Thousands of our cells successfully divide every day. They replicate their DNA and then separate so that each of the new cells has the same complement of chromosomes. If the process goes wrong and cells end up with too many or too few chromosomes, they may turn cancerous.
A complex network of molecular checks and balances ensures that chromosomes end up exactly where they should. Central to this network is an enormous multi-molecular machine called the kinetochore.
Fred Hutchinson Cancer Center molecular biologist Changkun Hu, PhD, was recently named a 2023 Jane Coffin Childs – HHMI Fellow by the Jane Coffin Childs Memorial Fund for Medical Research. Hu’s support from the JCC Fund will enable him to better understand how kinetochore components assemble to create a working kinetochore, and how this process is orchestrated.
“The kinetochore has hundreds of proteins [fitting together]. How such a big complex assembles together is not clear,” said Hu, a postdoctoral fellow in the lab of Basic Sciences Division Director Sue Biggins, PhD. “My project focuses on the temporal order [of the assembly] for each of its subcomplexes.”
Because cell division is so important to human health, the kinetochore’s assembly is carefully regulated and there’s still a lot to learn about that process. A better understanding of that regulation could help scientists see how it goes wrong in diseases like cancer, and guide development of new cancer treatments, Hu said. He has developed a new method that uses TIRF microscopy (short for total internal refection fluorescence microscopy) to chart the order in which kinetochore components come together.
“Changkun completely shifted fields and has already made major progress on his postdoctoral project that the JCC is funding,” said Biggins. “He is a highly motivated postdoc that we are fortunate to have in the lab.”
The JCC Fund was established in 1937 to support fundamental investigations of human disease and honors Jane Coffin Childs, who died of cancer in 1936. The JCC Fund’s Board of Scientific Advisors selects the best and brightest postdoctoral fellows to receive three years of salary support. As a JCC Fund Fellow, Hu is following in his mentor’s footsteps: Biggins also received a JCC Fund fellowship early in her scientific career. She is the current director of the JCC Fund’s BSA, a position which is not involved in the fellow selection process.
When a cell divides, it duplicates each chromosome, forming a closely connected pair that must separate so that each daughter cell gets a single copy of every chromosome. This process is so critical that there are stringent biological checkpoints in place, and most cells that can’t sort their chromosomes will die. But very rarely, cells improperly segregate their chromosomes and make it past these checkpoints.
This can cause big problems: If a daughter cell gets an extra chromosome, it may get extra copies of cancer-promoting genes that help it (and all of its daughter cells) grow unchecked. Conversely, if a daughter cell loses a chromosome, all of its progeny may lack genes that help prevent cancer. Aneuploidy, an abnormal number of chromosomes in a cell, is a hallmark of cancer.
The kinetochore is the complicated molecular machine that prevents aneuploidy and ensures that chromosomes get separated and sorted properly. It segregates chromosomes but also puts the breaks on cell division if there are problems.
Biggins’ team discovered how the kinetochore fulfills these dual roles.
“Sue discovered that you need the right tension for the kinetochore to work,” Hu said.
Tension is the clue that the chromosome-separation apparatus is set up correctly. Right before cell division, chromosome pairs are neatly lined up in the middle of the cell. In yeast, the simple organism Hu uses to study the kinetochore, each chromosome has its own kinetochore. (In more complex cells, like human cells, several kinetochores bind along the same chromosome, making the process tougher to study.) Long molecular ropes called microtubules reach from each end of the cell to attach to kinetochores like grappling hooks.
If the two kinetochores on a chromosome pair are grabbed by microtubules pulling from opposite sides, this creates a tug of rope-like tension. In this case, the opposing microtubules can haul chromosome duplicates apart and ensure they each end up in separate daughter cells. But if one microtubule doesn’t attach, or both microtubules attach to the same kinetochore (leaving the kinetochore on the other chromosome bare), there’s no tension and cell division falters.
To make sure this process works, the kinetochore must correctly assemble and bind to DNA. But how do cells get an assembled, DNA-bound kinetochore from hundreds of loose kinetochore components? There remains much to learn about this complicated process, Hu said.
“Is there a step-by-step assembly process [for each protein], or does it pre-assemble into larger complexes that are then recruited to the DNA?” Hu said.
His outline of kinetochore assembly will be a major step to understanding how the process is regulated and will help scientists see how it breaks down and allows aneuploidy to occur.
Hu is using TIRF, a type of microscopy that will allow him to track the individual molecules as they come together to form the kinetochore. He can tag different kinetochore components with a molecule that fluoresces, and then use TIRF to track that fluorescence over time and space as the components come together into a full kinetochore. There are about 13 subcomplexes that make up a complete kinetochore, and Hu’s goal is to tag at least one representative protein from each. After repeating the tagging-and-tracking process with a protein from each subcomplex, Hu will be able to build a model of kinetochore assembly.
His approach is not limited to the kinetochore, he said. After he maps kinetochore organization, Hu is planning on applying his TIRF strategy to other large molecular complexes, such as those that are involved in repairing damaged DNA.
JCC Fund Fellowship offers more than financial support, Hu said. He’s looking forward to the travel support the fellowship provides and connecting with the wider community of JCC Fund fellows, particularly at their annual meeting.
Sabrina Richards, a staff writer at Fred Hutchinson Cancer Center, has written about scientific research and the environment for The Scientist and OnEarth Magazine. She has a PhD in immunology from the University of Washington, an MA in journalism and an advanced certificate from the Science, Health and Environmental Reporting Program at New York University. Reach her at srichar2@fredhutch.org.
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