Gene editing, brain slicing, and glowing proteins, all in the pursuit of improved treatment for a disease that has befuddled the medical community for years. Methods like this used to be the stuff of science fiction. Now, it’s just another day in the life for Tyler Smith, an IGB Woese Undergraduate Research Scholar majoring in molecular and cellular biology.

Growing up in North Aurora, IL, Smith was exposed to a wide range of scientific fields from an early age.

“My dad is a computer scientist, and we had a lot of half destroyed computer parts laying around that I would spent time tinkering with,” recalls Smith. “I also had a huge interest in space, and often took trips to the planetarium with my mom”.

Smith’s interest in molecular biology emerged after taking AP Chemistry in high school, motivating him to explore the biomedical applications of chemistry in his degree.

These interests led Smith to the Gaj lab (BSD), which aims to use gene editing tools to develop treatments for a variety of neurological disorders including Alzheimer’s disease, the focus of Smith’s work. The most common gene editing method is CRISPR-Cas9, which makes use of a bacterial immune response machinery to open up DNA strands and insert an engineered piece of DNA into a specific location in the genome.

“There is a gene called GPR3 that, when upregulated, leads to the formation of more amyloid beta plaques, which are one of the key causes of Alzheimer's disease,” describes Smith. “Our goal was to knock down that gene with our CRISPR therapy and see its effect on Alzheimer’s symptoms and amyloid beta plaque buildup”.

To do this, Smith acquired mice that contained five mutations known to be associated with Alzheimer’s disease. He used CRISPR to treat the mice with either a therapeutic gene that inhibits GPR3 or a control that has no effect on GPR3. All mice were then tested for two behaviors - anxiety-like behavior and spatial learning. Smith was then tasked with developing a protocol to analyze these behaviors. “It came back to my upbringing and exposure to computer science through my dad’s work,” says Smith. “This helped me identify a software that could analyze the videos of mouse behavior and tailor it to fit our study”.

With the behavioral work completed last spring, Smith’s focus this past summer was on the molecular side. First, he performed immunohistochemistry (IHC) on the brains of mice from both treatment groups to verify that the CRISPR reached the target and assess the effects of the treatment on levels of GPR3. IHC begins with a process called sectioning, where the brain is sliced into many thin layers. Next, the brains are stained with green fluorescent protein, a protein that lights up under blue to ultraviolet wavelengths and is designed to bind to a protein of interest (GPR3 in this case). Thus, by imaging the brains under a microscope and counting the number of cells that fluoresce under these wavelengths, Smith was able to get an estimate of expression levels of GPR3 across treatments. Finally, he also performed an additional staining, this time targeting the amyloid beta plaques, to identify if the treatment reduced buildup of this common Alzheimer’s symptom.

For Smith, this staining and imaging process was both one of his most challenging and most rewarding aspects of this project. “It was a more meticulous process than I thought it would be. There are times where I wouldn’t see anything on an image, and senior lab members would tell me that there is actually something there,” says Smith. “However, it is really exciting when you do get a good image and can see your hypotheses turning into actual results.”

Analysis of these results are still ongoing, but Smith’s successful implementation of these methods has already established a proof of concept for this potential treatment. Next steps include genetic sequencing to further evaluate the editing rate of the CRISPR method, allowing Smith to assess how successful the CRISPR edits were and if any unintended edits were made at other locations in the genome.

Outside of his research, Smith enjoys playing video games like Spiderman and Final Fantasy, and playing with his dog, Chewy, and his cat, Swish. After graduating, Smith plans to take some time off of school and work a job in industry before applying to PhD programs.