Petri dish. The researchers say they gaining important insights into the cell biology of long-term memory.

They discovered that the long-term memory for sensitization in the marine snail can be erased by inhibiting the activity of a specific protein kinase -- a class of molecules that modifies proteins by chemically adding to them a phosphate (an inorganic chemical), which changes the proteins' structure and activity. The protein kinase is called PKM (protein kinase M), a member of the class known as protein kinase C (PKC), which is associated with memory.

The research has important potential implications for the treatment of post-traumatic stress disorder, as well as drug addiction, in which memory plays an important role, and perhaps Alzheimer's disease and other long-term memory disorders.

"Almost all the processes that are involved in memory in the snail also have been shown to be involved in memory in the brains of mammals," said Glanzman, who added that the human brain is far too complicated to study directly.

PKM is rare in that while most protein kinases have both a catalytic domain, which is the part of the molecule that does its work, and a regulatory domain, akin to an on-off switch that can be used by other signaling pathways to shut off the activity of the kinase, PKM has only the catalytic domain -- not the regulatory domain.

"This means that once PKM is formed, there is no way to shut it off," said Glanzman, who is a member of UCLA's Brain Research Institute. "Once it is activated, PKM's continual activity maintains a memory until PKM degrades."

Glanzman decided to study PKM in the marine snail, which has simple forms of learning and a simple nervous system, so that he could understand in precise detail how PKM's activity maintains a long-term memory, a process that is not well understood.

Glanzman and his colleagues -- researchers Diancai Cai, lead author of the study; Kaycey Pearce; and Shanping Chen, all of whom work in his laboratory -- studied a simple kind of memory called sensitization. If marine snails are attacked by a predator, the attack heightens their sensitivity to environmental stimuli -- a "fundamental form of learning that is necessary for survival and is very robust in the marine snail," Glanzman said.

"The advantage of Aplysia," he said, "is that we know the neurons that produce this reflex; we know where they are in the nervous system."

The scientists removed the key neurons from the snail's nervous system and put them in a Petri dish, thereby recreating in the dish the two-neuron "circuit" -- a sensory neuron and a motor neuron -- that produces the reflex.

"The point is to reduce the problem so we can study on a fundamental biological level how PKM is maintaining long-term memory," Glanzman said.

They succeeded in erasing a long-term memory, both in the snail itself and in the circuit in the dish. They are the first scientists to show that long-term memory can be erased at a connection between just two neurons.

"We found that if we inhibit PKM in the marine snail, we will erase the memory for long-term sensitization," Glanzman said. "In addition, we can erase the long-term change at a single synapse that underlies long-term memory in the snail."

The scientists administered electric shocks to the snails' tails. Following this training, when the scientists gently touched a snail's siphon (an organ in their mid-section used in respiration), the animal responded with a reflexive contraction that lasted about 50 seconds. A week later, when the scientists touched the siphon, the reflex still lasted 30 seconds or more, rather than just the second or two the reflex normally lasts without the shock training. This constituted a long-term memory.

Then, once the marine snail had formed the long-term memory, the scientists injected an inhibitor of PKM into the snail and 24 hours later touched the siphon; the marine snail responded as though it had never received the tail shocks, with a very brief contraction.

"The long-term memory is gone," Glanzman said.

Life scientists agree that learning is due to changes in the synaptic connections, some of which strengthen and some of which weaken, in the brain. This new research opens the door to learning how the changes in synaptic connections are maintained and what role PKM plays in this memory maintenance. Glanzman and his colleagues are now conducting detailed analyses.

During the long-term memory, new synaptic connections grow between the sensory neuron and the motor neuron. If the scientists inhibit PKM, will those synaptic connections disappear?

"We're going to study that," Glanzman said. "Now we can study the cell biology of how PKM maintains long-term memory. Once we know that, we may be able to alter long-term memories. This has implications for psychiatric disorders that are related to memory. Post-traumatic stress disorder is a hyper-induction of a long-term memory that won't go away."

Targeting specific memories

Is there a way to turn the traumatic memory down?

"This is the first step toward figuring that out," Glanzman said. "Even after we know this, we will still need a way to target the memory. We have captured the memory in the dish, but we also have to know where in the brain the memory is."

Does he think it will become possible to target and weaken specific traumatic memories?

"I do," Glanzman said. "Not in the immediate future, but I think we will be able to go into one's brain, identify the location of the memory of a traumatic experience and try to dampen it down. We can do this in culture, and there is no essential difference between the synapse in culture and the synapse in your brain. We have captured the memory in the dish; now we have to figure out a way to target the memories in human brains. Once we know the neural circuit that contains the memory, then we need a selective way to inhibit the activity of PKM in that circuit."

People have different brain circuits -- collections of neurons and synapses that join neurons -- for different memories, Glanzman believes. Scientists may seek to inhibit PKM in a particular circuit. The goal would be to find the brain circuit that is predominantly associated with a traumatic memory and target PKM in that circuit.

If you boost rather than inhibit PKM activity, might that have a beneficial affect for patients with Alzheimer's disease? Alzheimer's disease appears to initially disrupt the synaptic basis of learning, Glanzman said, and PKM might be involved in that disruption.

Just as scientists are seeking to target and kill cancer cells without damaging healthy cells, Glanzman intends to study whether it is possible to weaken only certain synapses associated with traumatic memories, while leaving other memories intact.

"The brain is the most complicated organ in the body," Glanzman said, noting that the brain has many trillions of synapses. "The research is complex, but this is the way we are going to understand how memories in our brains last a lifetime, or at least part of the way. It will take a lot of research, but I think it will be feasible."

Next steps include studying the relationship between PKM and the synapses and how the structure of synapses changes when PKM is inhibited.

"That is going to tell us how long-term memories are maintained," Glanzman said. "This is the first step. The more we know about how long-term memory is induced in the brain and how our memories are maintained in the brain, the more we are going to be able to treat long-term memory loss."

The experiments are very difficult, and Glanzman praised co-authors Cai, Pearce and Chen as "unbelievably skilled."

For 28 years, Glanzman has studied learning and memory in the marine snail, which is substantially larger than its garden variety counterpart and has approximately 20,000 neurons in its central nervous system; humans have approximately 1 trillion. However, the cellular and molecular processes seem to be very similar between the marine snail and humans.

"The fundamental mechanisms of learning and memory are identical, as far as we can tell," Glanzman said.

Glanzman's research is funded by a Senator Jacob Javits Award in the Neurosciences from the National Institute of Neurological Disorders and Stroke (NINDS) and by the National Institute of Mental Health.

The marine snail processes information about its environment and is capable of learning when an environment is safe and when it is not, learning to escape from predators, and learning to identify food. The marine snail is native to California, living in tidal waters off the coast.

Glanzman is also studying learning at the synaptic level in the zebra fish.

In earlier research, Glanzman's team identified a cellular mechanism in the Aplysia that plays an important role in learning and memory. A protein called the NMDA (N-methyl D-aspartate) receptor enhances the strength of synaptic connections in the nervous system and plays a vital role in memory and in certain kinds of learning in the mammalian brain as well. Glanzman's demonstration that the NMDA receptor plays a critical role in learning in the marine snail was entirely unexpected.

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Electron Microscopy: New Type of Genetic Tag Illuminates Life in Never-Before-Seen Detail

Electron Microscopy: New Type of Genetic Tag Illuminates Life in Never-Before-Seen Detail

  11:01:00 PM    Science Lover

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By modifying a protein from a plant that is much favored by science, researchers at the University of California, San Diego School of Medicine and colleagues have created a new type of genetic tag visible under an electron microscope, illuminating life in never-before-seen detail.

Confocal fluorescence images of miniSOG-targeted 
endoplasmic reticulum (A), Rab5a (B), zyxin (C), 
tubulin (D), ²-actin (E), ±-actinin (F), mitochondria
(G), and histone 2B (H) in HeLa cells; scale bars, 
10 µm. (Credit: Xiaokun Shu, Varda Lev-Ram, Thomas 
J. Deerinck, Yingchuan Qi, Ericka B. Ramko, Michael W. 
Davidson, Yishi Jin, Mark H. Ellisman, Roger Y. Tsien.
A Genetically Encoded Tag for Correlated Light and 
Electron Microscopy of Intact Cells, Tissues, and 
Organisms. PLoS Biology, 2011; 9 (4): e1001041 
DOI: 10.1371/journal.pbio.1001041)

Led by Nobel laureate Roger Tsien, PhD, Howard Hughes Medical Institute investigator and UCSD professor of pharmacology, chemistry and biochemistry, a team of scientists radically re-engineered a light-absorbing protein from the flowering cress plant Arabidopsis thaliana. When exposed to blue light, the altered protein produces abundant singlet oxygen, a form of molecular oxygen that can be made visible by electron microscopy (EM).

The findings are published in the online, open access journal PLoS Biology.

Tsien was co-winner of the 2008 Nobel Prize in chemistry for his role in helping develop and expand the use of green fluorescent protein (GFP), a protein from jellyfish that is now widely employed in light microscopy to peer inside living cells or whole animals and observe molecules interacting in real-time. Tsien said the development of the small, highly engineered Arabidopsis protein, dubbed "miniSOG," may elevate the abilities of electron microscopy in the same way that GFP and its relatives have made modern light microscopy in biological research much more powerful and useful.


"The big advantage of EM is that it has much higher spatial resolution than light microscopy. You can get up to a hundred-fold higher useful magnification from EM than from light microscopy," said Tsien. The result has been extraordinarily detailed, three-dimensional images of microscopic objects at resolutions measuring in the tens of nanometers, tiny enough to meticulously render the internal anatomy of individual cells. But current EM technologies do not distinguish or highlight individual proteins in these images. Although individual proteins can be tagged with GFP or other fluorescent proteins to aid localization by light microscopy, there has been no equivalent technology for the higher-resolution images provided by EM.

To create this ability, the scientists began with a protein from Arabidopsis that absorbs incoming blue light. It's normal function is to trigger biochemical signals that inform the plant how much sunlight it is receiving. "We rationally engineered the protein based on its atomic model so that it changes incoming blue light into a little bit of green fluorescence and a lot of singlet oxygen," said the paper's first author, Xiaokun Shu, now an assistant professor at UC San Francisco. Established methods were then used to convert singlet oxygen production into a tissue stain that the electron microscope can "see." The scientists tested the modified protein's utility as an EM marker by first using it to confirm the locations of several well-understood proteins in mammalian cells, nematodes and rodents, and then used miniSOG to successfully tag two neuronal proteins in mice whose locations had not been known.

Tsien is optimistic that miniSOG will grant new powers to electron microscopy, permitting scientists to pursue answers to questions previously impossible to ask. MiniSOG will especially be useful to scientists who investigate cellular and subcellular structures including neuronal circuits at nanometer resolution in multicellular organisms since previous methods have great difficulty in achieving both efficient labeling and good preservation of the structures under study. While EM can provide much higher useful magnification than light microscopy, EM will not replace light microscopy. "When we use miniSOG, we see the tagged proteins plus the landmarks that we are used to navigating by," said Tsien. "On the other hand, EM has the disadvantage that it gives a snapshot of cells before we killed them (to make the image), whereas light microscopy can show the dynamics in live cells. Each technique has different complementary strengths and weaknesses."

Co-authors of the paper include: Varda Lev-Ram, UCSD Department of Pharmacology; Thomas J. Deerinck, National Center for Microscopy and Imaging Research, Center for Research on Biological Systems, UCSD; Yingchuan Qi and Yishi Jin, Howard Hughes Medical Institute, UCSD and UCSD Division of Biological Science; Ericka B. Ramko and Michael W. Davidson, National High Magnetic Field Laboratory and Department of Biological Science, Florida State University; and Mark H. Ellisman, National Center for Microscopy and Imaging Research, Center for Research on Biological Systems, UCSD and UCSD Department of Neurosciences

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