Â鶹ÒùÔº

Like its namesake, Janelia's GENIE Project Team makes wishes come true. Luckily for biologists, this genie doesn't grant just three requests.

For more than a decade, GENIE has collaborated with Janelia's tool developers and tool users to take innovative biological sensors from novel prototypes to —a feat likely impossible for a single lab on its own.

Now, the team is unveiling their latest collaborations. Working with the tool developers and Janelia's support teams, GENIE created next-generation sensors for detecting and —key neurotransmitters that enable neurons to communicate with each other. Through work with Janelia's Project Technical Resources support team, they've also aimed at controlling and studying specific neural circuits in fruit flies.

"That's the essence of the project teams: to do higher throughput, larger person-hour projects that no Janelia lab is set up to do or wants to do," says GENIE Screening Manager Jeremy Hasseman. "That's where those collaborations are really key."

With its dedicated team of scientists, close collaborations with Janelia's labs, and access to Janelia's support teams, GENIE has developed a winning formula for accelerating the development of best-in-class sensors for measuring neuronal activity.

The team collaborates with tool developers to screen and test hundreds of variants, creating state-of-the-art sensors that allow researchers worldwide to track neuronal activity in living animals at spatial and temporal scales not previously possible. These sensors not only enable new discoveries about how the brain works but also free up Janelia's tool developers to create new inventions.

"These projects are at the scale of a team, not the scale of a person," says GENIE Project Team Director Glenn Turner. "Tool developers are incredibly skilled at making different sensors, and if they can exploit that and build a bunch of different things that are good, then, maybe, we can make them a little bit better."

A team approach

Janelia's Project Teams bring together groups of scientists for years at a time to take on large-scale efforts that have the potential to transform entire scientific fields but can't be accomplished in a traditional academic environment or by a single lab. The resulting tools, data, and knowledge are shared widely with the scientific community.

GENIE was created to speed up the development of genetically engineered protein sensors for neurophysiology—tools that enable neuroscientists to track signals from large groups of neurons. These sensors use specially designed proteins to detect specific biological signals associated with neuronal activity and convert them into a measurable output, like fluorescence.

Engineered protein sensors can take years to create, and the first iteration is often not optimized for use in biological research. Improving on the initial prototype can be tedious—perhaps impossible—for a single lab to accomplish on its own.

"A lot of sensors kind of die in that space: It looks good as a biochemical tool but then it doesn't really translate and it's really hard to make it work," Turner says.

To ensure its products are optimized, GENIE works closely with Janelia's tool developers to figure out how to improve the sensors. With help from Janelia's support teams, they create hundreds of variants and screen them in cultured neurons, which are a good proxy for how well the sensor will work in vivo.

The team identifies which mutations are beneficial for whatever attribute they are looking to improve—like speed or brightness—and then combine those to see which groupings work best. From there, the team tweaks the variants until they work well for their intended purpose.

To test the new sensors in vivo, GENIE works with Janelia's biology labs, giving the researchers early access to the tools while also ensuring that the sensors work well before releasing them to the larger scientific community.

"We're pushing the science forward here as well as outside Janelia," Hasseman says.

Improving neurotransmitter detection

Two new sensors for detecting neurotransmitters exemplify GENIE's approach. The development of iGABASnFR2, the latest sensor for detecting the neurotransmitter GABA, was a typical collaboration between GENIE and a Janelia lab—in this case the lab of Loren Looger, a former Janelia Group Leader. Looger and Janelia Senior Scientist Jonathan Marvin developed the prototype, iGABASnFR, which was the first sensor for detecting the inhibitory neurotransmitter GABA.

The research is in the journal eLife.

"You see neural activity and generally that's excitatory, but somewhere underneath that excitation is some kind of inhibition that's sculpting those excitatory signals. This allows you to actually see that," Turner says.

Researchers inside and outside of Janelia had used the prototype GABA sensor with mixed results. GENIE was able to work with the developers to create and screen nearly 4,000 variants to make the sensor more sensitive and faster, enabling it to be more useful to biologists. They also created a version that starts bright but dims when it binds to GABA, allowing researchers to use it in conjunction with other indicators.

"This was a great example of taking a prototype that people had used outside the building and had tried with very mixed results," Hasseman says. "We were able to find things that improved the sensitivity quite a bit and extend what people were able to use it for."

was developed to detect glutamate—one of the most important and abundant excitatory neurotransmitters in the brain. The sensor was developed by former Janelia researchers Kasper Podgorski and Abhi Aggarwal.

When it was released in 2022, the sensor's third iteration was the fastest and brightest indicator available for measuring glutamate as it enters a neuron's dendrites, but the team knew it had the potential to be improved even further. Some biologists also wanted a version that fluoresced for longer, so they could use it to study larger areas.

GENIE's work allowed the team to collaborate with the tool developers, now at the Allen Institute, to target many more sites on the molecule. By screening more than 9,000 variants, they developed , a more sensitive, brighter, and faster version of the third-generation sensor. Details have been published on the pre-print server bioRxiv.

They also created iGluSnFR4s, which is just as sensitive and bright but turns off slower, allowing the recording of signals across large numbers of synapses simultaneously.

"It was a very natural endpoint because it was quite a bit better and opened up new experiments for a lot of people," Turner says. "At this point, the sensor goes out into the wild and people give feedback on it and say if they want it better and what they would want."