In a new  study, ¶¶ŇőÂĂĐĐÉä Boulder Physics Professor Meredith Betterton and her team explored a key motor protein in cells called kinesin-5, which helps organize the cell division machinery, and they discovered how a small chemical modification plays a big role in controlling its power.

In our cells, kinesins act like tiny “motors” to help move chromosomes, the packages of our genetic information, to opposite sides of the cell so it can split. Kinesins propel across a larger cellular structure—the mitotic spindle—made of long, thread-like filaments called microtubules. Kinesin-5 helps these filaments slide against each other, building a sturdy structure that pulls chromosomes into place.

“It’s a bit like having a crane that moves very heavy objects, despite being quite small itself,” Betterton explains.

For this process to work, kinesin-5 must apply just the right amount of force. Too little force and the spindle won’t form properly, but too much force can cause damage. It’s a balancing act, and this new study reveals how cells achieve this balance.

“Imagine a city full of moving parts—traffic lights, cars, pedestrians—all needing to be in sync to avoid chaos,” says Betterton. “Similarly, cells have systems in place to control these movements. This research helps us understand how those controls work to keep our cells, and us, healthy.”

A Deeper Dive into Phosphorylation

Previous research shows that one way that cells control kinesin-5’s activity is through a process called phosphorylation, where a small chemical group is added to the kinesin-5 protein. This modification acts like a “gear shift” for the protein, helping it adjust its activity levels.

“Think of it as putting the motor into the right gear, like a car,” adds Betterton.

For kinesin-5, scientists have found nine different areas on the “tail” of the kinesin-5 protein that can be phosphorylated. While one or two of the sites have been heavily studied, Betterton and her team decided to take a different approach, focusing on all nine sites at once, wondering if these modifications are the key to controlling the motor’s power.

“In some cases, proteins are affected by only a single phosphorylation site really dramatically, but that hadn't been seen in kinesin-5 motor proteins,” Betterton explains. “But in other cases, there can be multiple sites that somehow work together, which is called combinatorial phosphoregulation. So, we decided to kind of go for what would be the biggest change we could potentially make, to mutate all of them together. By altering all nine sites, we hoped to understand how much it would affect kinesin-5’s ability to move and organize the spindle.”

Modifications, Models, and Microscopy

The team created two types of mutations to investigate the role of phosphorylation in the kinesin-5 protein. The first mutation, known as the alanine or “9A” mutation, replaced nine phosphorylation sites with alanine, a non-phosphorylatable amino acid, effectively blocking phosphorylation at these locations.

The second mutation, called the aspartate or “9D” mutation, substituted the same sites with aspartate, an amino acid that partially mimics phosphorylation by carrying a similar negative charge. These two mutations allowed the team to compare the effects of blocking versus mimicking phosphorylation on kinesin-5's motor function and force generation during spindle assembly. By tracking these altered proteins with a green fluorescent protein (GFP) tag, the team watched their behavior in live cells under a microscope.

The team also built computer models and ran theoretical simulations to test whether these phosphorylation changes matched the effects they saw in the cells.

“The fact that we’re able to use not only fluorescent microscopy but also computer modeling and theoretical simulations is something that has developed over the course of my career,” Betterton adds. “When I started in the department more than 20 years ago, I only did simulations and theory work. I started this experimental part of my lab after I got tenure with support from the physics and MCDB [the Molecular, Cellular & Developmental Biology] departments. The fact we’ve been able to do work like this and have papers like this is exactly the new direction I wanted my lab to take.”

From their multidisciplinary work, the researchers saw that cells without properly phosphorylated kinesin-5 had a hard time forming the usual spindle structures. Normally, about 80% of cells show microtubule sliding in the spindle that can form protrusions, but when the phosphorylation was blocked, very few cells showed proper sliding.

“This dramatic drop really showed us how essential phosphorylation is for building a functional spindle,” said Betterton. “Without this fine-tuning, the whole system fails.”

The results confirmed that phosphorylation allows kinesin-5 to apply the right amount of force as the cell needs it, ensuring a stable, functioning spindle for cell division.

Looking at the Bigger Implications for Human Health

These findings have potential real-world implications for health. Problems in cell division are a hallmark of many diseases, including cancer, where cells divide uncontrollably.

“Understanding how motor proteins are controlled gives us insights into diseases where this regulation fails,” said Betterton.

This study may open up new strategies for cancer treatments that target proteins like kinesin-5 to interrupt abnormal cell division.

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Among the recipients is ¶¶ŇőÂĂĐĐÉä Boulder Physics Professor Meredith Betterton, who, alongside collaborator Jeffrey Moore from ¶¶ŇőÂĂĐĐÉä Anschutz, received funding for their project on tubulinopathies, genetic diseases that disrupt brain and nervous system development due to mutated tubulin proteins.

“You can think of tubulin as being like a brick that is stacked next to other bricks to build a road (the microtubule),” Betterton explained. “One of the puzzles about tublinopathies is that the mutation usually occurs in one tubulin gene out of many, so it affects only a minority (usually 25% or less) of the subunits. We aim to understand how a mutation in one small part of a tubulin gene can cause catastrophic defects at the cell and tissue level, ultimately impacting patients.”

Betterton's and Moore’s research proposes that tubulin mutations influence structural changes in neighboring tubulins, amplifying the mutation's effects and creating serious health issues for individuals.

“This award is very exciting for my lab and me because it will provide seed funding for a new direction for our work,” Betterton added. “It’s a fantastic opportunity to potentially help people affected by these diseases.”

Highlighting the collaborative nature of the project, Betterton emphasized the importance of interdisciplinary research: “We will work with the Moore lab at ¶¶ŇőÂĂĐĐÉä Anschutz to conduct a combined experimental and theoretical study. This award is meaningful because it supports a new idea predicted by our theoretical work, now finding support in experiments. As a theoretical physicist, being able to predict an important new effect is something we all hope to do in our work.”

The AB Nexus program continues cultivating a culture of collaboration and innovation at the University of Colorado. Its vision is to tackle the toughest challenges in human health through teamwork across diverse fields.

As Vice Chancellor Thomas Flaig noted in the award announcement: “Solving the toughest challenges in human health requires teamwork across a wide range of fields, and we’re very proud of how this program has helped to inspire so many new interdisciplinary research projects across our campuses.”

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