SynBioFromLeukipposInstitute

Article | Published: 04 March 2019

DNA-based communication in populations of synthetic protocells
Alex Joesaar, Shuo Yang, Bas Bögels, Ardjan van der Linden, Pascal Pieters, B. V. V. S. Pavan Kumar, Neil Dalchau, Andrew Phillips, Stephen Mann & Tom F. A. de Greef 
Nature Nanotechnology (2019) | Download Citation

Abstract

Developing molecular communication platforms based on orthogonal communication channels is a crucial step towards engineering artificial multicellular systems. Here, we present a general and scalable platform entitled ‘biomolecular implementation of protocellular communication’ (BIO-PC) to engineer distributed multichannel molecular communication between populations of non-lipid semipermeable microcapsules. Our method leverages the modularity and scalability of enzyme-free DNA strand-displacement circuits to develop protocellular consortia that can sense, process and respond to DNA-based messages. We engineer a rich variety of biochemical communication devices capable of cascaded amplification, bidirectional communication and distributed computational operations. Encapsulating DNA strand-displacement circuits further allows their use in concentrated serum where non-compartmentalized DNA circuits cannot operate. BIO-PC enables reliable execution of distributed DNA-based molecular programs in biologically relevant environments and opens new directions in DNA computing and minimal cell technology.

Structure-guided protein engineering of Cas12a yields variants that have increased activity and that can edit sites with previously inaccessible PAMs.

Type V CRISPR-Cas systems are distinguished by a single RNA-guided RuvC domain-containing effector, Cas12. Although effectors of subtypes V-A (Cas12a) and V-B (Cas12b) have been studied in detail, the distinct domain architectures and diverged RuvC sequences of uncharacterized Cas12 proteins suggest unexplored functional diversity. Here, we identify and characterize Cas12c, -g, -h, and -i. Cas12c, -h, and -i demonstrate RNA-guided double-stranded DNA (dsDNA) interference activity. Cas12i exhibits markedly different efficiencies of CRISPR RNA spacer complementary and noncomplementary strand cleavage resulting in predominant dsDNA nicking. Cas12g is an RNA-guided ribonuclease (RNase) with collateral RNase and single-strand DNase activities. Our study reveals the functional diversity emerging along different routes of type V CRISPR-Cas evolution and expands the CRISPR toolbox.

Genome-reduced bacteria often show impaired growth under laboratory conditions. Here the authors use adaptive laboratory evolution to optimise growth performance and show transcriptome and translatome-wide remodeling of the organism.

An epidemic of Lassa fever in Nigeria that has killed 69 people this year is on track to be the worst ever recorded anywhere. Now, in the hope of reducing deaths from Lassa in years to come, researchers in Nigeria are trying out a new diagnostic test based on the gene-editing tool CRISPR.

The test relies on CRISPR’s ability to hunt down genetic snippets ― in this case, RNA from the Lassa virus ― that it has been programmed to find. If the approach is successful, it could help to catch a wide range of viral infections early so that treatments can be more effective and health workers can curb the spread of infection.

Scientists in Honduras and California are testing CRISPR diagnostics for dengue viruses, Zika viruses and strains of human papillomavirus (HPV) associated with cancer. And a study to explore a CRISPR test for the Ebola virus is pending in the Democratic Republic of the Congo.

A robust, user-friendly test could reduce the death rates from Lassa fever, which can be as high as 60%, says Jessica Uwanibe, a molecular biologist developing a Lassa diagnostic at Redeemer’s University in Ede, Nigeria. “I’m working on something that could save a lot of lives.”

The emerging field of RNA nanotechnology harnesses the versatility of RNA molecules to generate nature-inspired systems with programmable structure and functionality. Such methodology ha

CRISPR–CasX represents a distinct RNA-guided platform that is functionally separate from Cas9 and Cas12a and is active for bacterial and human genome modification.

A bacterial enzyme that researchers often use to modify other organisms’ genomes can also be harnessed to tackle invaders.

The Cas9 enzyme naturally cuts DNA at targeted sites, guided by short ‘CRISPR’ sequences. David Savage at the University of California, Berkeley, and his colleagues added a trigger that allows proteases — enzymes that chew up proteins — to activate Cas9.

The team used a linker to join the two ends of the Cas9 protein together in such a way that the protein’s activity was locked. The resulting enzyme couldn’t function unless a protease cleaved the linker.

The researchers inserted the circularized Cas9 into cultured human cells and expressed a protease made by flaviviruses, a genus that includes the Zika and dengue viruses. The protease activated the Cas9 enzyme, which the researchers had programmed to cut a specific repetitive DNA sequence in the genome, causing the cells to die. The approach could be used to create other Cas9 variants, the authors say.

In addition to coding for protein sequences, RNA molecules encode a diverse set of gene-regulatory elements. RNA switches are one class of gene-regulatory elements that control protein expression in a manner that is dependent on the concentration of specific ligand molecules. These allosteric gene-regulatory elements have been shown as useful tools in engineering diverse cell types to display novel function. In particular, RNA switches have been used as genetically encoded biosensors and conditional controllers to direct cellular decisions based on the system’s changing environment. A significant focus in the field has been the generation of novel RNA switches that are tailored for different biological systems. We review approaches that have been used to generate RNA switches, which leverage the unique physical properties of RNA and the myriad ways in which RNA can modulate gene expression.

If humanity is going to push the boundaries of space exploration, we're going to need plants to come along for the ride. Not just spinach or potatoes, though--plants can do so much more than just feed us.

A science experiment aiming to demonstrate plants' capabilities in space has arrived at the International Space Station and is ready to begin relaying data. The experiment, according to University of Utah chemistry professor and project chief scientist Ming Hammond, will assess in real-time whether plants engineered to bio-manufacture specific proteins, in a process called synthetic biology, can do so in space. The experiment began on Dec. 18 and will run through Dec. 28.

See photos of the team and images of the installed experiment from the space station here.

"There's a lot of promise, potential and hope that we can use the tools developed in synthetic biology to solve problems," Hammond says, "not just that you would find in space, but where you have extreme limitation of resources."

Hammond's involvement in this experiment, called Hydra-1, began at the University of California, Berkeley, before her recent move to the U. She and Berkeley graduate student Rebekah Kitto joined with a "very multidisciplinary team", Hammond says, of scientists and engineers looking to perform synthetic biology experiments in space.

Synthetic biology is a field that engineers biological systems. In this case, the team is looking at plants as potential bio-factories. Every organism naturally produces countless proteins as part of its biological function, so why not engineer a plant to produce, say, a needed medication or a polymer that could be useful in future long-term space exploration missions?

"The benefit is that you can take seeds with you," Hammond says. "They're very lightweight. They grow and gain biomass using the CO2 that we breathe out. And if those plants can produce proteins on demand--we know that plants are able to produce anti-viral and anti-cancer antibodies on a large scale."

Synthetic biology is already established on Earth. But translating that same technology to spaceflight requires a different set of considerations. Hammond and her team encountered many of these constraints when adapting their experiment to operate within a small cube-shaped enclosure, and without tending from the space station crew. The enclosure is the same size (10 cm on a side) as the small low-cost CubeSats that are rising in popularity.

For an experiment on earth, researchers could test samples of plants as they grew to see if they were producing the desired protein. But that's not an option in space - in the early stages of planning, the team didn't even know if they'd get the experiment back at the end.

So the team decided to engineer plants to change color as they produced the target protein, and monitor the progress with a camera. It's an elegant and innovative solution, based on a previously published method, but adapted for the constraints of a cube in space.

"We had to take something that worked beautifully in the most carefully controlled and nurturing conditions," Hammond says, "and get it to work under very stringent, harsh and challenging conditions without human intervention in the plant cube." The plant cube was designed with the forward vision of preparing for plant growth studies on the Moon, and the Hydra-1 mission is a technology development step towards that goal.

The team of collaborators spans two continents, with other partners at NASA Ames Research Center, the International Space University, and the University of Strasbourg. But as one of the experiment's lead scientists, Hammond will be monitoring data from the experiment, and conducting the matching Earthbound control experiment, here at the University of Utah. The control experiment will grow the same plants, engineered to produce the same protein, but lag behind the experiment on the space station by one day, so that researchers can match both experiments' temperature conditions.

The entire experiment will take 10 days. "By day four or five we should know if the experiment worked," Hammond says. "There's so many variables that we can't know the answers to." In January, the cube will return to Earth and will be further analyzed in Strasbourg. "I plan for this experience to prepare us for doing more chemistry experiments in space!" Hammond says.

On the engineering side, the Hydra-1 experiment will help develop a framework, through a commercial company called ICE-Cubes, to commercialize similar cube-based space science experiments in the future.

It takes a lot of time and effort to put equipment in space, and Hammond appreciates the many hours of work that the team has put in over the past two years. "We are a small but dedicated group of volunteers," she says. "In the past two weeks, people worked nonstop to fix last-minute things that came up before launch. I'm just really proud of the effort that everyone has undertaken to get us to this point."

Hammond and her family traveled to the NASA Kennedy Space Center to watch the Dec. 5 launch of her experiment, which was nestled within a SpaceX Falcon 9 rocket on a resupply mission to the International Space Station. "I think most of us feel the allure and excitement that is stirred by the wonder of looking up and thinking about humans living and working at the space station," she says. "It was an amazing opportunity to share the launch with my 6-year-old son and other family members. Of all the things I've done in science this, for them, is the one that probably inspires the most interest and awe."

Read the International Space University's release here.