Tag Archives: Unviersity of Chicago

Minimalist DNA nanodevices perform bio-analytical chemistry inside live cells

A comparison of minimalist versus baroque architecture is one of the more startling elements in this March 24, 2016 news item on Nanowerk about a scientist working with DNA (deoxyribonucleic acid) nanodevices,

Some biochemistry laboratories fashion proteins into complex shapes, constructing the DNA nanotechnological equivalent of Baroque or Rococo architecture. Yamuna Krishnan, however, prefers structurally minimalist devices.

“Our lab’s philosophy is one of minimalist design,” said Krishnan, a professor of chemistry at the University of Chicago. “It borders on brutalist. Functional with zero bells and whistles. There are several labs that design DNA into wonderful shapes, but inside a living system, you need as little DNA as possible to get the job done.”

That job is to act as drug-delivery capsules or as biomedical diagnostic tools.

A March 24, 2016 University of Chicago news release by Steve Koppes, which originated the news item, provides some background information before launching into the latest news,

In 2011, Krishnan and her group, then at the National Centre for Biological Sciences in Bangalore, India, became the first to demonstrate the functioning of a DNA nanomachine inside a living organism. This nanomachine, called I-switch, measured subcellular pH with a high degree of accuracy. Since 2011, Krishnan and her team have developed a palette of pH sensors, each keyed to the pH of the target organelle.

Last summer, the team reported another achievement: the development of a DNA nanosensor that can measure the physiological concentration of chloride with a high degree of accuracy.

“Yamuna Krishnan is one of the leading practitioners of biologically oriented DNA nanotechnology,” said Nadrian Seeman, the father of the field and the Margaret and Herman Sokol Professor of Chemistry at New York University. “These types of intracellular sensors are unique to my knowledge, and represent a major advance for the field of DNA nanotechnology.”

Chloride sensor

Chloride is the single most abundant, soluble, negatively charged molecule in the body. And yet until the Krishnan group introduced its chloride sensor—called Clensor—there was no effective and practical way to measure intracellular stores of chloride.

“What is especially interesting about this sensor is that it is completely pH independent,” Seeman said, a significant departure from Krishnan’s previous scheme. “She spent a number of years developing pH sensors that work intra-cellularly and provide a fluorescent signal as a consequence of a shift in pH.”

The ability to record chloride concentrations is important for many reasons. Chloride plays an important role in neurobiology, for example. But calcium and sodium—both positively charged ions—tend to grab most of the neurobiological glory because of their role in neuron excitation.

“But if you want your neuron to fire again, you have to bring it back to its normal state. You have to stop it firing,” Krishnan said. This is called “neuronal inhibition,” which chloride does.

“It’s important in order to reset your neuron for a second round of firing, otherwise we would all be able to use our brains only once,” she said.

Under normal circumstances, the transport of chloride ions helps the body produce thin, freely flowing mucus. But a genetic defect results in a life-threatening disease: cystic fibrosis. Clensor’s capacity to measure and visualize protein activity of molecules like the one related to cystic fibrosis transmembrane could lead to high-throughput assays to screen for chemicals that would restore normal functioning of the chloride channel.

Nine diseases

“One could use this to look at chloride ion channel activity in a variety of diseases,” Krishnan said. “Humans have nine chloride ion channels, and the mutation of each of these channels results in nine different diseases.” Among them are osteopetrosis, deafness, muscular dystrophy and Best’s macular dystrophy.

The pH-sensing capabilities of the I-switch, meanwhile, are important because cells contain multiple organelles that maintain specific values of acidity. Cells need these different microenvironments to carry out specialized chemical reactions.

“Each subcellular organelle has a specific resting value of acidity, and that acidity is crucial to its function,” Krishnan said. “When the pH is not the value that it’s meant to be, it results in a range of different diseases.”

There are 70 rare diseases called lysosomal storage disorders, which are progressive and often fatal. Each one—including Batten disease, Niemann-Pick disease, Pompe disease and Tay-Sachs disease—represents a different way a lysosome can go bad. She likened a defective lysosome to a garbage bin that never gets emptied.

“The lysosome is basically responsible for chewing up all the garbage and making sure it’s either reused or got rid of. It’s the most acidic organelle in the cell.” And that acidity is crucial for the degradation process.

Although there are 70 lysosomal storage diseases, small molecule drugs are available for only a few of them. These existing treatments—enzyme-replacement therapies—are expensive and are only palliative treatments. One goal of Krishnan’s group is to demonstrate the utility of their pH sensors to discover new biological insights into these diseases. Developing small molecule drugs—which are structurally simpler and easier to manufacture than traditional biological drugs—could help significantly.

“If we can do this for one or two lysosomal diseases, there’ll be hope for the other 68,” Krishnan said.

Here are links to and citations for the 2015 and 2011 papers,

A pH-independent DNA nanodevice for quantifying chloride transport in organelles of living cells by Sonali Saha, Ved Prakash, Saheli Halder, Kasturi Chakraborty, & Yamuna Krishnan. Nature Nanotechnology 10, 645–651 (2015)  doi:10.1038/nnano.2015.130 Published online 22 June 2015

An autonomous DNA nanomachine maps spatiotemporal pH changes in a multicellular living organism by Sunaina Surana, Jaffar M. Bhat, Sandhya P. Koushika, & Yamuna Krishnan. Nature Communications 2, Article number: 340  doi:10.1038/ncomms1340 Published 07 June 2011

The 2015 paper is behind a paywall but the 2011 paper is open access.

When twice as much (algebra) is good for you

“We find positive and substantial longer-run impacts of double-dose algebra on college entrance exam scores, high school graduation rates and college enrollment rates, suggesting that the policy had significant benefits that were not easily observable in the first couple of years of its existence,” wrote the article’s authors.

The Mar. 21, 2013 news release on EurekAlert which includes the preceding quote recounts an extraordinary story about an approach to teaching algebra that was not enthusiastically adopted at first but first some reason administrators and teachers persisted with it. Chelsey Leu’s Mar. 21, 2013 article (which originated the news release) for UChicago (University of Chicago) News (Note: Links have been removed),

Martin Gartzman sat in his dentist’s waiting room last fall when he read a study in Education Next that nearly brought him to tears.

A decade ago, in his former position as chief math and science officer for Chicago Public Schools [CPS], Gartzman spearheaded an attempt to decrease ninth-grade algebra failure rates, an issue he calls “an incredibly vexing problem.” His idea was to provide extra time for struggling students by having them take two consecutive periods of algebra.

In high schools, ninth-grade algebra is typically the class with the highest failure rate. This presents a barrier to graduation, because high schools usually require three to four years of math to graduate.

Students have about a 20 percent chance of passing the next math level if they don’t first pass algebra, Gartzman said, versus 80 percent for those who do pass. The data are clear: If students fail ninth-grade algebra, the likelihood of passing later years of math, and ultimately of graduating, is slim

Gartzman’s work to decrease algebra failure rates at CPS was motivated by a study of Melissa Roderick, the Hermon Dunlap Smith Professor at UChicago’s School of Social Service Administration. The study emphasized the importance of keeping students academically on track in their freshman year to increase the graduation rate.

Some administrators and teachers resisted the new policy. Teachers called these sessions “double-period hell” because they gathered, in one class, the most unmotivated students who had the biggest problems with math.

Principals and counselors sometimes saw the double periods as punishment for the students, depriving them of courses they may have enjoyed taking and replacing them with courses they disliked.

It seemed to Gartzman that double-period students were learning more math, though he had no supporting data. He gauged students’ progress by class grades, not by standardized tests. The CPS educators had no way of fully assessing their double-period idea. All they knew was that failure rates didn’t budge.

Unfortunately, Leu does not explain why the administrators and teachers continued with the program but it’s a good thing they did (Note: Links have been removed),

“Double-dosing had an immediate impact on student performance in algebra, increasing the proportion of students earning at least a B by 9.4 percentage points, or more than 65 percent,” noted the Education Next article. Although ninth-grade algebra passing rates remained mostly unaffected, “The mean GPA across all math courses taken after freshman year increased by 0.14 grade points on a 4.0 scale.”

They also found significantly increased graduation rates. The researchers concluded on an encouraging note: “Although the intervention was not particularly effective for the average affected student, the fact that it improved high school graduation and college enrollment rates for even a subset of low-performing and at-risk students is extraordinarily promising when targeted at the appropriate students.” [emphasis mine]

Gartzman recalled that reading the article “was mind-blowing for me. I had no idea that the researchers were continuing to study these kids.”

The study had followed a set of students from eighth grade through graduation, while Gartzman’s team could only follow them for a year after the program began. The improvements appeared five years after launching double-dose algebra, hiding them from the CPS team, which had focused on short-term student performance. [emphasis mine]

Gartzman stressed the importance of education policy research. “Nomi and Allensworth did some really sophisticated modeling that only researchers could do, that school districts really can’t do. It validates school districts all over the country who had been investing in double-period strategies.”

I’m not sure I understand the numbers very well (maybe I need a double-dose of numbers). The 9.4% increase for students earning a B sounds good but a mean increase of 0.14 in grade points doesn’t sound as impressive. As for the bit about the program being “not particularly effective for the average affected student,” what kind of student is helped by this program? As for the improvements being seen five years after the program launch. does this mean that students in the program showed improvement five years later (in first year university) or that researchers weren’t able to effectively measure any impact in the grade nine classroom until five years after the program began?

Regardless, it seems there is an improvement and having suffered through my share algebra classes, I applaud the educators for finding a way to help some students, if not all.