engineer biological

Stanford: Researchers engineer biological "devices" to program cells

Researchers engineer biological "devices" to program cells

Stanford bioengineer Christina Smolke, PhD, and colleagues have built biological "devices" capable of sensing disease states in cultured human cells and fine-tuning their own functions in response to the cell's internal signals, according to research recently published in Science. An article published today in the Stanford Report further describes the design and functionality of the new tools, which are called "sensor-actuator" devices:

Stanford researcher's engineered molecule changes itself to detect and attack diseased cells

Assistant Professor of Bioengineering Christina Smolke has engineered biological molecules that regulate a cell's behavior by adjusting their own forms and functions in response to the internal conditions of the cell. These tools can be used to facilitate medical research and biotechnology today and could one day be used as diagnostic and therapeutic aides.
L.A. Cicero
Christina Smolke

BY SUSAN YOUNG
Imagine if your doctor could look for cancer in your body just by checking for green glowing cells, alerting her to the presence of the disease. Imagine further that she could convince any cancerous cells in your body to commit suicide, while leaving your healthy cells unaffected.
In Friday's issue of Science, a Stanford researcher reported engineered biological "devices" that could one day offer these kinds of diagnostic and treatment options. The devices built by Assistant Professor of Bioengineering Christina Smolke, along with a graduate student and a postdoctoral researcher, can sense disease states in cultured human cells and fine-tune their own functions in response to a cell's internal signals.

These autonomous biological tools are called "sensor-actuator" devices because they sense what's happening in a cell and act upon what they detect.

The researchers built these devices by combining different pieces of DNA into one long stretch. The DNA is then put into cells that convert it to RNA, a slightly different version of genetic material that is frequently made by cells. The RNA molecule can then be read like a recipe by the cell's protein-making molecular machinery.

The sensor-actuator devices are built with efficient redesign in mind. Each piece of the device, whether the sensor or the protein-recipe actuator, can be swapped out for another version. This way, researchers can conveniently build a device to fit their particular needs. "You can fan out with lots of different outputs and you have lots of different inputs you could potentially link into," said Smolke. The input could be any number of protein signals inside a cell and the output could be instructions for the cell to create a molecule that's easily detected by a researcher – as in the case of the green-glowing cancer cells.
Or the output could cause a diseased cell to kill itself.

The sensor part of the RNA molecule can detect whether a certain protein is present simply by binding to it. The proteins these devices detect are chemical messengers, communicating information gathered inside and outside of the cell into the nucleus, which acts like the cell's control center.

Smolke and her team used the molecular devices to sense disease-like states, such as inflammation and cancer, in cultured human cells. "We have a lot of these different signaling pathways in our cells and many diseases are associated with mistaken signaling through these pathways," said Smolke.
The RNA sensor-actuator devices can "listen in" on the messages communicated by the cell and act accordingly. Depending on whether or not the device binds to the input protein, the RNA molecule could keep its original structure, or cut out a piece of itself and thus change the genetic information it contains.

When the RNA is read by the cell's protein-making machinery, the final product will depend on the RNA's information content.
Powerful tool for cells
The process by which the RNA device can remove part of itself is called "alternative splicing." Alternative splicing is an everyday process for many cells and is a powerful way to generate a diverse array of proteins inside a cell.

In the sensor-actuator devices described in the study, the optional piece of the RNA that could be cut out contained a "stop" message that instructed cells to stop making a protein before it was complete. When this "stop"-containing piece was removed, the device produced instructions for a whole and functional protein, one that, for instance, could glow green. In this way, the device could alter its output based upon the state of the cell.

"This is the first time a sensor-actuation device has been developed to respond to protein inputs and control an alternative splicing event linked to gene expression," said Smolke.
"With the application of this device, you encode a certain level of intelligence that allows it to go into the cell and first assess whether the cell is diseased or not based upon disease markers. If yes, then it can then specifically activate therapeutic effects in that cell."
One such therapeutic effect is the ability to specifically kill diseased cells. The researchers engineered an actuator module with an output that converted an inactive drug into an active form that causes cells to die. The sensor-actuator device only made the drug-activating output protein when the cell was diseased. Otherwise, the "stop" signal was left in the device and acted like a safety trigger preventing the death of healthy cells.

But the power of alternative splicing is not limited to just functional and non-functional outputs. "Instead of just yes/no, alternative splicing could modulate function," said Smolke. Proteins could be modified to have slightly different functions in response to different cell states. "There's a lot of richness in alternative splicing that could be used to develop more complex genetic circuits, beyond the demonstrated examples, that we might begin to implement in human cells," she said.

Smolke began the study at Caltech, where she was an assistant professor of chemical engineering. She moved to Stanford mid-project in 2009, where she completed much of the data analysis. Caltech student Stephanie Culler and postdoctoral researcher Kevin Hoff also contributed to the report.

The study was funded by the Caltech Joseph Jacobs Institute for Molecular Engineering for Medicine, the National Institutes of Health, the U.S. Department of Defense, the Alfred P. Sloan Foundation and the Bill and Melinda Gates Foundation.
Susan Young is a science-writing intern at the Stanford News Service

Adult stem AIDS

Adult stem cell therapy cures AIDS

Even as Barack Obama prepares to reward his liberal university friends by opening public coffers to fund new labs fronted by futile and morally bereft human embryo experimentation comes this news from Germany, as reported by the Wall Street Journal and Reuters:

The startling case of an Aids patient who was cured after undergoing a bone marrow transplant to treat leukemia is stirring new hope that researchers might someday find a cure for Aids....

The patient, a 42-year-old American living in Berlin, is still recovering from his leukemia therapy, but he appears to have won his battle with Aids. Doctors have not been able to detect the virus in his blood for more than 600 days, despite his having ceased all conventional Aids medication. Normally when a patient stops taking Aids drugs, the virus stampedes through the body within weeks, or days.

"I was very surprised," said the doctor, Gero Hütter.

The breakthrough appears to be that Dr Hütter, a soft-spoken hematologist who isn't an Aids specialist, deliberately replaced the patient's bone marrow cells with those from a donor who has a naturally occurring genetic mutation that renders his cells immune to almost all strains of HIV, the virus that causes Aids.

The reality, once again, is that adult stem cells have been treating patients for decades, and almost every day now there is another breakthrough. Let's see where AIDs activists now demand where the money is spent.

 

Not Exactly Rocket

Gonorrhea has picked up human DNA (and that’s just the beginning)

Millions of people pick up gonorrhea every year, but the bacteria that cause the disease (Neisseria gonorrheae) have picked up something in return. They carry a little bit of human DNA within their genomes. It seems that the microbe behind the clap is partly human.

The human side of N.gonorrheae is a ‘LINE-1 (L1) sequence’ – a short piece of DNA that can copy and paste itself into new locations in the human genome. It has no obvious function beyond making more copies of itself, but it is very good at that. There are around half a million L1 sequences in the human genome and together, they make up a fifth of our DNA. And one of these sequences managed to hop into N.gonorrheae.

Mark Anderson and Steven Seifert from Northwestern University discovered the out-of-place DNA because the full genomes of 14 strains of N.gonorrheae have been completed and are publicly available. Within this database, they found a small DNA fragment that’s almost a perfect match to a human L1. After sequencing many more strains of the bacterium, Anderson and Seifert found the rogue L1 in around one in nine of them, and in none of their close bacterial relatives.

N. gonorrheae can invade the cells of its host, but it can’t break into the nucleus where most of the DNA is stored. So how did it manage to smuggle in an L1? Anderson and Seifert think that the fateful event happened when an infected cell died and broke apart, exposing its own DNA to the bacterium living inside it. Even then, there’s no easy route into N. gonorrheae’s genome. L1 can hop around a genome but only if it has the right landing sites. A bacterial genome doesn’t provide any. How it got in is anyone’s guess, but Anderson and Seifert speculate that the bacterium could have glued a fragment that contained L1 into a broken chunk of its own genome.

This complicated chain of events could explain why transfers of DNA from humans to bacteria are so rare (indeed, this is the first example of such a swap). Other genetic exchanges are far more common. Bacteria can swap genes as readily as humans swap opinions and N.gonorrheae’s own genome is a melting-pot of genes from several species. This “horizontal gene transfer” is a great way of injecting rocket fuel into evolution. By trading genes, bacteria can gain new powers in a single bound, including both offensive and defensive abilities.

These swaps are so pervasive that a sixth of the genome of Escherichia coli – a common gut bacterium – is made up of borrowed genes. On rare occasions, bacteria also trade DNA with their hosts. In one famous case, a species called Wolbachia managed to insert its entire genome into that of the fruit fly it infects. It’s still there to this day, creating a genetic fusion of fly and microbe. Transfers in the opposite direction, from host to bacterium, are rarer.

It’s not clear if N.gonorrheae’s human loan is actually doing anything, but it certainly hasn’t changed very much since it first leapt into the bacterium. All of the L1s in all of the different strains are very similar, and still in the same place. Some evolutionary pressure could be stopping the fragment from changing, implying a possible use. But it’s not clear what that might be – after all, Anderson and Seifert couldn’t find any differences between the strains that have human L1 and those that don’t. Alternatively, the L1 sequence might have hopped across very recently and hasn’t had time to change. This could explain why only a ninth of the bacteria have it.

There is another obvious possibility: the human DNA could have come from a human. As geneticists handle their samples, bits of skin or hair can fall in, adding human DNA to an otherwise pristine sample. This is a big problem. In a different study, Mark Longo from the University of Connecticut found human sequences in over 450 other genomes, from bacteria to wheat to zebrafish.

Longo found these stray sequences by accident. He was originally searching the zebrafish genome for the remains of ancient viruses that embedded themselves in the genomes of our ancestors. These genetic fossils are found in a variety of different animals and they have similarities that reflect their shared history. But Longo found something different – short pieces of DNA called Alu elements that are unique to humans and other primates. These sequences weren’t just superficially similar. They exactly matched their human counterparts and they couldn’t possibly exist in a zebrafish. They were clearly contaminants.

Like L1s, Alu can move around the genome. But Longo ruled out the possibility that this mobile DNA had hopped from humans to other species, just as L1 had done into gonorrhea. He looked for, and found, stretches of DNA that flank Alu in the human genome and that can’t hop from place to place. These sequences hadn’t integrated themselves into the different genomes. They were like stray pieces from a different jigsaw puzzle that had ended up in the wrong box. At least a fifth of published genomes contain these vagrant pieces.

This could cause problems for scientists who study animal evolution. But the challenges and the stakes are even higher when human DNA contaminates human samples. This might seem odd, but consider that we’ll soon reach a point when individual people can have their genomes sequenced cheaply. Doctors could make medical decisions based on these results and they could do so wrongly if one person’s genes are contaminating another’s.

Meanwhile, Jonathan Eisen, who studies microbe genomes, says the contamination problem also vexes scientists who study species that can easily swap genes. “On the one hand, contamination has possibly contributed to mistaken claims of lateral gene transfer in the past,” he says. “However, filtering out all “weird” DNA will lead one to miss real LGT. In the end this is going to be very hard to prevent completely.”

But there are certainly steps that scientists can take. In Longo’s study, it’s telling that only the genomes of flu viruses, which are handled with extreme care, were free of contamination. Longo checked 172 flu genomes and couldn’t find a single trace of Alu.

Rachel O’Neill, who led Longo’s study, says that scientists can reduce the odds of contamination by handling samples with the greatest, “forensics-level” care. They can also run independent tests in different labs to check their conclusions. Eisen adds that the ideal way to weed out stray sequences is to truly finish genome sequences (many “full” genomes are actually patchy drafts) and to check and cross-check everything.

In the end, this is about alerting the scientific community to the scale of the problem and encouraging people to focus on quality control and validation, as well as technological speed. O’Neill says, “The most important thing is to be aware of the possibility.” Eisen agrees, citing the importance of “[educating] everyone about possible problems so that we do not see 1000 papers on weird lateral transfers over the next few years.”

So could the human DNA in gonorrhea just be one of these contaminants? It’s unlikely. Anderson and Seifert went back to the original strains whose genomes were sequenced and analysed them again. They got the same result, and the fact that the stray L1 appears at the same position in three different strains sealed the case. This seems to be one of those “weird lateral transfers” that’s actually genuine.