21. MRI reveals genetic activity

MRI reveals genetic activity

Alan Jasanoff, an MIT associate professor of biological engineering and leader of the research team.

Doctors commonly use magnetic resonance imaging (MRI) to diagnose tumors, damage from stroke, and many other medical conditions. Neuroscientists also rely on it as a research tool for identifying parts of the brain that carry out different cognitive functions.

Now, a team of biological engineers at MIT is trying to adapt MRI to a much smaller scale, allowing researchers to visualize gene activity inside the brains of living animals. Tracking these genes with MRI would enable scientists to learn more about how the genes control processes such as forming memories and learning new skills, says Alan Jasanoff, an MIT associate professor of biological engineering and leader of the research team.

“The dream of molecular imaging is to provide information about the biology of intact organisms, at the molecule level,” says Jasanoff, who is also an associate member of MIT’s McGovern Institute for Brain Research. “The goal is to not have to chop up the brain, but instead to actually see things that are happening inside.”

To help reach that goal, Jasanoff and colleagues have developed a new way to image a “reporter gene” — an artificial gene that turns on or off to signal events in the body, much like an indicator light on a car’s dashboard. In the new study, the reporter gene encodes an enzyme that interacts with a magnetic contrast agent injected into the brain, making the agent visible with MRI. This approach, described in a recent issue of the journal Chemical Biology, allows researchers to determine when and where that reporter gene is turned on.

An on/off switch 

MRI uses magnetic fields and radio waves that interact with protons in the body to produce detailed images of the body’s interior. In brain studies, neuroscientists commonly use functional MRI to measure blood flow, which reveals which parts of the brain are active during a particular task. When scanning other organs, doctors sometimes use magnetic “contrast agents” to boost the visibility of certain tissues.

The new MIT approach includes a contrast agent called a manganese porphyrin and the new reporter gene, which codes for a genetically engineered enzyme that alters the electric charge on the contrast agent. Jasanoff and colleagues designed the contrast agent so that it is soluble in water and readily eliminated from the body, making it difficult to detect by MRI. However, when the engineered enzyme, known as SEAP, slices phosphate molecules from the manganese porphyrin, the contrast agent becomes insoluble and starts to accumulate in brain tissues, allowing it to be seen.

The natural version of SEAP is found in the placenta, but not in other tissues. By injecting a virus carrying the SEAP gene into the brain cells of mice, the researchers were able to incorporate the gene into the cells’ own genome. Brain cells then started producing the SEAP protein, which is secreted from the cells and can be anchored to their outer surfaces. That’s important, Jasanoff says, because it means that the contrast agent doesn’t have to penetrate the cells to interact with the enzyme.

Researchers can then find out where SEAP is active by injecting the MRI contrast agent, which spreads throughout the brain but accumulates only near cells producing the SEAP protein.

Exploring brain function

In this study, which was designed to test this general approach, the detection system revealed only whether the SEAP gene had been successfully incorporated into brain cells. However, in future studies, the researchers intend to engineer the SEAP gene so it is only active when a particular gene of interest is turned on. 

Jasanoff first plans to link the SEAP gene with so-called “early immediate genes,” which are necessary for brain plasticity — the weakening and strengthening of connections between neurons, which is essential to learning and memory. 

“As people who are interested in brain function, the top questions we want to address are about how brain function changes patterns of gene expression in the brain,” Jasanoff says. “We also imagine a future where we might turn the reporter enzyme on and off when it binds to neurotransmitters, so we can detect changes in neurotransmitter levels as well.”

Assaf Gilad, an assistant professor of radiology at Johns Hopkins University, says the MIT team has taken a “very creative approach” to developing noninvasive, real-time imaging of gene activity. “These kinds of genetically engineered reporters have the potential to revolutionize our understanding of many biological processes,” says Gilad, who was not involved in the study.

The research was funded by the Raymond and Beverly Sackler Foundation, the National Institutes of Health, and an MIT-Germany Seed Fund grant. The paper’s lead author is former MIT postdoc Gil Westmeyer; other authors are former MIT technical assistant Yelena Emer and Jutta Lintelmann of the German Research Center for Environmental Health.

20. Image of the Day

Image of the Day: Colorful Partnership

Xanthoria parientina, the fungal part of common orange lichen, is being sequenced as a model organism for lichen-forming fungus.

19. Scientists Synthesize First

Scientists Synthesize First Functional “Designer” Chromosome in Yeast

Study reports major advance in synthetic biology

https://www.youtube.com/watch?feature=player_embedded&v=Fo-LKmflRvo#t=0

An international team of scientists led by Jef Boeke, PhD, director of NYU Langone Medical Center’s Institute for Systems Genetics, has synthesized the first functional chromosome in yeast, an important step in the emerging field of synthetic biology, designing microorganisms to produce novel medicines, raw materials for food, and biofuels.

Over the last five years, scientists have built bacterial chromosomes and viral DNA, but this is the first report of an entire eukaryotic chromosome, the threadlike structure that carries genes in the nucleus of all plant and animal cells, built from scratch. Researchers say their team’s global effort also marks one of the most significant advances in yeast genetics since 1996, when scientists initially mapped out yeast’s entire DNA code, or genetic blueprint.

“Our research moves the needle in synthetic biology from theory to reality,” says Dr. Boeke, a pioneer in synthetic biology who recently joined NYU Langone from Johns Hopkins University.

“This work represents the biggest step yet in an international effort to construct the full genome of synthetic yeast,” says Dr. Boeke. “It is the most extensively altered chromosome ever built. But the milestone that really counts is integrating it into a living yeast cell. We have shown that yeast cells carrying this synthetic chromosome are remarkably normal. They behave almost identically to wild yeast cells, only they now possess new capabilities and can do things that wild yeast cannot.”

In this week’s issue of Science online March 27, the team reports how, using computer-aided design, they built a fully functioning chromosome, which they call synIII, and successfully incorporated it into brewer’s yeast, known scientifically as Saccharomyces cerevisiae.

The seven-year effort to construct synIII tied together some 273, 871 base pairs of DNA, shorter than its native yeast counterpart, which has 316,667 base pairs. Dr. Boeke and his team made more than 500 alterations to its genetic base, removing repeating sections of some 47,841 DNA base pairs, deemed unnecessary to chromosome reproduction and growth. Also removed was what is popularly termed junk DNA, including base pairs known not to encode for any particular proteins, and “jumping gene” segments known to randomly move around and introduce mutations. Other sets of base pairs were added or altered to enable researchers to tag DNA as synthetic or native, and to delete or move genes on synIII.

“When you change the genome you're gambling. One wrong change can kill the cell,” says Dr. Boeke. “We have made over 50,000 changes to the DNA code in the chromosome and our yeast still live. That is remarkable. It shows that our synthetic chromosome is hardy, and it endows the yeast with new properties.”

The Herculean effort was aided by some 60 undergraduate students enrolled in the “Build a Genome” project, founded by Dr. Boeke at Johns Hopkins. The students pieced together short snippets of the synthetic DNA into stretches of 750 to 1,000 base pairs or more. These pieces were then assembled into larger ones, which were swapped for native yeast DNA, an effort led by Srinivasan Chandrasegaran, PhD, a professor at Johns Hopkins. Chandrasegaran is also the senior investigator of the team’s studies on synIII.

Student participation kicked off what has become an international effort, called Sc2.0 for short, in which several academic researchers have partnered to reconstruct the entire yeast genome, including collaborators at universities in China, Australia, Singapore, the United Kingdom, and elsewhere in the U.S.

Yeast chromosome III was selected for synthesis because it is among the smallest of the 16 yeast chromosomes and controls how yeast cells mate and undergo genetic change. DNA comprises four letter-designated base macromolecules strung together in matching sets, or base pairs, in a pattern of repeating letters. “A” stands for adenine, paired with “T” for thymine; and “C” represents cysteine, paired with “G” for guanine. When stacked, these base pairs form a helical structure of DNA resembling a twisted ladder.

Yeast shares roughly a third of its 6,000 genes—functional units of chromosomal DNA for encoding proteins — with humans. The team was able to manipulate large sections of yeast DNA without compromising chromosomal viability and function using a so-called scrambling technique that allowed the scientists to shuffle genes like a deck of cards, where each gene is a card. “We can pull together any group of cards, shuffle the order and make millions and millions of different decks, all in one small tube of yeast,” Dr. Boeke says. “Now that we can shuffle the genomic deck, it will allow us to ask, can we make a deck of cards with a better hand for making yeast survive under any of a multitude of conditions, such as tolerating higher alcohol levels.”

Using the scrambling technique, researchers say they will be able to more quickly develop synthetic strains of yeast that could be used in the manufacture of rare medicines, such as artemisinin for malaria, or in the production of certain vaccines, including the vaccine for hepatitis B, which is derived from yeast. Synthetic yeast, they say, could also be used to bolster development of more efficient biofuels, such as alcohol, butanol, and biodiesel.

The study will also likely spur laboratory investigations into specific gene function and interactions between genes, adds Dr. Boeke, in an effort to understand how whole networks of genes specify individual biological behaviors.

Their initial success rebuilding a functioning chromosome will likely lead to the construction of other yeast chromosomes (yeast has a total of 16 chromosomes, compared to humans’ 23 pairs), and move genetic research one step closer to constructing the organism’s entire functioning genome, says Dr. Boeke.

Dr. Boeke says the international team’s next steps involve synthesizing larger yeast chromosomes, faster and cheaper. His team, with further support from Build a Genome students, is already working on assembling base pairs in chunks of more than 10,000 base pairs. They also plan studies of synIII where they scramble the chromosome, removing, duplicating, or changing gene order.

Detailing the Landmark Research Process

Before testing the scrambling technique, researchers first assessed synIII’s reproductive fitness,  comparing its growth and viability in its unscrambled from  — from a single cell to a colony of many cells — with that of native yeast III. Yeast proliferation was gauged under 19 different environmental conditions, including changes in temperature, acidity, and hydrogen peroxide, a DNA-damaging chemical. Growth rates remained the same for all but one condition.

Further tests of unscrambled synIII, involving some 30 different colonies after 125 cell divisions, showed that its genetic structure remained intact as it reproduced. According to Dr. Boeke, individual chromosome loss of one in a million cell divisions is normal as cells divide. Chromosome loss rates for synIII were only marginally higher than for native yeast III.

To test the scrambling technique, researchers successfully converted a non-mating cell with synIII to a cell that could mate by eliminating the gene that prevented it from mating.

Funding support for these experiments was provided by National Science Foundation, the National Institutes of Health, and Microsoft. Corresponding federal grant numbers are MCB-0718846 and GM-077291. Additional funding support was provided by fellowships from La Fondation pour la Recherche Médicale, Pasteur-Roux, National Sciences and Engineering Research Council of Canada, U.S. Department of Energy, and grants from the Exploratory Research Grant from the Maryland Stem Cell Research Fund and the Johns Hopkins University Applied Physics Laboratory.

Besides the teams at NYU Langone and Johns Hopkins, other scientific teams involved in the global Sc2.0 research effort are based at Loyola University in Baltimore, Md; BGI in Shenzhen, China; Tianjin University in China; Tsinghua University in China; MacQuarie University in Sydney, Australia; the Australian Wine Institute in Adelaide, Australia; the National University of Singapore; Imperial College, London, England; and the University of Edinburgh in Scotland.

18. BRCA1

BRCA1 Linked to Brain Size

The BRCA1 gene, most famous for its link to breast cancer risk, may also play a role in neural development and influence brain size, according to a study published this week (March 17) in PNAS. Researchers have long known that BRCA1-knockout mice die soon after birth, but did not fully understand why.

A team led by Inder Verma of the Salk Institute for Biological Studies in La Jolla, California, had previously seen that BRCA1 is highly expressed in the neuroectoderm, which is populated with neural stem cells. So the researchers engineered mice in which BRCA1 was knocked out in just the neural stem cells. They found that the brains of these mice were a third the size of normal mouse brains, and had especially small regions for learning, memory, motor control and sensation. Mature mutant mice had severe ataxia.

Upon closer inspection, the researchers found that neural stem cells were dying off at a high rate. They showed that BRCA1 prevented DNA breaks and that without it, excessive DNA damage triggered cell destruction. Brain cells that did survive looked like disorganized and deformed cancer cells.

“This is important fundamental basic science about how the genome is protected in rapidly proliferating cells in the brain,” Huda Zoghbi, a neuroscientist at Baylor College of Medicine in Houston, Texas, who was not involved with the study, told ScienceNOW.

17. Origins of Lactase

Origins of Lactase Persistence in Africa

Large-scale sequencing effort confirms several mutations that confer lactase persistence in Africans, while haplotype analysis sheds light on the trait’s origins.

Maasai man, who took part in the study, with goats in Tanzania

A study of lactase persistence in African populations led by researchers at the University of Pennsylvania bolsters geneticists’ knowledge of variants influencing the human ability to break down lactose, a disaccharide and the primary carbohydrate found in milk, into the monosaccharides glucose and galactose. The team’s work, published today (March 13) in The American Journal of Human Genetics, includes analyses of the genetic backgrounds of the variants as well as their geographical distribution to suggest potential pathways by which lactase persistence-associated single-nucleotide polymorphisms (SNPs) might have arisen in Africa.

“This is the largest study to date of the genetic basis of lactose tolerance across Africa,” said Penn’sSarah Tishkoff, who led the study.

“The work is interesting as many ethnic groups from Africa were studied and haplotypes were constructed that gave data about the migrations of the LP [lactase persistence] alleles,” Irma Järveläof the University of Helsinki, who was not involved in the work, told The Scientist in an e-mail.

Tishkoff and her colleagues identified three known variants—C-14010, G-13907, and G-13915—in people from diverse populations throughout Africa, which they confirmed were significantly associated with lactase persistence. They also suggested two new SNPs associated with lactase persistence, but because the potential SNPs are closely linked to the known lactase persistence-associated variants, the team has not yet been able to isolate their potential effects, Tishkoff said.

The continued expression of lactase beyond childhood enables humans to drink milk and reap its nutrients into adulthood. The enzyme is encoded by the gene LCT, found on human chromosome 2.LCT expression, scientists had previously found, is regulated by several genetic variants found within introns 9 and 13 of another gene, MCM6, which lies upstream. In adults of European ancestry, the SNP C/T-13910, within MCM6’s intron 13, is the primary genetic element behind lactase persistence. Other research groups had previously found that this region of intron 13 is a transcription factor binding site and enhancer of LCT expression. In Africa and the Middle East, however, there are pastoral populations who drink milk, yet lack T-13910, leading scientists to investigate whether some other variants conferred lactose tolerance in these groups.

With that question in mind, the researchers sequenced regions of introns 9 and 13 of MCM6 and about two kilobases of the LCT promoter in 819 individuals from 63 African populations as well as 154 people from Europe, Asia, and the Middle East. A subset of these study participants took a lactose tolerance test, which involved drinking a lactose solution and undergoing periodic blood glucose screens.

The researchers also determined the genetic background of each variant by genotyping four microsatellite repeats in the regions of interest and reconstructing the layouts of the individual chromosomes, or haplotypes, on which the variants lie. By comparing the frequencies of each variant in different geographic regions, both in Africa and in other parts of the world, they were able to hypothesize where the African lactase persistence-associated mutations originated.

“We were able to infer an East African origin for the C-14010 variant, a NE [Northeast] African origin for the G-13907 variant, and a Middle Eastern origin of the G-13915 variant,” wrote Penn’s Alessia Ranciaro in an e-mail to The Scientist. “We demonstrate that analysis of the geographic distribution of the LP variants are highly informative for reconstructing historic migration events and to trace the history of pastoralism in Africa.”

For example, the researchers found C-14010 in people from both Eastern and Southern Africa, leading them to speculate the variant may have spread southward throughout history.

While the researchers did identify the European variant T-13910 in North African populations, they did not find a significant link between the variant and lactose tolerance.

Dallas Swallow of University College London, who has also studied lactase persistence within African populations but was not involved in the present study, cautioned that it is still difficult to determine how lactase persistence-associated variants spread. 

“If you think about the frequency of a particular allele, it’s very tempting to assume that the place where that allele is found most frequently is the place where it originated. But the problem is that you don’t really know where the ancestors of those people who are living there now were living then,” she said. “One of the issues [that] is particularly relevant to pastoralist people is that they might move around a lot, they migrate.”

Swallow proposed combining sequencing data with computer models of gene spread and relevant historical information as another approach to tracing the spread of lactase persistence throughout history.

A. Ranciaro et al., “Genetic origins of lactase persistence and the spread of pastoralism in Africa,” The American Journal of Human Genetics, doi:10.1016/j.ajhg.2014.02.009, 2014.

16. Engineered Microbes Act as Sensors

Engineered Microbes Act as Sensors

Engineered probiotic E. coli colonize a mouse intestine.

Synthetic biologists programmed Escherichia coli to sense and record environmental stimuli, such as the antibiotic anhydrotetracycline,  in the mouse gastrointestinal tract. In their paper, published their work in PNAS this week (March 17), Harvard Medical School’s Pamela Silver and her colleagues wrote that their work provides proof of the principle “that E. coli can be engineered into living diagnostics capable of nondestructively probing the mammalian gut.”

Silver and her colleagues constructed a two-part genetic memory system, based on the phage lambda cI/Cro genetic switch, which they used to equip E. coli cells to sense and record exposure to anhydrotetracycline as they pass through the mouse gut.

“This is a really exciting advance,” MIT’s Chris Voigt, who was not involved in the work, told New Scientist. “This is the first use of a genetic circuit in a real environment. It is remarkable that they were able to engineer the cells to perform a computational operation—albeit a simple one—in this environment.”

The researchers are now working to construct additional switches to sense signs of toxicity or inflammation, for example. “This work lays a foundation for the use of synthetic genetic circuits as monitoring systems in complex, ill-defined environments, and may lead to the development of living diagnostics and therapeutics,” they wrote in PNAS.

15. Gut Microbes Gobble Cocoa

Gut Microbes Gobble Cocoa

Microbes like Bifidobacterium and lactic acid bacteria that reside in the human gut “feast on chocolate,” said Louisiana State University’s Maria Moore, an undergraduate student who assisted on a research that examined the effects of dark chocolate on gut bacteria, in a statementreleased at the American Chemical Society’s annual meeting being held in Dallas, Texas, this week. “When you eat dark chocolate, [these bacteria] grow and ferment it, producing compounds that are anti-inflammatory,” Moore added.

In in vitro experiments meant to model the human digestive tract, LSU food scientist John Finley’s team tested three cocoa powders, subjecting the substances to anaerobic fermentation using human fecal bacteria. The researchers found that, within the mock gut, the cocoa’s fiber content “is fermented and the large polyphenolic polymers are metabolized to smaller molecules, which are more easily absorbed,” Finley said in the statement. They also found that the smaller molecules “exhibit anti-inflammatory activity.” And gut microbes could help reduce a person’s feeling of hunger after having digested cocoa. “The microbes break down the fiber into short fatty chain acids, which get absorbed and can have an effect on satiety,” Finley told NPR’s The Salt blog.

The researchers have yet to confirm their results in vivo and cautioned that the apparent benefits of consuming cocoa powder don’t extend to gobbling dark chocolate bars from the corner store. “Our results don’t translate to a Hershey bar,” Finley told NPR. The British Heart Foundation’s Christopher Allen told BBC News that even though heart-healthy compounds like flavanols are found in dark chocolate, these molecules “are often destroyed by processing, and by the time a chocolate bar lands on the supermarket shelf it will also contain added extras such as sugar and fat.”

14. Bacteria’s Role in Bowel Cancer

Bacteria’s Role in Bowel Cancer

The development of serrated polyps depends on bacteria present in the gut, a mouse study shows.

Clostridium botulinum

Changes to the microbial composition of the gut can drastically alter the development of certain bowel tumors, according to a study published today (March 3) in The Journal of Experimental Medicine. Researchers from New York City’s Icahn School of Medicine at Mount Sinai worked with a mouse model that develops tumors called serrated polyps in the cecum, the part of the large intestine proximal to the colon. The polyps arise in part because the mice are genetically engineered, via a pair of transgenes, to overexpress the growth factor HB-EGF. But genetics, the researchers found, are not the whole story. Their work revealed that bacteria are also required for tumor development—the ceca of transgenic mice raised on an antibiotic cocktail did not form polyps.

“We were able to show that tumor formation was dependent on the microbiota present in that particular area of the intestine,” said Sergio Lira, who led the study. “In the presence of antibiotics, or of a slightly different cecum microbiota, the tumors did not develop.”

“This study adds to our knowledge of links between the gut microbiome and colon cancer, where causation is now established in several animal models and correlations are intriguing in humans (although causation in humans are not yet proven),” Rob Knight, a microbial ecologist at the University of Colorado, Boulder, who was not involved in the work, told The Scientist in an e-mail.

“There’s a growing body of information that constituents in the microbiota play a role in chronic inflammation and in cancer development,” said Martin Blaser, a professor of internal medicine and microbiology at the New York University School of Medicine, who did not participate in the study. “This study supplies yet another model of the same phenomenon.”

Since researchers determined that the bacterium Helicobacter pylori can cause some stomach cancers, a growing body of evidence has suggested that certain bacteria influence cancer development.

After finding that antibiotics prevented polyp formation, the researchers tried feeding the antibiotic-treated mice stool from their untreated counterparts to determine if bacteria alone could reverse the effects of the drugs. After ingesting the gut bacteria from the untreated mice, the once germ-free mice developed polyps.

The researchers also transplanted early embryos of the transgenic mice into females of another, cancer-free mouse strain, Swiss Webster. Inoculated at birth with the bacteria of their surrogate mothers, these transplanted mice did not develop tumors until 25 weeks, whereas the genetically identical controls had tumors by 12 weeks. This showed that small changes in the gut microbiota could have a large influence on tumor growth.

“This essentially suggests that if you have a genetic mutation,” Lira said, “the same genetic mutation in different individuals may have a different outcome.”

When the researchers examined the animals’ bowels, they found both that bacteria had invaded the intestinal epithelium and that the connections between the epithelial cells—as indicated by the presence of cell adhesion molecules, including E-cadherin—were weakened where the polyps had formed, compared with adjacent tissue. Most of the tumor-dwelling bacteria belonged to the Clostridiales family, Lira said. The researchers also observed an upregulation of inflammatory molecules near the polyps.

One outstanding question, Lira said, is how microbes affect the intestinal epithelium: Do the bacteria make it more permeable or just capitalize on its pre-existing weak spots?

“We need now to go back and do longitudinal experiments throughout the development of the tumors to try to understand what is causing the permeability changes that we have observed,” he said.

G. Bongers et al., “Interplay of host microbiota, genetic perturbations, and inflammation promotes local development of intestinal neoplasms in mice,” The Journal of Experimental Medicine, doi: 10.1084/jem.20131587, 2014.

13. Protein Protects Aging Brain

Protein Protects Aging Brain

Along with symptoms of cognitive decline, Alzheimer’s disease patients often have an accumulation of plaques and tangles of proteins in parts of their brains. But a long-standing question in neurology is why some elderly people develop dementia and others do not. Researchers are also left to wonder why some people have Alzheimer’s disease-like brain pathology yet show no cognitive symptoms.

A study published today (March 19) in Nature provides new clues that could help solve both puzzles, showing that a previously unknown stress response kicks in later in life to protect aging neurons. Researchers at Harvard Medical School found that a protein called REST, which is well characterized as a transcription factor that represses neuron-specific genes during embryogenesis, is switched on during middle- and late-adulthood, helping to protect neurons of the hippocampus and cortex from oxidative stress and the aggregated and misfolded proteins characteristic of Alzheimer’s and other neurodegenerative diseases.

“This work establishes REST as a regulator protein we have to pay a lot of attention to in the context of neurodegeneration,” said Susan Lindquist, a molecular biologist at the MIT Whitehead Institute for Biomedical Research, who was not involved in the study.

Analyzing brain samples from more than 300 individuals who had cognitive function ranging from normal to Alzheimer’s disease, the researchers showed that levels of REST were strongly correlated with measures of cognition. Many of the brain samples were from individuals who had died while taking part in a long-term clinical, pathological study and had undergone extensive neuropsychiatric assessments. Tissue from those individuals who retained robust cognitive function at the end of life yet also had Alzheimer’s-related plaques had three-fold higher levels of REST in their brains compared to those who had similar pathology but showed signs of dementia.

“This raises the possibility that the structural pathology may not be sufficient to cause Alzheimer’s disease,” said geneticist Bruce Yankner, who led the work. “A failure of the brain stress response, which REST might mediate, may also be required.”

REST first came to the researchers’ attention when it emerged among the most active genes in aging human cortices. Using neural cell lines, they next found that REST coordinates the activities of genes that may protect the brain from stress, upregulating the FOXO1 transcription factor—which is known to promote longevity—and binding and repressing cell death-related genes and those that mediate phosphorylation of tau and the generation of amyloid plaques in Alzheimer’s disease.

The researchers showed that while primary mouse neurons without REST were highly sensitive to hydrogen peroxide and other stressors, the REST protein could rescue the cell degeneration and cell death phenotype. Mice that lacked REST in their brains showed apoptosis and neurodegeneration in the hippocampus and cortex by their eighth month of life, but not earlier.

The researchers also identified an ortholog of REST in the nematode C. elegans, called SPR-4. SPR-4 mutants showed increased sensitivity to oxidative stress, which could be rescued by expression of either the human REST gene or wild-type SPR-4. When a spr-4 mutant was crossed with the C. elegans model that expresses amyloid-b, the resulting worms exhibited accelerated neurodegeneration. “That REST functions in C. elegans shows this system is likely deeply rooted in evolutionary biology,” said Lindquist.

Of course, there are many unanswered questions. Chiefly, whether the REST pathway is a master stress-regulator in neurons—analogous to the heat shock response in other cell types—remains to be seen.

Todd Golde, a neuroscientist and pathologist who studies neurodegenerative diseases at the University of Florida, said the study presents a compelling case that reduction of neuronal REST plays a role in the susceptibility of neurons to stress but cautioned that it doesn’t show how exactly the protein is involved in the pathological processes that lead to Alzheimer’s disease. “The unanswered question is whether it’s a key driver of the degenerative process or just one of many factors that contribute to the downstream degenerative process.”

Golde is also somewhat skeptical of the C. elegans and mouse model results. “The experiments in these model organisms show that REST has a role in stress and aging response and can regulate neuronal viability in these systems, but it doesn’t really inform on whether REST plays a neuroprotective role in the aging human brain as these are not very good models of human Alzheimer’s disease,” he said.

Yankner and his colleagues are now working on follow-up experiments to better understand whether REST is indeed a regulator of brain aging. One goal is to isolate the signals that turn REST on in the aging brain, which the researchers suggested could be linked to Wnt signaling. Another is to determine whether REST has a more general stress response function in the context of other neurodegenerative diseases, or in cases of traumatic brain injury or stroke.

The researchers are also screening compounds to target REST and other components of its defined chromatin-remodeling complex for drug development purposes.

“In addition to targeting abnormal proteins in the brain of patients with neurodegenerative diseases, our study suggests that boosting the physiological defense system may also to be important, and that a multipronged therapeutic approach might be beneficial in neurodegenerative disorders, analogous to current cancer therapies,” Yanker said.

T. Lu et al. “REST and stress resistance in ageing and Alzheimer’s disease,” Nature, doi:10.1038/nature13163, 2014.

12. DNA Mugshots

DNA Mugshots

DNA left at a crime scene could one day tell investigators what the perpetrator looked like. Researchers reported last week (March 20) in PLOS Genetics that they can make rough 3-D facial sketches based on 24 variants of 20 genes.

A team led by Mark Shriver of Pennsylvania State University mapped 7,000 anatomical points on 3-D models of the faces of 592 people of mixed European and West African ancestry in three regions (United States, Brazil, and Cape Verde). The researchers noted variations in distance between points and then hunted for variants in the genomes of study participants, focusing on genes associated with embryonic head development and facial deformity. They used two dozen genes to create a computer program that converts gene sequence data into a simulated facial model.

Predicting facial structure based on genes is much more complex than predicting hair or eye color, or skin pigment. “One thing we’re certain of: there’s no single gene that suddenly makes your nose big or small,” Kun Tang, a biologist who studies genes linked to facial structure at the Shanghai Institutes for Biological Sciences in China, who was not involved in this study, told Nature News.

Forensic use of the technology isn’t ready for prime time yet, but the researchers say it should be available in a matter of years. “I believe that in five to 10 years’ time, we will be able to computationally predict a face,” study coauthor Peter Claes of the University of Leuven told New Scientist.

11. Discovering Archaea

Discovering Archaea, 1977

FINGERPRINT: X-ray film “fingerprints” of digested small subunit ribosomal RNAs, such as this one annotated by Carl Woese, led Woese to the discovery of the three domains of life.

In a letter to Francis Crick dated June 24, 1969, Carl Woese, a microbiologist at the University of Illinois, wrote that he wanted to use “the cell’s ‘internal fossil record’”— specifically, the RNA of the cell’s translation machinery—“to extend our knowledge of evolution backward in time by a billion years or so.” Soon enough, Woese’s team had RNA sequencing up and running in the lab.

The protocol consisted of digestion and two-dimensional electrophoresis of radioactive small-subunit (16S or 18S) ribosomal RNA. When exposed to X-ray film, the separated fragments generated a unique fingerprint, which Woese interpreted based on the position of the spots.

“Literally every single day he sat in front of those fingerprints and analyzed them,” says George Fox, a postdoctoral fellow in the Woese lab from 1973 to 1977 and now a professor of biology and biochemistry at the University of Houston. After secondary and tertiary digestion of the spots, Woese and Fox determined the sequences of the oligonucleotide fragments—each about 6 to 14 nucleotides long—and recorded them on 80-column IBM punch cards. The team then compared the catalog of sequences from each organism using a computer program developed by Fox.

“Each one of those spots was a puzzle,” explains Fox. “If you do the same puzzle many times, you start to recognize it.” Initially, the researchers sequenced ribosomal RNA from readily available laboratory strains of bacteria, and Woese grew to expect the same spots that appeared over and over again on the film. But later, the lab’s ability to grow methanogens—microorganisms that produce methane and were not well-studied at the time—led to a eureka moment.

“When we did that first methanogen catalog, he started analyzing the data and all of a sudden the things he expected to find weren’t there,” says Fox. Woese immediately noticed that the fingerprints of two methanogen species, then referred to as bacteria, were missing two distinct spots. Upon secondary examination, these species also lacked fragments universal to previously examined bacteria. It became clear these organisms belonged to a distinct group.

In 1977, Woese and Fox published a landmark paper in PNAS containing just one table illustrating the relationships between the fragment catalogs of 13 species. The data revealed three distinct groups, which Woese and Fox described as “urkingdoms”: eubacteria, archaebacteria, and urkaryotes—their name for the presumed ancestor of the eukaryotes. Though some scientists balked at the idea of three urkingdoms replacing the conventional two, the three domains of life—archaea, bacteria, and eukarya—eventually became widely accepted.

With the more recent availability of vast amounts of genomic data, some now challenge the three-domain tree of life, and instead support a tree where bacteria and archaea are the sole domains, with eukaryotes branching from the archaea. But Woese’s three domains have not lost their stronghold in biology dogma just yet.

Fox wonders if the archaea would have been missed if a different technique had been employed. Fingerprinting gave “a much more black-and-white outcome than you would have if you were actually using modern sequencing,” he says. With the percent identity comparisons that scientists use today, “archaea would probably just be considered an odd niche of bacteria.”

10. Is Cannabis Really

Is Cannabis Really That Bad?

Though some studies point to negative consequences of pot use in adolescents, data on marijuana’s dangers are mixed.

Marijuana is a tricky drug, alternately demonized as a gateway drug and lionized for its medical promise. And while the juries remain out on both sides of the coin, one thing is clear: its use is on the rise. According to the US Department of Human Health and Services, the number of people in the United States who admit to smoking pot in the last month climbed from 14.4 million in 2007 to over 18 million in 2011.

This increase may in part be due to the lack of strong evidence supporting the suspected risks of cannabis use. Indeed, though marijuana smoke carries carcinogens and tar just as tobacco smoke does, definitive data linking marijuana to lung damage is lacking. And a recent long-term study that seemed to conclusively link chronic marijuana initiated in adolescence to a lowered IQ in New Zealanders was quickly challenged by a counter-analysis that pointed to socioeconomic status as a confounding factor. According to survey data from the Centers for Disease Control and Prevention, cannabis use increases in teenagers as marijuana’s perceived risks decline, and researchers—and undoubtedly some parents—are anxious to get to the bottom of the matter.

Take a deep breath

In 2012, a study at the University of California, San Francisco (UCSF) calculated that even smoking a single joint every day for 20 years might be benign, though most participants only smoked two or three joints each month. “I was surprised we didn’t see effects [of marijuana use],” said UCSF epidemiologist Mark Pletcher, who led the study.

One assessment of various epidemiological studies points to small sample size and poor study design as reasons for scientists’ inability to nail down a link between cannabis and cancer risk. But some suspect that such a link doesn’t exist, and that marijuana may even have cancer-preventive effects. A 2008 study, for example, suggested that smoking marijuana may reduce the risk of tobacco-associated lung cancer, calculating that people who smoke both marijuana and tobacco have a lower risk of cancer than those who smoke only tobacco (though still a higher risk than non-smokers).

But even Pletcher isn’t sanguine about marijuana’s effects on the lungs, and suspects that there may still be long-term lung damage that can be hard to detect. “We really can’t reassure ourselves about heavy use,” he explained.

Your brain on drugs

There is some evidence to suggest that stoned subjects exhibit increased risk-taking and impaired decision-making, and score worse on memory tasks—and residual impairments have been detected days or even weeks after use. Some studies also link years of regular marijuana use to deficits in memory, learning, and concentration. A recent and widely discussed report on the IQs of New Zealanders followed since birth found that cannabis users who’d started their habit in adolescence had lower IQs than non-users.

In this study, led by researchers at Duke University, “you could clearly see as a consequence of cannabis use, IQ goes down,” said Derik Hermann, a clinical neuroscientist at the Central Institute of Mental Health in Germany who was not involved in the research.

But not 4 months later, a re-analysis and computer simulation at the Ragnar Frisch Center for Economic Research in Oslo countered the Duke findings. Ole Rogeberg contended that socioeconomic factors, not marijuana use, contributed to the lower IQs seen in cannabis users.

Rogeberg’s conclusion counters a sizeable literature, however, which supports a link between pot use and neurophysiological decline. Studies in both humans and animals suggest that people who acquiring a marijuana habit in adolescence face long-term negative impacts on brain function, with some users finding it difficult to concentrate and learn new tasks.

Notably, most studies on the subject suggest that while there may be negative consequences of smoking as a teen, users who begin in adulthood are generally unaffected. This may be due to endocannabinoid-directed reorganization of the brain during puberty, Hermann explained. The intake of cannabinoids that comes with pot use may cause irreversible “misleading of the neural growth,” he said.

In addition to the consequences for intelligence, many studies suggest that smoking marijuana raises the risk of schizophrenia, and may have similar effects on the brain. Hermann’s group used MRI to detect cannabis-associated neuron damage in the pre-frontal cortex and found that it was similar to brain changes seen in schizophrenia patients. Other studies further suggest that weed-smoking schizophrenics have greater disease-associated brain changes and perform worse on cognitive tests than their non-smoking counterparts.

But much of this research can’t distinguish between brain changes resulting from marijuana use and symptoms associated with the disease. It’s possible that cannabis-smoking schizophrenics “might have unpleasant symptoms [that precede full-blown schizophrenia] and are self-medicating” with the psychotropic drug, said Roland Lamarine, a professor of community health at California State University, Chico. “We haven’t seen an increase in schizophrenics, even with a lot more marijuana use.”

In fact, other research suggests that cannabis-using schizophrenics score better on cognitive tests than non-using schizophrenics. Such conflicting reports may be due to the varying concentrations—and varying effects—of cannabinoids in marijuana. In addition to tetrahydrocannabinol (THC), a neurotoxic cannabinoid that is responsible for marijuana’s mind-altering properties, the drug also contains a variety of non-psychoactive cannabinoids, including cannabidiol (CBD), which can protect against neuron damage. Hermann found that the volume of the hippocampus—a brain area important for memory processing—is slightly smaller in cannabis users than in non-users, but more CBD-rich marijuana countered this effect.

A deadly cocktail?

While data supporting the harmful effects of marijuana on its own are weak, some researchers are more worried about the drug in conjunction with other substances, such as tobacco, alcohol, or cocaine. Some studies suggest, for example, that marijuana may increase cravings for other drugs, leading to its infamous tag as a “gateway drug.” A study published earlier this month supported this theory when it found that, at least in rats, THC exposure increases tobacco’s addictive effects. Furthermore, marijuana may not mix well with prescription drugs, as cannabis causes the liver to metabolize drugs more slowly, raising the risk of drug toxicity.  

Despite these concerns, however, Lamarine thinks it’s unlikely that the consequences of cannabis use are dire, given the amount of research that has focused on the subject. “We’re not going to wake up tomorrow to the big discovery that marijuana causes major brain damage,” he said. “We would have seen that by now.”

9. New Approach to Killing Cancer

New Approach to Killing Cancer

Researchers have found that a drug called Vacquinol-1 causes cancer cells to burst open, an achievement that represents “an entirely new mechanism” of killing cancer, according to the authors of the study, published in Cell this week (March 20). The drug prompted cells to over-develop vacuoles, become perfectly round, break apart, and die. “They have shown very exciting results,” said Ravi Bellamkonda, a neuroscientist at the Georgia Institute of Technology, in an e-mail to The Verge.

Mice with glioblastoma—a tough-to-treat brain cancer in humans—that were given Vacquinol-1 orally for five days ended up with either smaller tumors than control mice or no tumors at all. Mice that were treated with the drug also lived longer than control mice; only two of eight mice given Vacquinol-1 died over the course of 80 days, whereas the median survival of the eight control mice was only 31 days.

The results are enticing, given that the average survival time for a human with glioblastoma is only 15 months. “We now want to try to take this discovery in basic research through preclinical development and all the way to the clinic,” Patrik Ernfors, a coauthor of the study and a researcher at the Karolinska Institute, said in a press release. “The goal is to get into a phase 1 trial.”

Bellamkonda pointed out that the levels of Vacquinol-1 used were “relatively high” and it's unclear whether side effects might emerge. “This said, enhancing survival by several fold in aggressive tumor models is encouraging,” he told The Verge. “I'd love to see more studies.”

8. Enhancer and Promoter Atlases

Enhancer and Promoter Atlases

Consortium annotates the human genome with cell type-specific information about transcription start sites, active enhancers, and their expression throughout the body.

Researchers at Japan’s RIKEN institute, in collaboration with scientists worldwide, have produced two atlases of genetic regulatory elements throughout the human genome, as reported in a pair of papers published today (March 26) in Nature. The first paper presents an atlas of transcription start sites, where RNA polymerase begins to transcribe DNA into RNA; the second maps active enhancers, non-promoter stretches of DNA that upregulate the transcription of certain genes. Sixteen additional papers related to this work—results from the fifth edition of the Functional Annotation of the Mammalian Genome (FANTOM) project—are also today being published in other journals, including Blood and BMC Genomics.

“Both papers are very significant,” said biochemist Wei Wang from the University of California, San Diego, who was not involved in the work. “This will be a very valuable resource for the community.”

“We made an encyclopedia of the definition of the normal cell: 185,000 promoters, 44,000 enhancers, and the majority of them are tissue-specific,” said the RIKEN Omics Science Center’s Yoshihide Hayashizaki, who led the promoter annotation project.

“This is a very broad survey of transcriptional activity in diverse cell types, [making it] a very valuable resource, and currently, quite unique,” said Zhiping Weng from the University of Massachusetts Medical School, who was not involved in the work. Weng noted that the only comparable resource is the Genotype-Tissue Expression Program (GTEX), which when compared with FANTOM, is “not nearly as comprehensive at this point,” she said. “Right now, this is the most comprehensive, extensive collection of transcription data available, especially in primary cell types. I find that to be very significant. I think a lot of people are going to find the data to be highly useful.”

FANTOM is one of several projects that aim to annotate the human genome and to determine how the expression of its genes can produce a variety of cell types. Members of the Encyclopedia of DNA Elements (ENCODE) project, in which some RIKEN researchers took part, used chromatin immunoprecipitation analyses and mapped DNase hypersensitive sites, among other things, to determine where transcription factors bind DNA and where chromatin is “open,” and therefore vulnerable to cleavage by DNAse. The ENCODE team used many cell lines and examined only a few cell types, whereas the FANTOM group studied myriad primary cell and tissue types, as well as cell lines.

“I see FANTOM and ENCODE being very complementary, because FANTOM mainly generates transcription data, and ENCODE generates a much wider diversity, much more types of data. But FANTOM has a huge representation in the cell type dimension, while ENCODE is primarily focused on cell lines and only a few types of primary cells and tissue types,” said Weng, who was part of ENCODE. “You can imagine two very big projects—they are very extensive in different dimensions.”

To create these atlases, the FANTOM researchers used cap analysis of gene expression (CAGE) to sequence the beginnings of RNA transcripts. By mapping CAGE tags onto the human genome, the RIKEN-led team identified the promoter regions upstream of the transcription start sites. The researchers used CAGE to identify promoters in human primary cells, as well as in tissue samples and immortalized cell lines. They found that many genes have multiple transcription start sites and that transcription begins at different locations in different cell types.  

Using CAGE, the team also identified the RNA sequences transcribed from enhancers. Other groups had previously shown that some enhancers are transcribed bidirectionally—from the center, outward in both directions, and from both DNA strands. The FANTOM team found evidence to suggest that bidirectional transcripts are signatures of active enhancers. About 75 percent of the enhancers detected by CAGE drove expression of a reporter gene in HeLa cells, a larger percentage than the untranscribed enhancers previously identified through ENCODE.

Wang said he was most interested in the enhancer atlas. “This was the very first time people have done this enhancer RNA [analysis] on such a large scale,” he said.

“Over so many cell types, this number [of active enhancers] is kind of at the low end,” Wang noted, “particularly if you compare with ENCODE and other annotations. . . . I think the reason is related to the low abundance of eRNAs [enhancer RNAs].”

Other groups had also found that enhancers produce RNA in low amounts. “My only concern is they probably missed a lot of active enhancers,” said Wang. “My understanding is they have both [a] high true-positive rate and also false-negative rate. So whatever they identified, I believe those are real, active enhancers, but they may also miss many active enhancers because [of] this low abundance of enhancer RNA.”                               

In the future, Hayashizaki said, knowledge of the enhancer and promoter usages that define different cell types raises the possibility of turning one cell type into another. It could also aid in predicting whether or not a particular cancer is going to metastasize, he added.

The FANTOM Consortium and the RIKEN PMI [Preventative Medicine and Diagnosis Innovation Program] and CLST [Center For Life Science Technologies] (DGT [Division of Genomic Technologies]), “A promoter-level mammalian expression atlas,” Nature, doi:10.1038/nature13182, 2014.

R. Andersson, et al., “An atlas of active enhancers across human cell types and tissues,” Nature, doi:10.1038/nature12787, 2014.

7. Close up of Crick and Watson's model

Close up of Crick and Watson's model

Close up of Crick and Watson's model of DNA from the Science Museum.